BIO181
LAB MANUAL
ESTRELLA MOUNTAIN COMMUNITY COLLEGE
Spring 2011
BIO181 Lab Manual
Created by Smith/Steele
EMCC Spring 2011
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Table of Contents
Table of Contents ................................................................................................................ 3 Safety Measures for Students In General Biology Classes ................................................. 5 Metric System Conversions ................................................................................................. 9 How big is a . . .? ....................................................................................................................................... 18 Lab Equipment ........................................................................................................................................... 22 Measuring Liquids ............................................................................................................. 25 The Scientific Method ........................................................................................................ 28 Week 1 Lab Report Instructions ........................................................................................ 33 GOOD GRAPH OR BAD GRAPH?? ................................................................................. 38 Brain Warm Up: Molecular Modeling ................................................................................. 44 Molecular Modeling ........................................................................................................... 45 Functional Groups of Organic Molecules .......................................................................... 52 Solvent properties of oil, water, and alcohol ...................................................................... 54 Brain Warm Up: Organic Macromolecules ........................................................................ 56 Testing for Organic Compounds ....................................................................................... 57 Denaturation and Coagulation of Proteins......................................................................... 68 Measuring pH .................................................................................................................... 70 LAB SUMMARY NOTES ................................................................................................... 71 Determining the Solute Concentration of Potato Cells ...................................................... 74 Osmosis Lab Report and Questions.................................................................................. 77 Cell Membranes and Osmosis Lab Stations ..................................................................... 81 Brain Warm Up: Cells ....................................................................................................... 83 Microscope and Cells Lab ................................................................................................. 84 Brain Warm Up: Calorimetry & thermodynamics .............................................................. 97 Food Calorimetry: Measuring the energy in Food ............................................................ 98 Brain Warm Up: Enzymes .............................................................................................. 109 Enzyme Lab Problem: Chemical Production ................................................................... 110 Enzyme Experiment Lab Report ..................................................................................... 126 How to use a Spec-20 ..................................................................................................... 128 Photosynthesis and Cellular Respiration Review Lab ..................................................... 131 Brain Warm Up: DNA Structure ...................................................................................... 136 DNA and RNA Structure.................................................................................................. 137 Brain Warm Up: DNA replication .................................................................................... 144 DNA Replication - PUZZLE ............................................................................................. 145 DNA replication - COMPUTER ........................................................................................ 148 The Cell Cycle .......................................................................................................................................... 150 Brain Warm Up: Gene Expression ................................................................................. 152 Gene Expression – Team 1............................................................................................. 154 Gene Expression – Team 2............................................................................................. 156 Recovering the Romanovs .............................................................................................. 160 Brain Warm Up: Meiosis................................................................................................. 162 Meiosis – Reebops ................................................................................................................................. 163 Lab Final Study Guide ..................................................................................................... 168 Created by Smith/Steele
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Safety Measures for Students In General Biology Classes
CAUTIONARY STATEMENT
In several of the laboratory exercises in Biology, you will use materials that are toxic,
flammable, reactive, or corrosive. Since exposure to any of these materials could pose a
health risk, it is necessary that you understand and carry out basic lab safety procedures.
If you are allergic to any of the chemicals used in specific labs, it is your responsibility to make
note of it and consult with your instructor.
If you are pregnant or have a compromised immune system (that is, you are on a
chemotherapy regimen, you are taking corticosteroids, you have leukemia, AIDS, active
tuberculosis, etc), the consequences of an exposure to certain chemicals are slightly greater
than for individuals with a normal immune system. If you are pregnant, or if you have reason
to suspect that your immune system is not fully functional, you should consult with your
instructor and your physician for advice about taking Biology at this time.
LABORATORY SAFETY MEASURES
Please follow the safety and preventative measures outlined below any time you are in the
lab. If you do not follow these guidelines, your instructor may ask you to leave the lab.
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Wear proper eye protection (goggles or safety glasses that are ANSI approved) any time
glassware or chemicals are used. Glasses without side shields are NOT sufficient. The
bookstore has safety glasses which meet current safety standards. You should NOT
wear contact lenses in lab.
Wear gloves and safety goggles for any labs that involve animal dissection or hazardous
chemicals.
Be able to read and interpret the hazard labels on chemicals provided in lab. Be sure to
read the hazard label before using each chemical. If you wish more information about
chemicals used in lab, consult your instructor.
Follow explicitly directions given by the instructor for the use and disposal of chemicals
used in the lab.
NO FOOD OR DRINKS ARE PERMITTED IN THE LABORATORY. No smoking is
permitted in the laboratory
Tie back long hair and do not wear loose jewelry. You should not wear shorts or
expensive clothes to lab. Open shoes, such as sandals should not be worn in lab.
Keep open flames away from flammable materials and chemicals and YOU! Familiarize
yourself with the location of the fire extinguisher and fire blanket in the room.
Use equipment carefully. Ask your instructor for assistance if you have questions or
problems.
Report to the instructor any injury to yourself or another student, no matter how minor.
Report to your instructor any spills so they can be cleaned up immediately.
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This means it is easier
to use!
What’s up with this?
by Smith/Steele
Why don’tCreated
we use the
metric system?
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MARS DISASTER: One Small Mis-step For NASA Can Lead To One Giant Step
Forward For America (October 1999)
The recent loss of the $125 million spent by NASA in developing the Mars Climate Orbiter
might be the catalyst that wakes up Congress to realize that the U.S. cannot continue using two
measurement systems. This realization could be the beginning of the completion of U.S.
metrication.
U. S. Metric Association (USMA) president, Lorelle Young, blames the problem on a much
larger issue, the failure of the U.S. to fully adopt the metric system. "This failure has much
greater implications for the American public," she said. "There is no question that we will
become a fully metric nation, but by foot-dragging the process we are depriving ourselves of
the many benefits the American people could be enjoying now. By supplying the world with
exports of metric goods we could greatly strengthen our economy and provide more jobs for
American workers, and our math and science education-reform efforts would greatly accelerate
if the metric system were taught consistently in our schools."
NASA has been using the metric system for years according to JPL Administrator, Tom Gavin.
NASA mistakenly thought Lockheed, its contractor, was using metric data, but it turned out they
were using inch-pound measures. Noel Hinners, vice president of flight systems for Lockheed
Martin Astronautics in Denver admitted "We should have had them [the measurements] in
metric units."
NASA reported that the minimum survivable altitude would have been between 85 and 100
kilometers (53 to 62 miles). But the mismatch of measurement units allowed the spacecraft to
come within 57 kilometers (35 miles) of Mars where the temperature was too high. NASA's plan
was to approach the planet at 140 kilometers (87 miles).
2009 December 31
All products sold in Europe (with limited
exceptions) will be required to have only
SI-metric units on their labels. Dual
labeling will not be permitted.
Implementation of the labeling directive,
previously 1999 December 31, was
extended by the EU Commission for 10
years, giving more time for companies to
comply and for U.S. regulations to allow
metric-only labeling on consumer products.
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Unit Conversions and the Metric System
BIO 181/ CHM130
Converting between units of measurement is an important skill to learn. The most
common systems of measurements are the English system and the metric system.
While many people in the United States still use the English system of measurements,
most scientists and many applications in healthcare use the metric system.
If you grew up in the United States, you are probably most familiar with the English
system of measurements.
1) Give an example of an English unit for:
a) length?
b) mass or weight?
c) volume?
In this lab you will learn how to convert between different units in the metric system and
convert between the English and metric system.
We are going to learn how to use unit equivalencies to convert measurements.
As long as you know the equivalency you can convert between any units, English Æ
English, English ÅÆ metric or metric Æ metric.
UNIT EQUIVALENCIES
You are familiar performing unit conversions when you convert something like “36 eggs”
into “dozens of eggs”. You know 36 eggs are equal to 3 dozen eggs, but that’s because
you already know the unit equivalency 12 eggs = 1 dozen eggs.
Here it is mathematically. DON”T FREAK OUT!! This is not hard math. Take a deep
breath and read this through.
Problem: Convert 36 eggs to dozen eggs
⎛ 1dozen ⎞
⎟⎟ = 3dozen
36eggs⎜⎜
⎝ 12eggs ⎠
The math you end up doing is 36 x 1 which is the same as 36 divided by 12 which
equals 3.
12
Notice that we write our unit equivalency as a fraction with the unit we want (dozens in
this case) in the answer on top and the unit we want to cancel or get rid of (individual
eggs) on the bottom.
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Here are some more UNIT EQUIVALENCIES
for converting between English units:
Any of these can be writing as fractions, for example:
1 ft
12inches
or
1gallon or
4 quarts
12inches
1 ft
1 ft = 12 in
1 cup = 8 fluid oz
1 gallon = 4 qts
1 yard = 36 inches
1T = 3teaspoons
1 lb = 16oz
Note that the fraction can be written either way up (look at the feet and inches
examples). Which one you use will depend on what the problem you are trying to solve
is asking for.
EXAMPLE PRACTICE PROBLEM:
Use the above unit equivalencies to practice converting English Æ English
measurements:
a) Your waist measures 28 inches. How many yards is this?
There are 3 steps to solving these problems
1. Write out the problem as shown below, with the quantity you are given (in this
example 28inches) on the left, then leave a gap to write the unit equivalency and
then “equals ? “ and the unit you are trying to convert into (in this example yards)
on the right:
28 inches
= ? yards
STEP ONE
2. Next, find the unit equivalency that relates inches to yards (look at the sticky note
at the top of this page). 1 yard = 36 inches
3. Write the unit equivalency as a fraction in the gap you left. Put the unit you
WANT in the answer (yards in this example) on the top of the fraction and the
unit you are starting with (inches in this example) on the bottom
28 inches x
1 yard
36 inches
= ? yards
STEP TWO
4. Now just do the math: 28 multiplied by 1 and divided by 36.
The inches units will cancel and that will leave you with the answer in yards
28 inches x
1 yard
36 inches
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= 0.77 yards
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NOW you try these two problems on your own. Use the space give to write out the steps
for each problem
b) You need to take 2 teaspoons of medicine. Convert this to tablespoons
c) You ordered a 22 ounce steak. How many pounds is it?
Now, you can use this exact same method to convert
from English to Metric units, or vice versa, provided
you know the unit equivalencies.
For example:
Problem: Convert 346.5lb to kg
1 in = 2.54 cm
1 oz = 28.4 g
1 gallon = 3.79 liter
1 kg = 2.2 lb
1 fl oz = 30 ml
1 pint = 500 ml
⎛ 1kg ⎞
346.5lb⎜
⎟ = 157.5kg
⎝ 2.2lb ⎠
Using the problems at the left,
show how units are cancelled.
Problem: Convert 23.6 cm to ft.
⎛ 1inch ⎞⎛ 1 ft ⎞
23.6cm⎜
⎟⎜
⎟ = 0.77 ft
⎝ 2.54cm ⎠⎝ 12inch ⎠
!!!! What is happening in that second example??
To go from cm to feet we are first converting from cm to inches (because we have an
unit equivalency for that: 1in =2.54cm) and then from inches to feet (because we have a
unit equivalency for that: 1ft = 12 inches). We have to do this in two steps because we
don’t have a unit equivalency for cm and feet directly.
HOW do you know which way up to put the equivalency fractions? You always put the
unit you are converting into on the top, so in this example the first step turns cm into
inches and the second step converts inches into feet.
Before you go on, check your understanding about the example problems above:
1. T or F. For the first problem “what you know” is the number of pounds while “what
you want to know” (or have to figure out) is the number of kilograms.
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2. T or F. For the second problem “what you know” is the number of inches while “what
you want to know” (or have to figure out) is the number of feet.
3. T or F. For all problems the unit you want has to end up being on the top of the last
unit equivalency or fraction that you use.
Practice Problems – use the unit equivalencies given on pages 10 and 11 to solve
these problems. Some problems will require you use more than one equivalency
fraction.
1) Convert your 28 inch waist to centimeters.
2) A person weighs 150 pounds. What is his weight in kilograms?
3) An average male has a lung capacity of 6 liters. Convert this to quarts.
4) A patient receives 1500mL of blood. How many pints did they receive?
5) A bottle of alcohol contains 375 ml. Convert this to fluid ounces.
6) A person’s height is recorded as 152.4 cm. How tall are they in feet?
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Now that you understand how to convert within different English system units and
between some English and metric units, we need to look at metric units in more detail.
Scientists and most countries use the Metric System of measurement. An advantage of
this system is that it is based on powers of 10; therefore it is easy to convert between
different metric units.
The practice problems you did earlier using unit equivalencies were actually done by a
technique called Dimensional Analysis. This technique is also used to do metric to
metric conversions.
The scale below shows you the common metric prefixes (centi, milli, micro etc…) and
their unit equivalencies to the base unit of a meter, liter or gram.
10-9
-3
10-6
-2
10
10
meter m
liter l or L
gram g
Base metric units
for length, volume
and mass
103
centi c
nano n
micro μ
milli m
kilo k
This scale is just a quick way of showing a bunch of
equivalencies for milli, micro, nano, centi and kilo all together.
For example the unit equivalency between meters and millimeters is:
1mm
10-3m
or
10-3m
1mm
PRACTICE PROBLEM: How many milliliters (mL) are there in 2 kiloliters (kL)?
Follow the steps just as before:
1. Rewrite the question:
2.0kL
= ? mL
2. The unit equivalencies on the scale allow us to convert to and from the base units of
meters, liters or grams, so in this case we are going to have to convert kiloliters to
liters, then liters to milliliters. We will need two equivalency fractions to do this.
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2.0 kL
x
103 L
1kL
EMCC Spring 2011
x
1 mL = ?mL
10-3L
Converts
kL to L
Converts
L to mL
3. Do the math… (you can use a calculator) …
The answer is 2,000,000 mL. (2.0 x 106mL)
2kL = 2 000 000.0 mL
Try one more: convert 450 ng to cg
STEP 1 Rewrite the question:
450ng
= ? cg
STEP 2 Find the appropriate unit equivalencies. You will need ng to g and then
g to cg.
STEP 3 place the fractions in the problem making sure you always put the unit you want
to convert into on the top
450ng x
10-9g
1ng
x
1cg
10-2 g
= ?cg
The number is finally written as: 0.000045cg – don’t forget the units! (4.5 x 10-5cg)
Finally:
How many Meters are in 4.1cm? Let’s say our partner in class has it set up like this:
4.1cm x 102cm
1m
4. Check your understanding: What is wrong?
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5. You tell your neighbor that a good rule to follow for using unit equivalencies is:
a) (number you want) x (unit you have)
(unit you want)
b) (number you have) x (unit you have)
(unit you want)
c) (number you want) x (unit you want)
(unit you have)
d) (number you have) x (unit you want)
(unit you have)
e) don’t ever use them and drop chemistry or biology so you don’t have to
Do the following 5 questions with your partner and DON’T FORGET UNITS when
you write your answers!!!
1. If we use meters to measure length, what do liters measure?
And what do grams measure?
2. How many nl are in one mL?
3. 4.5 g = ________mg?
4. Convert one cg into g?
5. How many mg are in 5 μg?
The rest are practice. The answers are opposite
6. How many cm are in 1km?
7. How many kL are in 50L
8. Measure the width of your hand in cm
9. Convert this to mm
10. How many grams of crackers are in the cracker box?
11. Convert this to µg and kg
12. A cookie recipe calls for 250mL of milk. How many liters is this?
13. Convert this to cL
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More Conversions
10-9
NAME:
10-6
BIO 181/ CHM130
10-3
10-2
meter m
liter L
gram g
103
centi c
nano n
micro μ
milli m
kilo k
1 ft = 12 in
1 cup = 8 fluid oz
1 gallon = 4 qts
1 yard = 36 inches
1T = 3teaspoons
1 lb = 16oz
Convert 345 cm into m
Convert 12.5 cm into µm
Convert 32.3 cm to inches.
1 in = 2.54 cm
1 oz = 28.4 g
1 gallon = 3.79 liter
1 kg = 2.2 lb
1 fl oz = 30 ml
1 pint = 500 ml
Convert 465 liters to gallons.
A person weighs 150 pounds. How much does he weigh in grams?
A bacterium is 0.0000087 inches wide. Convert this into centimeters.
How many 325 mg tablets can be produced from 2.50 kg of ibuprofen?
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One baked potato provides an average of 31.0 mg of vitamin C. If 5.0 lb. of potatoes
has 15 potatoes, how many milligrams of vitamin C are provided per pound of potatoes?
In body fluids such as blood or saliva, a pathogen can survive over 15 days at room
temperature. How many hours can the pathogen survive in liquid at room temperature?
You are working with a drug and only have 0.38 kg of the drug available. You are doing
an experiment on mice and each one needs to receive 5 µg (5 mcg). You need to
determine how many µg (mcg) of the drug you have so you know how many mice you
can use.
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How big is a . . .?
EMCC Spring 2011
NAME:
Turn on the computer and log in to http://www.cellsalive.com/howbig.htm
Click on ‘Start the Animation’. You will notice that the scale for the image is in
millimeters. Complete this worksheet, writing your answers on the lines provided.
Space is provided so you can show your working if necessary.
Q1: How many millimeters are in a meter?
Q2: Estimate the size of the head of the pin.
Click on the 10 on the magnification bar. Check the size of the head of the pin. Change
it if you want. Take your pointer over the words “human hair” and “dust mite” using the
list on the right hand side. You can see that those are the two things you can see when
you magnify 10 times.
Now click on the 100 times.
Q3: Estimate the width of a dust mite in microns (micrometer = micron). (Yes,
you are going to have to think about how to do this! – Use the scale bar on the
image just as you would use the scale on a map to measure distances)
Q4: How many microns are in one meter?
Click on 1000 times. Now you see a group of objects.
Q5: Estimate the diameter of the lymphocyte which is a type of white blood cell.
Click on the 10,000 times. The green cells and yellow cells are types of bacteria.
Q6: There are 4 Staphylococcus cells in this picture. Estimate the diameter of
one cell.
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Click on 100,000 times.
Q7: Estimate the length of the Ebola virus in nanometers. When Ebola virus
infects a human body, it liquefies most of the internal organs and causes the
patient to bleed from any opening. You usually die within one to two weeks. It
kills 90% of the people it infects.
Q8: How many nanometers are in one meter?
Click on 1,000,000 times.
Q9: Rhinovirus is the type of virus that causes the common cold. There are
approximately 114 different types of this virus. Estimate its diameter. Use the
appropriate metric unit for your answer.
Q10: Now, use one of the rulers to measure the width of one of your index
fingernails in mm.
Many organisms are carried under fingernails – let’s see how many cold viruses
(rhinoviruses) would fit across your fingernail. Use the answers from Q9 and
Q10 and do the math – How many will fit? Yep, you are going to have to do some
conversions!
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Becoming more familiar with metric measurements
BIO 181/ CHM130
LENGTH
It is useful to be able to estimate lengths in the metric system. Examine a ruler, and get
an idea of the length of a millimeter, centimeter, and meter.
1a) An adult's height may be about
1) 1.7 km
2) 1.7 mm
3) 1.7 cm
4) 1.7 m
b) The longer edge of a credit card is about
1) 8.5 µm
2) 8.5 cm
3) 1.7 cm
4) 1.7 m
c) The thickness of the wire in a paper clip is about
1) 1 mm
2) 10 mm
3) 1 cm
4) 10 cm
d) The width of this sheet of paper is about
1) 22 m
2) 22 µm
3) 22 cm
4) 22 ml
VOLUME
Volume is a measure of how much space an object takes up. Examine the containers
of blue liquid on the back bench to get an idea of what a Liter, milliliter and microliter
look like.
2) Think about the relative sizes of liters and milliliters and circle the appropriate volume.
a) One aluminum can of soda is about
1) 3.5 ml
2) 35 ml
3) 350 ml
4) 3.5 l
b) A small glass of orange juice is approximately
1) 2.0 µL
2) 2.0 ml
3) 200 ml
4) 20 l
c) A gallon of milk is about equal to
1) 380 ml
2) 3.8 ml
3) 0.38 ml
4) 3.8 µL
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BIO 181/ CHM130
How to quickly check your work – Is your answer absurd?
CONCEPT:
Many small things fit inside one big thing. Small things are fractions of
big things.
Let’s say this mouse is 90 mm in length and the elephant is 9m in length.
1. What is the length of the mouse in meters? First JUST LOOK at the four
possible answers:
a) 90,000m
b) 0.9m
c) 90m
d) 0.09m
This question is asking you to convert 90mm into meters. Without even doing the math
two options are absurd. Which really do not make sense AND WHY?
Try these questions without a calculator or even doing any working with a pencil. In
each case just read the question and circle the answers that can be immediately
eliminated because they are obviously absurd. There are at least two absurd answers
in each list.
2. How many nL are in 5mL?
a) 50 000nL
b) 0.5nL
c) 0.000005nL
d) 5 000 000nL
3. Convert 750mm to km
a) 0.0075km
b) 750km
c) 0.00075km
d) 75 000km
4. A tablet that contains 0.2mg of drug is equivalent to what dose in mcg?
a) 200mcg
b) 0.0002mcg
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c) 0.0000002mcg
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d) 20mcg
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NAME:
Lab Equipment
You are a lab manager and you must identify lab equipment that is labeled in a store
room. In addition, you must provide to your supervisor the function of each piece. While
you are familiar with most of the equipment, you still use a reference sheet to help you.
Unfortunately, there is one item that you do not have on your sheet, so do your best.
Fill in the following chart:
Item
number
Name
Function
001
002
003
004
005
006
007
008
009
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Lab Equipment Reference Sheet
flask
beaker
Bunsen
burner
pipet
Test tube tongs
hot plate
graduated cylinder
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ring stand
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NAME:
Measuring Liquids
You will be using several pieces of equipment designed for measuring liquids. They
include pipets, flasks, beakers and graduated cylinders.
1) Use the beaker at your table and measure out 100mL of water from the tap.
2) Now, pour it into the graduated cylinder – is it exactly 100mL?
3) The graduated cylinder is more accurate than the beaker – WHY?
4) a. Measure 2mL of water using a 1mL pipet and put it into the small cylinder – is it
exactly 2mL?
b. Measure 2mL of water using a 5mL pipet and put it into the small cylinder (make
sure the cylinder is as dry as possible) – is it exactly 2mL?
c. Was there any difference that you could tell using the two different pipets?
Circle the statements that are true:
a) Beakers are used to measure the mass of a liquid.
b) There is a greater error when using a beaker to measure a liquid compared to
a graduated cylinder.
c) Flasks, pipets and cylinders are used to measure the volume of the liquid.
d) Unmarked pipets will provide accurate measurements.
e) A pipet will be better for measuring small volumes of liquid since the diameter
is smaller.
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Brain Warm Up: Salt Water Boiling
INTRODUCTION
A. Answer the following True/False questions – we will go over them in class.
a.
As a liquid heats up, the molecules break apart into atoms
b.
When water boils and evaporates you are actually seeing Hydrogen and Oxygen
gas escaping as the water breaks down
c.
Boiling water is undergoing a chemical reaction
d.
You can tell a chemical reaction is happening because of all the bubbling action
e.
Adding salt to the water is creating new molecules of saltwater
f.
Once water boils the liquid will stay at the same temperature regardless of how
long it boils
B.
What is the molecular formula for water?
Draw the structural formula for water
Draw what a water molecule looks like at 100°C
Describe what a water molecule is doing at:
•
0°C
•
37°C
•
105°C
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The Scientific Method
You are going to investigate how the temperature at which a solution boils is affected by
the amount of salt in the solution. That is, how hot does the solution need to get before
it boils. You may be familiar with this idea – have you ever been taught to put salt into
the water before you cook macaroni? What role might the salt be playing? Today, you
get to investigate whether or not salt might alter the boiling point temperature of water
by determining the boiling point temperature of different concentrations of NaCl
solutions.
Flip to Page 24 and read the experimental procedure, then continue reading below
Look at the following VARIABLES - These are things you could potentially change
and/or measure during your experiment today:
a.
b.
c.
d.
the volume of the solution you’re heating
the amount of time each solution takes to boil
the amount of salt you put into the solution
the temperature at which the solution boils
Some variables are set or determined by the scientist and some are measured (the
outcome is unknown).
Answer the following questions concerning these variables.
1. One of them will be kept constant, that is, it will be the same for each
solution you investigate. Which one is it?
2. One of them will be different between solutions AND it is the one that you
are going to investigate and measure. Which one is it?
3. In order to investigate the variable in #2, you will need a piece of lab
equipment to measure it – what will you need?
4. For this specific experiment, will you be measuring variable b?
5. There is one variable left. This variable will be set or determined for you.
Which one is it?
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Generating a Hypothesis
Remember, a scientist starts by observing a phenomenon and wondering about it. The
phenomenon you are going to think about and try to explain is the effect of the salt NaCl
(there are many salts, NaCl is just one) on the boiling point temperature of water. You
probably have some kind of impression or ideas in your head from family or your own
experimentation when you are cooking pasta. So, first, think about the question: How
does adding NaCl to water affect its boiling point? Next, you will generate a
hypothesis: an explanation of what you think will happen. What you think will
happen is called a prediction. These two are easily put together as follows:
I predict that as the molarity (concentration) of the NaCl solution increases, the boiling
point will:
, because
After the word because is your explanation or your hypothesis. It should be based on
any knowledge you have of ions, molecules, bonds, and solutions. Try to think at the
molecular level. Remember, you are dealing with a solution which means one
substance (NaCl) is surrounded or dissolved in another substance (H2O). A picture
showing this is on p. 51 of your textbook and is helpful. If you have no knowledge of
molecules, atoms, etc. then do the best you can using any observations you have made
previously. You still must talk about the water and salt as two substances in the same
container and what may be happening between them as the temperature of the solution
rises. It is also helpful to think about what is actually happening at the boiling point That is: what is different between a solution that is actually boiling compared to one that
is just hot.
These animations will help you also:
Dissolving salt:
http://www.mhhe.com/physsci/chemistry/essentialchemistry/flash/molvie1.swf
Boiling water – scroll down to steam:
http://www.visionlearning.com/library/module_viewer.php?mid=120&l=&c3=
BEFORE YOU DO THE EXPERIMENT, YOU MUST HAVE
YOUR HYPOTHESIS APPROVED BY YOUR INSTRUCTOR
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Experimental Procedure
1. Obtain four 125 or 220mL flasks and label them A, B, C, and D. You are going to
experiment with the following concentrations of NaCl in water: 0M, 1M, 3M, and 5M.
FYI: M or molarity is referring to concentration. It is the way scientists describe the
concentration of solutions. It refers to the amount (or number of moles) of a substance
dissolved in a liter of a liquid (usually water). The higher the molarity, the more
concentrated the solution. So a 5M NaCl solution is more concentrated (or ‘salty’) than a
3M NaCl solution.
Before you measure your solutions, you need to know how scientists use graduated
cylinders to do this as accurately as possible. Look at the following diagram:
25
20
When a liquid is in a graduated cylinder the liquid (usually a solution made with water)
clings to the side of the cylinder and is ‘pulled up’ at the edges making a curve.
Scientists measure liquids using the low point or the meniscus of the liquid in the
cylinder. This is shown by the dotted line in the diagram above.
The volume of this solution is actually
mL and NOT
mL
2. Now, using a graduated cylinder, measure out 100mL of distilled water and put it
into flask A.
This flask contains which molarity of NaCl?
3. Measure 100mL of the remaining NaCl solutions and put them into the following
flasks:
B – 1M NaCl solution
C – 3M NaCl solution
D – 5M NaCl solution
4. Use a hot plate – set it on 8 or high. You may place more than one flask on the hot
plate at a time.
5. When the flask has just reached a rolling boil, put the thermometer into the solution.
DO NOT TOUCH THE SIDES OR BOTTOM with the thermometer. KEEP IT
SUSPENDED in the boiling solution to take your measurement.
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Results
Record your results in the following table:
Boiling Point Temperatures of Different Molar Concentrations of NaCl Solutions.
Flask
Solution
A
0M NaCl
B
1M NaCl
C
3M NaCl
D
5M NaCl
Boiling Pt Temperature (oC)
Questions:
1. Did the amount of NaCl in the water affect the boiling point?
2. Did the results SUPPORT or REFUTE your hypothesis?
3. At the side of the room are containers that show you the amount of NaCl that was
added per 100mL for each molar solution of NaCl you investigated. Looking at these
amounts of NaCl, what do you think about the amount of salt you or anyone else
adds to boiling water to help cook pasta?
YOU ARE GOING TO SUBMIT A LAB REPORT NEXT WEEK IN LAB.
PAGES 26-28 CONTAIN THE DIRECTIONS AS TO WHAT YOU NEED
TO WRITE UP AND TURN IN.
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Week 1 Lab Report Instructions
20 points
• Your lab report should be on separate paper – do not turn in
these pages or you will lose all of the points!!! Make sure you
staple the pages of your report together - unstapled reports will
be docked 2 points (a whole letter grade!)
Keep the lab report in the following order or you will lose points
1. 3pts. An introduction to the experiment: what are you studying and why?
2. 3 pts. Your prediction and hypothesis.
3. 2pts. The table of your data.
4. 5pts. Your impeccable line graph of your data - THIS MUST BE DRAWN BY
HAND ON GRAPH PAPER (there is graph paper on page 27). Read about
Graphing the data below
5. 5pts. Answers to questions below: #8a-e
6. 2 pts. Your conclusion – was your hypothesis supported or refuted? - USE
THESE TERMS AND, EXPLAIN why, using your results
Other criteria: NEAT AND ORGANIZED. I want to see the content ordered as outlined
above. For instance if your graph is after your conclusion, you will lose points.
Graphing the data – learning more about variables
In any experiment there are at least two variables: an independent variable and a
dependent variable. The independent variable is what you, the experimenter, sets or
determines; you purposely set it, do an experiment and measure the outcome, and then
change this variable for the next experiment. The dependent variable is what you
measure during the experiment. It DEPENDS on the independent variable.
In your experiment, which was the independent and dependent variable?
Whenever possible, scientists graph data to show the relationship between the two
variables. Graphs are an easy, direct way to show trends that would be hard to pick
out from a table of figures. From a graph you can easily determine what effect one
variable (for example: the amount of salt in a solution) has on another (for example: the
boiling point temperature).
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Using the graph paper provided on page 29, graph your data following these directions:
1. Determine your axes: x (the horizontal axis) is for the independent variable;
y (the vertical axis) is for the dependent variable.
2. Draw the lines for your axes using a ruler.
3. Now, label your axes and don’t forget the units of measurement for each one.
4. Look at your data and decide on your scales. It is not necessary to start at zero for
each scale (although you may if you wish). However, whatever number you choose to
start at, you must keep the scale evenly spaced along its entire length. Your data points
need to be spread out nicely, not necessarily using the entire piece of graph paper, but
not squished either or too small: the first three are not acceptable, the last one is the
best:
5. Plot the points of your data on your graph and then use them to estimate a straight
line through the data. If you do not know how to do this, ASK!
IF YOU JUST CONNECT THE DOTS INSTEAD OF ESTIMATING A STRAIGHT LINE
OF BEST FIT, YOU LOSE ALL POINTS FOR THE GRAPH.
6. All graphs must have a title. Use one of the following as a title for your graph. Two of
them are good, you can use one of them.
a. Boiling Saltwater Experiment
b. The Boiling Temperature of Saltwater
c. The Effects of Increasing Concentrations of NaCl on the Boiling Point of
Water
d. How to Cook Pasta
e. Increasing Concentrations of NaCl Solutions vs Boiling Point
7. NEATNESS COUNTS, points will be taken off for a messy graph.
8. Answer the following questions:
a. Using your graph, what do you predict the boiling point of a 2M solution of NaCl to be?
b. Using your graph, what do you predict the boiling point of a 4M solution of NaCl to be?
c. Using your graph, what do you predict the boiling point of a .5M solution of NaCl to be?
Turn over…
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In answering questions a – c above, you were asked to predict data within the range of
your experimental data but for concentrations that you did not directly measure. This is
called interpolation of data, and the ability to do this is one of the reasons that
graphing data is so useful to scientists.
Now, you will make predictions by extrapolation of your data.
FIRST, Draw a dotted line extending the line on your chart:
8d. Using your graph, what would you predict the boiling point of a 6M solution of NaCl
to be?
8e. Using your graph, what would be the molar concentration of a NaCl solution which
boiling point was 109°C?
TURN IN YOUR COMPLETED LAB REPORT
AT THE BEGINNING OF THE NEXT LAB PERIOD
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GOOD GRAPH OR BAD GRAPH??
This is a screen shot from an actual website promoting a brand of contact lenses. Review the
information presented in the text and in the graph and then answers the questions below.
1. Can you tell what was measured to create this graph?
2. What data does the Y (vertical) axis measure?
3. Does this graph help you to understand anything about 02 Optix lenses or corneal oxygen
deficiency?
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A graph should allow a reader to look just at the image and understand roughly what you
were investigating (summarized in the TITLE of the graph), and identify any trends in your
results (from the data plotted on the graph) – By now you should realize that the graph from
the O2 Optix website is a BAD GRAPH! – Don’t worry if you weren’t able to answer any of the
questions on the first page. You should not need to read the text of a lab report, website
or whatever, to understand what the graph is showing
Every graph needs:
•
•
•
•
•
A title
Labels and units for both axes (e.g. Distance traveled (meters); Time spent in car (hours) )
The scale should be even – (although you do not have to start at 0)
The scale should be chosen to make sure the data is well spread out on the graph and
easy to interpret.
The axes should be drawn with a ruler
Read the text below, taken from page two of the O2 Optix website.
Use the information in the text to improve the copy of the graph below by adding
everything a good graph needs (check out the list at the top of this page)
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You’ll be using a line graph, rather than a bar graph, to present your data from this week’s
experiment. Take a look at the graph below. It’s an example of a REALLY BAD line
graph.
kg
6
4
2
1
3
5
7
9
10
Fertilizer
Make a list of all the things that are wrong with this graph: (I can think of at least 5
problems with it)
•
•
•
•
•
•
•
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This is actually a graph of data collected in an experiment that tested how the yield of a
corn plant varied depending on how much fertilizer was applied. Here is the raw data from
the experiment.
Amount of fertilizer (grams)
Mass of corn cobs produced (kg)
0
1.6
1
1.9
3
2.6
5
3.0
7
3.2
9
3.8
10
4.7
Use the blank grid below to draw a perfect line graph to display this data. Once you have
plotted all the points on your graph DO NOT join them up…. Read the next section of this
worksheet and then draw a line of best fit for this data.
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Drawing a line of best fit.
A line of best fit is added to a graph when you think that the data points should lie in a
straight line or slope, even if they don’t. Basically it shows where a straight line would go if
the data were perfect and all the points did line up. It averages out all the little errors from
your experiment that make the data points not quite lie on a straight line. To draw a line of
best fit, simply take a ruler and lie it across your graph then move the slope up and down
until roughly half of the data points lie above the line and half lie below it. It is just a
estimation! Then draw that line in on your graph.
Remember, a line of best fit
•
•
•
•
Does not need to go through (0,0)
Does not need to go through the first and last data point
Does not necessarily need to go through all or any of the points
Should be drawn with a ruler
Add a line of best fit to your graph of the corn plant data
Using Graphs to estimate data that you didn’t collect
While one of the purposes of graphs is to display data in a pictorial format so you can see
trends easily, one of the other uses of graphs is to allow you to predict data that you didn’t
collect during your experiment. For example, in the experiment with the corn plants they
didn’t test what the yield would be if 2g of fertilizer was applied, nor if 6.5g were applied,
nor if 12g were applied. BUT a graph would help you to predict what would happen if you
had done those experiments.
To determine the corn yield from a plant treated with 2g of fertilizer, find 2g on the
appropriate axis of your graph, now go up from that point to where you intersect with the
line of best fit. Next read across from that intersection to see what the yield of corn would
be.
Use your perfect line graph to predict how many kg of corn would be produced from corn
plants treated with:
6.5g of fertilizer
12g of fertilizer
If you are reading between data points you actually have collected (for example 6.5g lies
between 5g and 7g which you collected data for), this is called interpolation. If you need
to extend the line of best fit further because you didn’t collect actual data out that far, then
that is called extrapolation (for example, you stopped collecting data at 10g of fertilizer, so
you need to extrapolate to predict what would happen with 12g of fertilizer).
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Graphing two sets of data on the same graph.
Sometimes an experiment will require you to collect two sets of data to compare – for
example the data collected below show the effects of adding fertilizer to both corn plants
and to soybean plants. In this example a scientist could be trying to determine which type
of plant most benefits from getting fertilized, or if it is worth spending money on fertilizer for
corn or on fertilizer for soybeans considering the increase in crop yields it will produce.
Amount of fertilizer applied (grams)
Mass of corn cobs produced (kg)
Mass of soy beans produced (kg)
0
1.6
0.5
1
1.9
0.6
3
2.6
0.6
5
3.0
0.7
7
3.2
0.8
9
3.8
1.0
10
4.7
1.0
This data should be plotted on the same graph – that is the graph will have two lines on it,
one for the corn data and one for the soybean data. By comparing the two lines you can
see the relative impact of fertilizer on the two types of plants.
Use the blank grid below to draw a perfect line graph to display this data. Draw lines of
best fit for each data set and include a key to indicate which line is for the soy bean data
and which is for the corn data.
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Brain Warm Up: Molecular Modeling
INTRODUCTION
A. Answer the following True/False questions – we will go over them in class.
a.
b.
c.
d.
Atoms are made of molecules
A covalent bond occurs when two atoms share a pair of electrons. This creates a
molecule
The number of electrons in an atom can be figured out using information from the
periodic table
The protons of an atom determine how that atom will interact to form bonds with
other atoms
e.
Lewis dot diagrams show how many electrons are in the outer shell of an atom
f.
Covalent bonds are very easy to break
B. Use your periodic table to determine:
The atomic number of Boron
The atomic mass of Boron
How many electrons in one atom of Boron
How many neutrons in one atom of Boron
Complete this Bohr model of Boron:
B
B
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Now draw a Lewis Dot
diagram of Boron:
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Molecular Modeling
Materials needed per group of two
•
1 molecular model kit
Objectives:
•
•
•
To observe relationships between different elements and their bonding
requirements
To become more familiar with empirical (molecular) and structural formulas as a
way to represent molecules
To construct models and corresponding formulas of functional groups found
commonly in organic molecules.
Procedure
1. This is the structural formula for methane. Build it with your kit.
*Use the wooden rods to join the atoms together unless you are
trying to show a double covalent bond in which case you should
use two of the flexible metal rods.
H
H
2. This molecule is composed of which two elements?
C
H
H
3. Draw the Lewis dot diagrams for one atom of each of these elements.
4. Why are there 4 hydrogen atoms surrounding the carbon atom in a molecule of
methane? (The Lewis dots you just drew actually show you the answer.)
5. What do the sticks on your model actually represent?
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6. The molecular formula for methane (the molecule you just built) is CH4.
The molecular formula for ethane is C2H6.
Draw structural formula of ethane AND build it with your kit.
7. Now, remove one H from your model and replace it with an oxygen atom, then reattach
the hydrogen atom to the new oxygen atom. (There should now be an oxygen atom
between one of the carbons and one of its hydrogens). Congrats! You just made
ethanol or drinking alcohol. Draw out its structural formula.
8. Look at the functional group handout (page 45) circle the functional group you just drew
in your structural formula from #7 and write its name next to it.
9. Draw a Lewis dot for an atom of oxygen in this box.
How many valence electrons does it have?
How many more electrons does it want?
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10. Look on your functional group handout (page 45) for the group called carboxyl.
a. Look at the O atom in the molecule.
How many covalent bonds does it form?
Why does it form this number of bonds?
b. The double covalent bond formed between oxygen and carbon means that, in total,
how many electrons are being shared between the Carbon atom and the Oxygen
atom?
c. A carboxyl group looks like it could be two different functional groups that are smaller
but it is NOT. Which two functional groups does it look like?
and
Remember, when you see this functional group it is one group:
11. Draw a Lewis dot diagram for one atom of nitrogen and one atom of sulfur.
12. This is cysteine, an amino acid that is found in hair. This is the
amino acid that makes hair stink when you burn it because it has
sulfur in it. Build this with your kit – you must build it in order to
answer the questions correctly. Remember to use the flexible
metal rods to create the double bond between the C atom and
the O atom in the carboxyl group in this molecule.
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a. Your kit contains four different color balls to represent H, O, N and C atoms. However
you also need a sulfur atom to build this molecule. One of the colors is going to have to
“double up” and represent sulfur as well. Which color ball are you going to use to
represent the sulfur atom in this molecule?
WHY??
b. Look at your model, how many covalent bonds does the nitrogen atom have?
c. Now, look at the drawing of cysteine above. Does it clearly show all of the covalent
bonds?
c. Draw out the molecule of cysteine more completely.
d.. There are three functional groups on the cysteine molecule. Circle all three on the
structural formula you drew in c above, and name them next to the functional group.
e. Now, look at the Lewis dots of sulfur and nitrogen from above. Do the Lewis dot
diagrams make sense with the completely drawn out molecule from c? That is, do the
bonds correspond to the Lewis dots? EXPLAIN.
f. Your Lewis dots of oxygen and of sulfur look the same yet we call them different
elements. How are they different?
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13. Now, for something larger.
a. Build a molecule of glucose in its linear form:
SHOW ME THIS MODEL BEFORE YOU GO ON.
b. Circle and name the two functional groups on C1 and C5
14. Use your model FROM 13 to fold up into the ring form of glucose. Here are the
directions written out AND a picture underneath to show how it folds.
•
The six corners of the hexagon are made up of the first five carbons of the chain
and the oxygen from the hydroxyl group of the fifth carbon. That oxygen will end up
bonded both to carbons #5 and to carbon #1 to complete the ring.
•
You will need to rearrange the atoms of the aldehyde group (C#1) and hydroxyl
group (C#5) to close the ring and still keep all the atoms satisfied.
SHOW ME THIS ONCE YOU HAVE MADE IT
15. Look at C1 and C5 in both the diagrams of glucose above. Do you see the functional
groups you circled in 13b in the molecule pictured on the left? CIRCLE THEM AGAIN.
Are they present in the molecule on the right?
16. What “function” do these two groups have? That is, what do they allow this molecule to
do?
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Next week you will have a quiz over functional groups, and molecular formulae. Use
this time right now to make a set of flash cards. You will need to know the
structural formula for each functional group and its name. For example:
O
carboxyl
C O
H
Make sure you understand why this line is here, and know how to spell the words correctly.
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Functional Groups of Organic Molecules
O
R
C
Aldehyde
**There are two types
of carbonyl groups
Ketone
H
R
C
O
R
H
Methyl group
R
C
H
H
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Solvent properties of oil, water, and alcohol
Any liquid may be a solvent, meaning that it can dissolve (separate and surround)
particles of another substance (solutes) to make a mixture (solution).
You are going to investigate whether or not certain substances dissolve in water, oil or
alcohol.
Procedure
a) Label three small test tubes “O” for oil, three “W” for water and three “A” for alcohol.
b) Put equal volumes of oil or water or alcohol into the appropriate test tubes (about
one quarter full – It doesn’t have to be exact).
c) Number the tubes in each set 1, 2, and 3. (E.g. O1, O2, O3, )
d) Using the flat toothpicks, place a small and equal amount of salt in Tubes O1, W1
and A1, a small and equal amount of sugar into all #2 tubes, and a small and
equal amount of β-carotene into all #3 tubes.
e) Cover each tube with a square of parafilm* and mix by tilting back and forth gently
for at least 30 seconds.
*IF you’ve never used parafilm before, ask your instructor for a demonstration of how to
use it.
Results:
In the table on the next page, DESCRIBE WHAT YOU SEE in each tube.
Avoid using the term “dissolve.” Stick to visual observations, such as:
• Are crystals visible?
• Did the crystals disappear in either or both liquids?
• Are there more crystals remaining in one liquid than in the other?
• Do the orange β-carotene powder smear and spread color through the liquid, or do
they remain intact and separate?
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SOLVENT
OIL
WATER
PROPANOL
SOLUTE
1
Salt
2
Sugar
3
β-Carotene
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Brain Warm Up: Organic Macromolecules
INTRODUCTION
Read the first section of the organic molecules lab and then answer the following True/False
questions – we will go over them in class.
a.
When the copper ion Cu2+ receives an electron and becomes Cu1+, a chemical
reaction has occurred.
b.
The glucose provides the electron to Cu2+.
c.
The glucose gets oxidized.
d.
A glucose molecule has double bonds when it is in the ring structure.
e.
f.
The reduction test is for detecting glucose only – this is the only monosaccharide
that exists.
A chemical reaction occurs when atoms are rearranged to make new molecules
called products.
A. How will you know that you do or do not have a reducing sugar in your test tube? That is,
what will happen to the solution in the tube when you do the test if you do have reducing
sugar or don’t have reducing sugar?
B. According to the figure on page 51, you have no idea what the glucose becomes once it is
oxidized. The only product that you know about for this reaction is:
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Testing for Organic Compounds
You and your partner have just obtained jobs working as lab techs behind the scenes for
EMCC’s science labs. (YES – these positions really exist – if you are interested in one of
these part-time work-study jobs talk to your instructor at the end of the semester!). In the
freezer you have found 3 boxes, labeled A, B and C. Box A contains bottles of soy flour,
glucose, non-fat milk, and white flour; Box B contains bottles of starch, corn syrup, sucrose
and egg white; Box C contains bottles of glucose, table salt, starch and gelatin.
Unfortunately all the labels have fallen off the individual bottles in each box and all the
liquids are similar looking whitish or clear fluids. You are going to have to test the contents
of each bottle from a box to figure out what is in each one!
However, since you are brand new you must first learn the tests that are used to detect the
each type of organic macromolecule. There are tests for monosaccarides, polysaccarides,
and proteins:
Organic molecule
Name of test or reagent
Carbohydrates – monosaccharides
Eg: glucose, fructose
Carbohydrates – polysaccharide
Starch only
Proteins
Benedict’s test
I
IKI (Iodine potassium iodide)
odine test
Biuret test
Your supervisor tells you that you are to work with your partner and perform each of these
tests so you can see the results for yourself and learn what a positive and negative test
looks like.
The first test you read about is the Benedict’s test. This is a test that will detect the
presence of monosaccharides in a liquid. First, you need to learn the chemistry behind this
test.
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The first thing you learn is that this test is also called the Reduction test for reducing
sugars. “Reducing sugars” is another name for monosaccharides. This is because
monosaccharides have either ketone or aldehyde groups that allow them to reduce copper
ions. You find the following figure in a text book:
e-
Cu1+
(RED)
Cu
2+
blue
SO42-
You recall your basic
inorganic chemistry:
Reduction: gaining of
electrons and
Oxidation: loss of
electrons
CuSO4
(BLUE)
Benedicts Reagent
Monosaccaride
eg. glucose
You also recall your basic
organic chemistry:
Monosaccharides in
solution “flip” in between
linear forms and ring
structures. It is only in the
linear form that they have
a double bond (carbonyl
group) that allows them to
reduce copper ions
You remember that reduction occurs when an atom or ion gains an electron. The atom or
ion that gained the electron(s) is said to have been reduced. The substance that gave
away or lost the electron is said to have been oxidized (LEO GER). Monosaccarides are
also called reducing sugars because they are able to give away or loose an electron from
the double bond in their carbonyl functional group. The sugar itself is oxidized when this
happens and the substance that picks up the electron is reduced.
Benedicts reagent is used in the reducing sugar test. Benedicts reagent is a solution that
contains copper sulphate: CuSO4. Copper ions can exist in two different states Cu2+ and
Cu1+ , depending on the number of electrons the ions have. Conveniently, Cu2+ is a blue
color and Cu1+ is a brick red color, so you know, just by looking at a tube, whether the
copper is in the 2+ or 1+ state. Benedicts reagent is a blue color so it contains Cu2+ ions.
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To do the benedict’s test you mix the blue colored Benedicts reagent with the substance you
are testing and heat it up. If a monosaccaride (AKA a reducing sugar) is present, it will lose
an electron and give it to the Cu2+ ions, turning them into Cu1+ ions – and the solution will
turn from blue (thru green and yellow/orange colors) to red.
Blue
Possible results:
no sugar
Green
Yellow/Orange
Brick red
Increasing amount of monosaccaride
Your supervisor tells you to do the Benedict’s test on five different solutions so you can see
what it looks like for yourself. You decide to do it on water (this should give you a negative
result) and then one solution that you KNOW contains a monosaccaride (this will show you
what a positive result looks like), one that you KNOW contains a polysaccharide (which
should give a negative result) one that you KNOW contains a disaccharide (which should
also give a negative result) and one other solution or substance of your choice.
After going over the chemistry of the Benedict’s test, you begin to recall certain facts about
the substances you could pick:
•
•
•
•
•
•
Glucose is a monosaccaride
Fructose is a monosaccaride
Sucrose is a disaccharide: it does NOT have a double bond for reducing copper ions
Sucrose is made of fructose and glucose
Starch and cellulose are polysaccharides and do NOT have many double bonds; they
have a few at the ends of their long molecules
Milk contains mainly a disaccharide, lactose. Milk also contains a small amount of
galactose which is a monosaccharide.
Use the table below to record the names of the specific solutions/substances that you are
going to use for your benedicts test and what you would expect the result of the test to be.
Substance
1
Water
2
KNOWN monosaccharide
3
KNOWN disaccharide
4
KNOWN polysaccharide
5
YOUR CHOICE
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You are now ready to perform your experiment:
Procedure for Benedict’s Test
1. Fill a 400mL beaker about half full of tap water and bring the water to a very slow boil on
your hot plate – set the hot plate at 8 then turn down when it just starts forming bubbles.
2. Get five clean test tubes and label them 1 - 5. Use the list of substances that you created
in the table at the bottom on page 59 - Put 2mL of each solution or liquid to be tested into
the appropriate tube.
3. Add 2mL of Benedict's Reagent to each test tube.
4. Thoroughly mix the contents of the tubes.
5. Put all the tubes into the boiling water bath at the same time and heat them for 30
seconds.
6. Promptly remove the tubes and observe for any color change. Let the instructor see if
they are ready - they may need to be returned to the water bath for slightly longer.
7. Record the results in Table 1 below.
TABLE 1. BENEDICT’S TEST. A negative result (no change in color) should be recorded
as a minus (–) sign. Various degrees of color change should be recorded as plus (+) signs
(e.g., red would be ++++, yellow would be +++ and so on).
Tube Material
Color after heating
Test result (positive/negative)
1
distilled water
____________________
____________________
2
___________
____________________
____________________
3
___________
____________________
____________________
4
___________
____________________
____________________
5
___________
____________________
____________________
Put your general findings about the benedicts test into the summary table on page 61
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Although the benedicts test cannot detect disaccharides such as sucrose, it is possible to
“pre-treat” disaccharides like sucrose with acid and this will break apart the disaccharide into
two monosaccarides. After the pre-treatment step, once you have monosaccharides in the
test tube, then if you do a benedicts test you will get a positive result.
Here is a basic diagram of what is going on:
G
F
Even though sucrose is
made of monosaccarides,
they are NOT single
monosaccarides - they have
been joined together to make
a disaccharide.
sucrose
So sucrose will give a negative benedict’s test…UNLESS…you first heat it up with acid. The
acid will break apart the disaccharide molecule to create two separate monosaccarides.
Once the sucrose has been broken down into its constituent monosaccarides, the Benedict’s
test will give a positive result.
G
acid and boiling
Sucrose
(disaccharide)
glucose
(monosaccharide)
+
F
fructose
(monosaccharide)
To confirm this, you want to see if you can get a positive reducing sugar result when sucrose
is pre-treated with heat and acid to hydrolyze it to glucose and fructose before doing the
benedicts test.
Procedure for Hydrolysis of Sucrose
1. Obtain three (3) clean test tubes. Label them A, B, and C.
2. Put 2 mL of 5% sucrose into tubes A and B. Put 2 mL of distilled water into Tube C.
3. Add 20 drops of 5% HCl (hydrochloric acid) to tubes A and C.
4. Add 20 drops of distilled water to tube B.
5. Place the three tubes into the boiling waterbath for 6 minutes. This step will hydrolyze any
α-glycosidic linkages that are present in the sample
CONCEPT CHECK!!
After step 5 of the procedure, which tube(s) A, B or C, should contain
molecules that can turn the benedicts solution red?
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6. Now test the solutions in each of the three tubes for reducing sugars, following the
procedure for the Benedict’s test as follows:
a) Add 2mL of Benedict's Reagent to each test tube.
b) Thoroughly mix the contents of the tubes.
c) Put all three tubes into the boiling water bath at the same time and heat them for 30
seconds.
d) Promptly remove the tubes and observe for any color change. Let the instructor see
if they are ready - they may need to be returned to the water bath for slightly longer.
Enter your results in table 2 below:
TABLE 2. HYDROLYSIS OF SUCROSE: REDUCING SUGAR TEST. Indicate by a plus
sign (+) if reducing sugar was present after the treatment to hydrolyse the sucrose; indicate
by a minus sign (—) if no reducing sugar was present.
Benedict’s Test Result
Tube
Material
Benedicts Test Result After Hydrolysis
A
sucrose + HCl
B
sucrose without HCl
__________________________
C
HCl without sucrose
__________________________
_________________
Are these the results you
would expect? Why/Why
not?
Which tube(s), A, B or C, would you expect to contain dissaccarides after step 5 of the
procedure?
Did your results confirm this? How can you tell?
Which tube(s), A, B or C, would you expect to contain monosaccarides after step 5 of the
procedure?
Did your results confirm this? How can you tell?
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You are now ready to conduct more tests – follow the procedures and learn how to do the
tests to detect starch and and proteins.
Procedure for IKI test to detect starch
To learn how to do the test for the presence of starch, chose a few solutions to test – at least
one that you know will give a positive test result and at least one that will give a negative test.
1. Obtain a porcelain spot plate.
2. Put 3 drops of the material to be tested into a depression of the spot plate. Write the
original color of each material tested.
3. Add 3 drops of IKI to each material in the spot plate.
4. Observe the color of the material after the addition of the iodine.
5. Record the color information in Table 3
TABLE 3
Substance
Original color
Color after IKI Test
Positive or Negative
for starch
Put your general findings about the IKI test into the summary table on page 61
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Procedure for Biuret Test to detect Proteins
To learn how to do the biuret test to detect proteins, chose a few solutions to test – at least
one that you know will give a positive test result and at least one that will give a negative test
result.
1. Put 1.5 mL of the test material into a clean test tube.
2. Add 1.5 mL of Biuret reagent to the tube and mix gently.
3. Observe the tube for a change of color. A purple/violet color should develop if protein is
present. When looking for comparative color changes, a white sheet of paper held behind
the test tube may help - Do not confuse the blue color of the buiret reagent with the violet
color you are seeking.
Record your results in Table 4.
TABLE 4. BIURET TEST.
Material
Original Color of Material
Positive/Negative
Color After Biuret Test
for Protein
_________
_________
_________
_________
Put your general findings about the biuret test into the summary table on page 61
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Analyzing the unlabelled samples
On the side bench are three boxes (A, B and C) each containing four bottles. Each bottle is
unlabeled, but box A contains bottles of non-fat milk, glucose, soy flour, and white flour; Box
B contains bottles of corn syrup, egg white, sucrose, and starch; Box C contains bottles of
glucose, gelatin, table salt, and starch. . Each pair will select one unknown from each of the
boxes and perform the various tests you just learned about to identify what’s in the bottle.
This is like a detective challenge. You will have to decide which tests you do on your
unknowns to narrow down which of the four possible solutions your unknown could be.
Record your results in the table below using the following scale:
Very positive
Positive
Slightly positive
Negative
Unknown sample test results
Benedict’s
Sample
#
Test
Unknown
from box A
Unknown
from box B
Unknown
from box C
Iodine
Test
+++
++
+
-
Biuret
Test
Identity of sample
Use the nutritional data table below to determine the identity of each of your unknown
samples.
*** MAKE SURE YOU REMEMBER WHAT EACH TEST IS TESTING FOR ***
Protein
Starch
Monosaccharides
(mg/100g )
(mg/100g)
(mg/100g)
720
0
54
Egg White
860
0
0
1% gelatin
0
0
910
1% glucose
Trace
0
825
1% corn syrup
3580
0
515
1% non-fat milk
830
600*
Trace
2% soy flour
0
0
0
1% sucrose
0
0
0
Sodium chloride
Trace
850
0
1% starch
210*
1522
100*
2% white flour
*These amounts are at the lower limits of what our tests can detect.
Sample
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Denaturation and Coagulation of Proteins
Cooking and baking is really all about chemistry. Extremes in pH and temperature effect
macromolecules such as proteins and fats. In the food industry both conditions are used to
help preserve foods: acids are used as preservatives in foods and pasteurization is a process
of heat treating foods that kills harmful bacteria. However, you have never really “seen” this
in a lab. You decide to investigate the effects of heat and acid on proteins since these
macromolecules are important as enzymes which control chemical reactions.
1. Obtain 2 clean test tubes and label them A and B. Do not reuse any tubes from the
other of today’s experiments. There should be a supply of fresh test tubes by the
fume hood.
2. Put 5mL of 5% egg albumen (kept in the fume hood) into Tube A. Place tube A into a
boiling water bath for about 5 minutes. Watch this tube carefully and remove it from
the water bath if the albumen starts to move up the tube.
3. Put 5mL of 5% egg albumen (kept in the fume hood) into Tube B. Add 20 drops of
concentrated hydrochloric acid (HCl). This is also in the fume hood at the back of the
room.
4. BE CAREFUL! Do not get acid on your hand or clothes.
5. Gently shake the test tube to mix the contents.
6. Wait 5 minutes and examine both tubes.
Draw a picture at the molecular level showing the difference between the tubes. SHOW
YOUR INSTRUCTOR. Here is some help: a picture of the protein solution before acid or heat
treatment:
water
Proteim
molecule
Protein solution – the
protein molecules
(diamonds) are
spread out throughout
the liquid
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Measuring pH
pH can have a great effect on biological and chemical reactions. For future reference for
your upcoming experiments as an EMCC lab tech you decide to determine the pH of various
common substances.
Use pH indicator strips to determine the pH of the solutions in the table below.
pH
To use a strip:
Coffee
1. Wet all three sections of the strip with the solution
you are testing.
2. Compare all the spots of color on the strip with the
columns of colors on the color chart on the strip
container.
3. Find which column of colors on the chart best
matches the series of colors on your test trip and
read off the pH from the chart.
milk
Orange juice
Coke
Detergent
Tap water
Record the pH of the substance you measured in the table, and add your data to whiteboard.
Complete the table by getting the rest of the pH measurements from your co-workers.
Answer the following questions:
Which solution is the most acidic?
Which is the most basic?
Is coffee more or less acidic than orange juice?
Which solution has the highest concentration of hydrogen ions?
What is the concentration of Hydrogen ions in coke?
Bleach has a hydrogen ion concentration of 10-13 M / L. What is the pH of bleach?
Does bleach contain more or less Hydrogen ions than milk?
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LAB SUMMARY NOTES
NAME OF TEST
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TYPE OF MOLECULE
IT TESTS FOR
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WHAT A POSITIVE
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WHAT A NEGATIVE
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LAB SUMMARY NOTES
Chemistry behind the Benedicts Test
Notes about pH
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Brain Warm Up: Osmosis and Membranes
INTRODUCTION
A. Answer the following True/False questions – we will go over them in class.
a.
b.
c.
d.
e.
f.
Water molecules are able to easily move across the phospholipid bilayer of a
plasma membrane
Diffusion is the movement of molecules from an area of their low concentration to
an area of their high concentration
Large molecules like sucrose and starch can cross a cell membrane easily
Water will move across a membrane from an area of low solute concentration to
an area of high solute concentration.
If a plant cell takes up too much water it may lyse
Osmosis refers to the movement of solutes such a glucose and sucrose across a
plasma membrane
B. The pictures below show two animal cells. The circles represent water molecules,
the triangles represent sucrose molecules.
For the diagram on the left, add the gradient
triangle, describe the cell and its
environment and say what will happen to
water in this case.
For the picture on the right, add water and
sucrose molecules according to the
description given, and add a gradient arrow
(triangle) to the diagram
The environment is hypotonic to
the cell.
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Determining the Solute Concentration of Potato Cells
In this experiment you are going to determine the solute concentration of potato cells. That is,
you are going to figure out the concentration of solutes within living potato cells. This is
important information for farmers and the food industry. Potatoes with higher amounts of
solutes and colloidals in their cells, when cooked, give a light fluffy texture and a full firm
appearance and they are preferred by fast food manufacturers for fries and baked potatoes.
MacDonald’s is the largest purchaser of potatoes in the US and spends considerable time
and money monitoring the chemical make-up of the potatoes they use, and determining how
changes in potato chemistry affect the quality of their fries. To ensure a market for their crop,
potato farmers must grow varieties of potatoes that will conform to MacDonald’s
requirements.
Procedure:
1. Prepare 3 dishes of potato slices as follows:
a. Using the same potato for all three dishes, trim away all the brown "skin" and use the
potato peeler to pare off a total of between 15 – 20g of potato
c. Place the thin slices into 3 petri dishes. Each dish must have very close to 5.0 grams of
potato in it. You will probably have to cut some slices to get to 5g in each dish.
d. Record the precise weight of the slices in each dish. THIS STEP IS VERY IMPORTANT!.
2. Cover the potato slices in each petri dish with one of the following solutions:
Dish 1 distilled water
Dish 2 0.3 M sucrose
Dish 3 0.5 M sucrose
See Fig. 1. Make sure that all the slices are submerged.
Fig. 1 Petri dish with potato
slices in solution.
3. Allow the dishes to sit undisturbed for at least 60 minutes.
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WHILE YOUR SLICES ARE SITTING IN THE PETRI DISHES, YOU ARE GOING TO DRAW
A PICTURE OF WHAT YOU PREDICT WILL HAPPEN AT THE MOLECULAR LEVEL TO A
POTATO CELL WHEN IT IS SUBMERGED IN DIFFERENT CONCENTRATIONS OF
SUCROSE.
Divide the mini-whiteboards into three areas and draw a potato cell
in each area. Each area will represent one of the three different
molarities of sucrose that the potato cells are bathing in.
This is a diagram of a potato cell. Do not draw the pieces of the
potato in the dish, but a single cell like this one in each of your
three areas on the white board.
You will create 3 “cell+environment” diagrams predicting the movement of WATER ONLY
across the cell membrane. These diagrams will be your prediction as to what will happen in
your dishes based on your hypothesis.
YOU MUST SHOW/PREDICT THE RELATIVE AMOUNTS AND LOCATIONS (INSIDE OR
OUTSIDE THE CELL) OF THE FOLLOWING MOLECULES IN YOUR DIAGRAMS:
Water
Starch
Sucrose
AND, ADD A CONCENTRATION TRIANGLE TO SHOW WHERE (INSIDE OR OUTSIDE
THE CELL) YOU PREDICT THERE ARE MORE SOLUTES AND YOU MUST ADD
ARROWS TO SHOW THE MOVEMENT OF WATER
How can you draw these molecules? Why, with different shapes, of course!. Such as
squares, triangles, circles etc. Think about which molecules would be outside the cell, and
which would be inside.
Under each drawing write three sentences describing:
a) Whether the cell is hypo-, iso- or hypertonic to its environment
b) Which direction water will move
c) What will happen to the weight of the potato cells (will it increase, decrease or
stay the same).
THE INSTRUCTOR MUST APPROVE YOUR DRAWINGS.
These drawings will be your hypothesis and predictions in a picture format. You MUST have
a copy of the drawings before you leave class to take with you since you will hand in
your drawings as part of a lab report.
While you are waiting to complete this experiment, complete the
Cell Membranes and Osmosis Lab Stations worksheets on pages
74 & 75 of this manual. The three stations are set up on the side
and front benches.
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4. After at least 60 minutes, remove the slices, blot off excess solution and reweigh each
group exactly as you did at the beginning of the experiment.
5. Record the new weights in Table 1. Review the significance of these changes in weight
and, in the space provided, indicate whether your data indicate each solution to be
hypertonic, hypotonic or isotonic compared to the potato cells.
Table 1. Weight of Potato Slices Before and After Submersion in Sucrose Solutions of
Different Molar Concentrations
Molar Concentrations of Sucrose
0.00
0.30
0.50
Original Weight (gm)
Final Weight (gm)
Change in Weight (+/- gm)
Tonicity of solution
compared to potato cells
6. Produce a lab report documenting your experiment, results and conclusions. Follow the
instructions on the next page to complete your report. Your report is due at the beginning
of the next lab session.
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Osmosis Lab Report and Questions
Your lab report must be done on a computer EXCEPT THE GRAPH. The ENTIRE REPORT
must be NEAT! Invest in a stapler!!!
Structure the report as follows:
1.
Introduction (5 points). What is the basic idea behind the lab? Introduce the
concepts that we have been studying and how we are experimenting with these
concepts in this lab. How is the data you collect going to enable you to determine
the solute concentration inside the cells? What is the point of weighing the slices
before and after? This should be about one or two paragraphs long. I DO
NOT want to read a description of the procedure with the slices and the dishes.
2.
Table of Results (5 points). Display your results in a table, including the
differences in mass you calculated.
3.
Graph of Results (10 points). Use the graph paper on page 69 and draw your
graph by hand. Graph the molar concentration of sucrose solution against the
change in mass of the potato pieces. Estimate a straight line thru the points and
mark an X where the line intersects with 0 weight loss/gain.
Don’t forget all the things that a good graph needs [e.g., variables on the
correct axes, scales evenly spaced, axes labeled (including units as
appropriate), points plotted accurately and line drawn, graph meaningfully
titled, graph paper and straight edge used.]
4.
5.
Analysis of results and conclusions (10 points):
Use your results table and graph to answer the following questions as precisely as
possible.
a.
Based on your graph, would you expect potatoes immersed in 0.2 M sucrose
to lose weight, gain weight or remain the same size?
If you expect a change in weight, how many grams will be gained or lost?
b.
Which potato cells would be expected to have the greatest turgor pressure,
those immersed in 0 M sucrose solution or those immersed in a 0.5 M
sucrose solution? Explain your answer
c.
What was the solute concentration inside your potato cells? How can
you tell? (Your answer to the first part of this question (the solute
concentration) should be a NUMBER, and the units are Moles)
d.
Did your data support or refute your hypotheses? (For this experiment your
hypotheses were the whiteboard drawings where you predicted what would
happen in each dish and why). Explain.
Attach your pictures from lab (your drawn hypothesis for what you predicted
would happen in each dish) to the back of your report for 3 extra bonus points!!.
Turn in your report at the next lab session
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Cell Membranes and Osmosis Lab Stations
Raisins and Strawberries (on side bench)
Examine the raisins and strawberries. The raisins have been submerged in distilled water for
48 hours, the strawberries have had sucrose sprinkled on them for 48 hours.
•
Describe the strawberry before the sucrose was sprinkled on it
•
Explain the condition (you are choosing one of the terms to use: hypotonic, hypertonic,
isotonic) of the strawberry cells compared to their environment once the sucrose was
sprinkled all over the strawberry. Remember to write your answer as a complete
sentence.
•
What does the strawberry look like after the sucrose has been on it for 48 hours? Why
does it look this way?
•
Describe the raisins before they were submerged in distilled water
•
Explain the condition of the raisin cells compared to their environment once they are
dropped into the distilled water.
•
What do raisins look like after they have been soaked in water for 24 hours? Why do
they look this way?
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Red Blood Cells (on front bench)
Examine the pictures of normal red blood cells and of red blood cells that have been bathed
in distilled water and in 0.5M Salt water
•
Which tube has an isotonic environment for the red blood cells?
•
Describe the environments of the other two tubes compared to the RBC’s
•
What will happed to the cells in tube 1 and why?
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Brain Warm Up: Cells
INTRODUCTION
Answer the following True/False questions – we will go over them in class.
a.
b.
c.
d.
e.
f.
Animal cells have cell walls.
Eukaryotic cells have a nucleus.
Plant cells do not have a cell membrane, only a cell wall.
Osmosis is the movement of water only across a membrane.
Vacuoles are storage organelles for plants.
Many molecules can move past a cell wall but will not easily move across the cell
membrane.
A. CELL STRUCTURE – You are going to draw cells, in particular, several plant cells.
a. What is the dark line on the outside representing?
b. What is the thin line on the inside representing?
_____
c. What could the other organelle inside this cell be?
Actually, when you see plant cells they look more like this with other organelles, dots, etc
inside them:
d. Why would they look like this (one thick outer line only visible) and not the picture above?
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Microscope and Cells Lab
Parts of the Compound Microscope
The compound microscope has a number of parts whose functions you must know in
order to use the instrument successfully.
1) Arm - the vertical piece that connects the lens system of the microscope to the stage and
lighting system. The microscope should always be carried with one hand firmly grasping
the arm.
2) Base - the flat bottom on which the microscope rests. The microscope should always be
carried with one hand firmly under the base and one hand around the arm.
3) Body - the housing that contains the optical system through which light passes from the
slide material to your eye.
4) Oculars - the eyepieces; one of the two sets of magnifying lenses found in the compound
microscope. Locate a number on the edge of the ocular lens - this indicates its
magnification.
5) Revolving Nosepiece - the thin stainless steel ring located at the bottom of the body,
capable of rotation. Attached to the nosepiece are the objective lenses. The revolving
nosepiece makes it possible to change objective lenses, thus, making it possible to
change the magnifications used.
6) Objective Lenses - the second type of magnifying lenses found on the compound
microscope. There are four objective lenses on these microscopes. The magnification of
each is indicated by a large number on the side of the lens.
YOU ARE NOT GOING TO BE USING THE 100X LENS.
a) scanning lens - the shortest objective lens. Magnification = ____
b) low power lens - the next to smallest lens. Magnification = ____
c) high power lens - the next to longest lens. Magnification = ____
Total Magnification =
Ocular lens power x Objective lens power =
10 X OBJECTIVE LENS POWER =
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7) Stage - the platform on which a slide containing material to be studied is placed. Notice
the hole in the center through which light passes. Specimens must be centered over this
hole.
8) Mechanical Stage - clamps for holding a slide and knobs which allow smooth controlled
movement of the slide forwards and backwards, and side to side. The mechanical stage
lies on top of the stage and the knobs are located below and to the right of the stage.
9) Condenser - a cone-shaped structure mounted directly underneath the stage. Lenses
within the condenser concentrate the beam of light traveling upwards from the light
source and cause the light to strike the material at an angle. The condenser may be
raised or lowered by means of a black condenser knob located beneath the stage on the
left side. However, the condenser should generally be left at its highest position.
10) Iris Diaphragm - a ring or thin black lever extending out from the condenser. Built into
the condenser is an iris diaphragm that regulates the amount of light permitted to pass
through the condenser and reach the stage. The iris diaphragm operates like the iris of
the eye; when it is open, more light passes through; when it is closed, less light passes
through. The diaphragm is opened and closed by moving the lever or rotating the ring
from side to side.
11) Light - the light source for these microscopes is built into the base of the instrument,
allowing light to travel straight up into the condenser.
12) Coarse and Fine Adjustment Knobs - pairs of knobs located at the base of the arm on
both sides of the microscope. The adjustment knobs are used in focusing the material
that is to be studied.
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Figure 1. Compound Light Microscope – Add Labels for the parts listed above
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A. Focusing and examining material - (We will go through this as a class)
1. Obtain a prepared slide and place it between the mechanical stage clamps by pulling
back the left clamp and placing the slide firmly against the horizontal bar and the right
hand clamp. Gently release the left clamp.
2. Using the mechanical stage knobs, move the slide so that the material on the slide is
directly over the lit circle. Do this while looking directly at the slide, not looking through
the microscope.
3. Using your scanning lens, raise the slide with the coarse adjustment knob until it is in
focus
4. Sharpen the focus by rotating the fine adjustment knob. The circular area observable
through the oculars is called the FIELD.
5. Observe the material carefully, and place in the center of the field that portion you wish to
examine in greater detail.
6. Carefully rotate the nosepiece one position so that the low power objective lens clicks into
position.
7. Refocus the material using the fine adjustment knob. Under low power it may be
necessary to use the coarse adjustment knob in refocusing.
8. Recenter the material and adjust the iris diaphragm, if necessary, to improve contrast and
make the material clearer and brighter. For some materials this is all the magnification
required, but frequently it will be desirable to magnify the material further.
9. Watching from the side CAREFULLY rotate the nosepiece another position allowing the
high power objective lens to click into place. IT IS ESSENTIAL THAT THE SLIDE IS
PERFECTLY FOCUSED UNDER LOW POWER BEFORE YOU SWITCH TO HIGH
POWER.
If the slide is properly focused under low power, the high power lens will fit into position.
If the slide is not properly focused under low power, the high power lens will not fit, and
the slide may be cracked.
The lenses on these microscopes are PARFOCAL, meaning that when material is in good
focus under low power, it should still be in rough focus under high power.
10. Using the FINE ADJUSTMENT ONLY refocus the slide. NEVER use the coarse
adjustment knob when the high power lens is in place.
11. Open the iris diaphragm slightly to brighten the field. The high power objective lens has a
smaller objective lens so less light enters the lens. Therefore, it is necessary to
compensate by increasing the amount of light passing through the condenser.
12. Move the slide around to examine in detail all areas of the material. Focus up and down
with the fine adjustment knob to see the three dimensional aspects of the material.
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B.
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Removing a slide
1. When you have finished studying a slide, rotate the nosepiece backwards through low
power to the scanning lens.
2. Lower the stage.
3. Carefully remove the slide from the mechanical stage clamps and return the slide to the
appropriate box.
C.
Putting the microscope away
1. When you have finished using the microscope for the day make sure:
•
the scanning lens is in position,
•
the stage is lowered,
•
the last slide has been removed from the stage.
2. Turn off the light and unplug the microscope. Cover the microscope with the dust cover.
3. Carefully return the microscope to the CORRECT cubicle in the microscope cabinet.
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Exercises with Prepared Slides
A.
Slide of Colored Threads
The vertical distance between the objective lens and the slide or specimen on the stage is
called the WORKING DISTANCE.
Working Distance
1. What is the relationship between magnification and working distance? (That is, as you
increase magnification, what happens to the depth working distance?)
Why is this significant for deciding which knob to use to focus your specimen when
using the high power (40X) objective lense?
The vertical space or “thickness” that is in focus at any given time is called the DEPTH OF
FIELD.
2. Carefully observe the slide under the scanning, low power, and high power objectives.
How many threads are in sharp focus simultaneously using the scanning lens?
How many threads are in sharp focus simultaneously using the low power lens?
How many threads are in sharp focus simultaneously using the high power lens?
3. What is the relationship between magnification and depth of field? (that is, as you
increase magnification, what happens to the depth of field?)
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4. Observing the slide under high power, use the fine adjustment knob to slowly move
through the stack of threads and determine which thread is on top, which is in the middle,
and which is on the bottom. (The slides are not all the same!)
Top:
_________________ Middle: ________________ Bottom: ________________
The area or the circle of light that you see at any given time is called the FIELD OF VIEW.
5. Rotate the nosepiece backwards to low power. Move the slide so that just one of the
threads is at the very edge of the field like the picture to the below. Carefully return to
high power but don’t move the slide again. Refocus.
Is the thread visible? _________________
Explain what happened and why.
How can this problem be avoided?
6. What is the relationship between magnification and field of view? (that is, as you increase
magnification, what happens to the diameter of the field of view?)
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B. Preparation of Wet Mounts
1. Place a drop of water on a clean slide.
2. Obtain a small amount of material and place it in the water on the slide. If the material to
be studied is thick, obtain a slice less than 1 mm thick.
3. Carefully cover the specimen with a cover slip. To avoid trapping air under the cover slip,
first put one edge of the cover slip on the slide in the water drop and then slowly lower the
cover slip onto the slide. Refer to Fig. 3.
4. Observe the slide under the microscope.
5. If a stain is to be used, add a drop at the edge of the cover slip. Draw the stain under the
cover slip by placing a piece of paper towel on the opposite side.
Fig. 3. Steps in Making Wet Mount
Making a Wet Mount to View Microorganisms in Pond Water
a) Make a fresh slide preparation of the pond water available. (You do not need to put a
drop of clean water on the slide first). Try to get good debris on your slide – small
organisms hang on to debris and live in it. Try to get anything close to the plants also.
b) Using low power, systematically examine all of the area under the cover slip.
c) When organisms are located, examine them in greater detail using high power.
d) If you want, try to identify your organism, there are some posters on the walls that
show pond water organisms.
Draw a SINGLE-CELLED ORGANISM from your pond water, use the space below:
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C. Estimating cell size
You will need to do all these steps for EACH cell type you are going to examine (Pages 80–
81: Cheek epithelium, Elodea, potato, red onion)
1. Estimate the size of a SINGLE cell. Make sure you can identify a single cell in the leaf,
onion, potato etc. since these cells are naturally packed together as tissues.
Ocular lens
10X
10X
10X
Objective lens
4X
10X
40X
Total magnification
40X
100X
400X
CONVERT mm measurements to µm. Remember:
Diameter of field of view
4mm
1.6mm
0.4mm
1mm = 1000µm
1µm = 0.001mm
You will need to have all cell sizes in µm on the sheets you turn in.
Practice:
this cell is observed using the 40X objective:
Estimate its size
2. Draw in PENCIL only. Pen is too messy when you are adding detail.
Diagram carefully. Each cell should have MORE detail than this:
i) Cheek Epithelium
a) Place a drop of water on a clean slide. Obtain the epithelial cells by gently rubbing
the inside of your cheek with the blunt end of a clean toothpick. Place the cells on
the slide by swirling the toothpick in the water. Now, add drop of methylene blue in
the middle, cover the slide with a cover slip.
Draw a SINGLE cell, use the space
Cheek
epithelium
SIZE:
Structures:
Cell membrane,
nucleus
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ii) Elodea leaf
a) Obtain a young (GREEN, NOT BROWN) leaf from the tip an Elodea plant. Put a
drop of water on a slide, place the leaf in the water, and cover with a cover slip.
b) Under low power observe that there are two layers of cells: one composed of large,
thick-walled cells, the second composed of smaller, thin-walled cells.
c) Under high power examine cells of the layer of small, thin-walled cells. Notice the
green chloroplasts. As the slide warms up, it may be possible to see the
chloroplasts move around the cell. This movement is called cyclosis or
cytoplasmic streaming. Notice, too, the large central vacuole in each cell. It is the
area of the cell in which there are no chloroplasts
Elodea
Draw a SINGLE cell, use the space
SIZE:
Structures:
Cell wall, cell
membrane,
chloroplast
central vacuole
(for TWO of these
you have to think
and visualize)
REVIEW: Now, put a couple of drops of NaCl on the Elodea, wait a couple of minutes,
draw what a cell looks like now:
Draw a SINGLE cell, use the space
Elodea – with NaCl
What is the environment
compared to the cell?
What happened?
Put your answers here:
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iii) Potato Tuber
a) With a razor blade or scalpel obtain a very thin slice of a white potato tuber. It
should be less than 1 mm thick. Put a large drop of water on a slide, place the
potato sample in the water, and cover with a cover slip. If the cover slip can “seesaw” back and forth over the potato, the potato slice is too thick.
b) Examine the slide under low and high power. Observe the obvious cell walls and
large oval bodies within the cells.
c) Put a drop of iodine (IKI) at the edge of the cover slip and allow the iodine to diffuse
under the cover slip.
d) Re-examine the cells. What color have the oval granules become? Based on their
reaction to iodine, what must the granules contain? These organelles called
leucoplasts.
Draw a SINGLE cell, use the space:
Potato
SIZE:
Structures:
Cell wall,
cell
membrane,
leukoplasts
iv) Red Onion
a) Put a large drop of water on a slide. Peel back a tiny section of a scale of THE RED
PART OF THE ONION, NOT WHITE PART. Place the specimen on a slide, and
cover with a cover slip.
b) Examine the colored cells under low and high power. Notice the central vacuole. It
is obvious because it contains the red/purple pigment.
c) Is the nucleus evident? If not, add a drop of iodine to the edge of the cover slip and
allow the iodine to diffuse under the cover slip. Reexamine the cells under high
power.
Draw a SINGLE cell, use the space:
Red onion
SIZE:
Structures:
Cell wall, cell
membrane,
nucleus
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The Dissecting Microscope
Dissecting microscopes are also used in laboratories to magnify specimens for examination.
However, these microscopes have several key differences compared to the compound
microscope that you have used so far.
Examine a dissecting microscope and add the following nine labels to the appropriate lines
on the picture below:
Carrying handle
Upper light source
Objective lens
Magnification control knob
Focusing knob
Lower light source
Ocular lenses
Light control
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Use of the dissecting scope
Use the dissecting scope to look at a range of objects. Try looking at money (bills and
coins), jewelry, your fingers, leaves or material you collect outside, or any specimens that are
available in the lab this week. As you use the scope, experiment with the magnification
control and the light sources to get different views of the objects.
Complete this chart comparing the dissecting scope to the compound microscope: put as
much information in the boxes to describe their differences as you can.
Compound microscope
Dissecting microscope
Light direction
(Where does it come
from/how does it
illuminate the
specimen?)
Thickness or
characteristics of
specimens that can
be viewed
Number of Objective
lenses
Range of
magnification.
In steps or
continuous?
What moves when
you focus an image?
What happens to the
image if you look at it
as you move the
specimen to the left?
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Brain Warm Up: Calorimetry & thermodynamics
INTRODUCTION
A.
Answer the following True/False questions – we will go over them in class.
a.
The piece of paper this page is printed on contains Ethermal and Echemical
b.
During the process of photosynthesis Elight is converted to Echemical
c.
A molecule of glucose contains Echemical
d.
An exergonic reaction results in products that have more Echemical than the
reactants.
e.
Energy can flow (be transferred) from one object to the next.
f.
One (science) calorie is equal to the amount of energy to raise 1gram of water 5oC
B.
The flame is going to burn up the paper - draw what the paper will look like afterwards.
Why does the paper look different?
Did a chemical reaction occur?
Was energy transferred?
Is there any energy in the paper before? What about afterwards? Explain
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Food Calorimetry: Measuring the energy in Food
In the following lab you are going to determine how much energy (or calories) is stored in the
chemical bonds in various food samples. You are going to do this measuring how much
energy is released when the chemical bonds in the food are broken.
Procedure:
Carry out the following procedure using a banana chip as the “food sample.”
1.
2.
3.
4.
Pour 10 mL of water into a test tube.
Mount the test tube in a test tube clamp and place a thermometer in the test tube.
Measure the initial temperature of the water and record it in the table below.
Zero the balance. Measure the mass of the food item on a small square of aluminum foil,
and record the value in the table below (be as precise as possible). Try to obtain a
sample such that its mass (along with the foil square) is between 1.0 and 1.5 grams.
Don’t forget to zero the balance BEFORE you put on the aluminum square – you want to
include the aluminum square in your weight.
5. Hold the food with a pair of metal forceps.
6. Ignite the food by placing it in the flame of a match or Bunsen burner. (Be sure the flame
is not near the test tube of water).
7. As soon as possible after it ignites, place the burning food beneath the test tube. The
flame should touch the tube (and will likely cause the tube to blacken). Continue to hold
the burning food under the test tube, repositioning it as necessary to keep the flame
directly under the tube, until the flame goes out. If the temperature should reach 80°C,
extinguish the flame prematurely.
8. After the temperature of the water stops rising, record its final temperature in the table
below.
9. Scrape any food residue from the forceps onto your square of aluminum foil. Measure
the final mass of the food residue+foil and record it in the table below (be as precise as
possible).
10. Calculate the change in temperature and the mass of food burned by subtracting the
initial from the final values.
Food Item
Final
Temp (°C)
(Tf)
Initial
Temp (°C)
(Ti)
Change in
Temp (°C)
(Tf -Ti)
Initial
Mass (Mi)
(grams)
Final Mass
(Mf)
(grams)
Mass of
food
burned (Mi
–Mf)
(grams)
Banana chip
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Answer the following:
1.
Where is the energy coming from to raise the temperature of the water?
2.
So, the increase of temperature of the water is actually from energy stored where?
3.
Draw a diagram showing the answers from 1 and 2. That is, sketch a cartoon showing
the flow of energy as it is transformed and transferred during this experiment.
4.
On your diagram label all the types of energy – that is, where are these types of energy
shown in your diagram? Add arrows between the energy types to show where the
conversions happen. LABEL THEM CLEARLY – even explain!
5.
Why was it important to keep the match or Bunsen burner away from the tube of water?
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Calculating Calories:
We can use this technique to compare the amount of energy in different foods. Calories are
a measure of energy. A calorie is defined as the amount of energy to raise the temperature
of 1 gram of water by 1°C.
1.
To calculate the energy (calories) released by combusting a food item, multiply the mass
of the water used (1 mL = 1 g of water) by the change in temperature. This gives you
the number of calories that were transferred to the water from the burning food sample.
Change in temperature x Grams of water used = Number of calories released
x
2.
3.
=
In order to be able to fairly compare the calories from one sample to another, we need to
adjust this value for the amount of food burned. Determine the amount of food actually
burned by subtracting the mass of the banana chip “after” from the mass of the banana
chip “before.”
Mass of food before — Mass of food after
=
–
=
Mass of food burned
Take the calories calculated in step #1 and divide by the change in mass calculated in
step #2. This gives you the number of calories per gram of banana chip.
Number of calories released /Mass of food burned (g) = calories (c) per gram of food
/
4.
=
Dietary calories (C) are actually kilocalories. To relate the caloric value you just
calculated to dietary Calories (C), divide your calories (c) per gram of banana chip by
1000.
/ 1000
calories (c) per gram of food
/
= Calories (C) per gram of food
1000 =
Record this value in the table on the next page.
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We are going to use this technique to compare the energy found in the following foods:
•
•
•
•
•
•
Banana chip
corn chip
pecan or walnut
Honeycomb™ cereal
oyster cracker
a wooden stick (toothpicks or a wooden applicator) - ok, not a food.
Before conducting this experiment, predict how the foods will compare in their stored
Echemical. List them in the order you think from greatest to least amount of energy.
GREATEST ENERGY
1.
LEAST ENERGY
2.
3.
4.
5.
Explain your reasoning.
Measure the energy found in the foods listed in the table below by using the procedure you
used previously. Use a clean test tube for each measurement.
Calculate the Calories per gram by following the steps on page 90.
Food Item
Final
Temp
(°C) (Tf)
Initial
Temp
(°C) (Ti)
Change in
Temp
(Tf -Ti)
Initial
Mass (Mi)
(grams)
Final
Mass (Mf)
(grams)
Mass of
food
burned
(Mi –Mf)
(grams)
Calories
(C) per
gram
Banana chip
Corn Chip
Pecan or
Walnut
Honeycomb
cereal
Cracker
Stick
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Answer the following:
1.
How do the foods compare in their Caloric content? Based on your data, rank them from
highest to lowest Calories per gram. How do your results compare to your predictions?
MOST CALORIES
1.
2.
LEAST CALORIES
2.
3.
4.
5.
What might explain any differences observed in the Caloric content of the different
foods? That is, why did the honeycomb cereal have a different amount of Calories/gram
than the pecan? Think about this at a molecular level!
Comparing Results to Nutritional Information
Let’s compare the energy content you have measured with the caloric values listed in the
nutritional information for these foods. We could find this data on the packaging or in a
nutritional database. We are going to use a nutritional database available on the internet.
You will need to go to the following web page:
http://www.nat.uiuc.edu/mainnat.html
1.
Click on NATS version 2.0. Select the age and gender of someone in your group from
the pull-down menu in Step 1.
2.
Type the name of the first food in Step 2 and click on the “Add food” button. The
database will list all the foods with that name in its descriptor. Select the one from the
list that best describes the food you used in the experiment, and click on “Add selected
food button.” (The pecans/walnuts used were dried, the crackers were “oyster”).
3.
Select “Gram” from the pull down menu of serving sizes and type “1” in the box for the
number of servings. Click on the “Add this amount” button.
4.
You should see the food listed on the “Personal Diet List.” Fill in your next food in the
space above the list and repeat steps 2-4 until you have listed all of the foods tested.
*Also look up Snickers – you will need this info later
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5. Once you have listed all of your foods, click on the “Analyze foods” button. This will
give you the nutritional analysis of all of the foods combined (as if you had made a recipe
using one gram of each of these items. Yuck). Scroll down and click on the “Display
nutrients for individual foods” button. This will display a table with the nutritional
breakdown of each of the foods listed separately.
Complete the following table to compare the experimentally determined Caloric values and
the nutritional values.
Food Item
Calories/gram
(Experimental)
(from page 91)
Calories/gram
(Nutritional)
(from database)
Banana Chip
Corn Chip
Pecan or
Walnut
Honeycomb
Cereal
Cracker
1.
How do these five foods compare in their Caloric content according to the nutritional
information from the database? Rank them from highest to lowest.
MOST CALORIES
1.
2.
LEAST CALORIES
3.
4.
5.
2.
How does this ranking compare to what you determined experimentally?
3.
Consider the highest Calorie foods tested versus the lowest Calorie foods tested. What
do you think might explain the difference in their Caloric content? Do you have any
evidence to support your idea?
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5.
Would you classify the combustion of the food items as an exergonic or endergonic
reaction? What is your evidence?
6.
Draw an energy diagram for the combustion of food. Do the products have more or less
energy than the reactants?
7.
How does this experiment demonstrate the “First Law of Thermodynamics?”
8.
How does this experiment demonstrate the “Second Law of Thermodynamics?”
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Today most Caloric determination of foods is analyzed by the quantity of each of the types of
molecules that can provide nutritional Calories (that is, the amount of carbohydrates, fats
and protein) and these values are multiplied by the corresponding number of Calories per
gram for these types of molecules.
Proteins and carbohydrates have approximately 4 Calories per gram, and fats have
approximately 9 Calories per gram.
9.
For example, 1 gram of a Snickers candy bar contains 0.1 g of protein, 0.22 g of fat and
0.60 grams of carbohydrate. Based on the Caloric information for proteins, fats and
carbohydrates listed above, how many Calories would you expect 1 g of Snickers to
contain? (Show your work)
10. You looked up the Caloric content of 1 g of Snickers candy bar earlier. How does this
value compare with the Caloric information calculated?
11. One Snickers bar has a mass of 61 grams. How many Calories would a Snickers bar
contain? (Show your work)
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“Burning Food” in Biological Systems
In this experiment, you have oxidized the food and converted its stored energy to thermal
energy by actually burning it. In the human body, food is enzymatically oxidized (you have
probably heard of people refer to us “burning our food”) by a process called cellular
respiration. In this process, the food is oxidized more slowly, and while some of the energy
is released as heat, some of it is transferred in coupled reactions to other energy-storing
molecules such as the molecule of ATP.
It is estimated that, at maximum, cellular respiration transfers about 37% of the energy of
food to ATP and the remainder is lost as heat. The ATP produced by cellular respiration
provides the necessary energy for other endergonic reactions in the organism.
1.
Suppose that cellular respiration were much more efficient. Supposed that 75% of the
calories in food were transferred to ATP. If your cells’ need for ATP remained the same,
how would this change your daily caloric requirements?
2.
Suppose that cellular respiration only transferred 5% of the energy to ATP. If your cells’
need for ATP remained the same, how would this change your daily caloric
requirements?
3.
Certain drugs and illnesses decrease the efficiency of cellular respiration. How would
these conditions affect the organism’s body temperature, assuming that the body still
required the same amount of ATP molecules to be made daily? How would these
conditions affect the organism’s use of food and stored energy reserves?
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Calorimetry of Wood
1.
Based on your results, what is the Caloric content per gram of wood?
2.
If there were a nutritional label on a package of wooden sticks (for example, popsicle
sticks, or toothpicks), it would read “Calories 0.” How might you explain the discrepancy
between the label and your results?
3.
The nutrition facts of All Bran is shown below. Calculate the number of Calories per
serving:
Nutrition Facts
Serving Size
1/2 cup (31 g; 1.1 oz)
Servings per package
About 17
Amount per
Serving
Total Fat ...................................................... 1 gram
Total Carbohydrate ................................ 24 grams
Fiber................................................... 11 grams
Sugars ................................................. 6 grams
Other carbohydrates ............................ 7 grams
Protein ....................................................... 4 grams
The Calories per serving listed on the box are 80. How does this compare with your
calculation?
How can you explain any difference observed?
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Brain Warm Up: Enzymes
INTRODUCTION
A. Answer the following True/False questions – we will go over them in class.
a.
Enzymes are made of carbohydrates.
b.
One enzyme can work in all kinds of environmental conditions, provided its
substrate is present
c.
Enzymes bind to specific substrates.
d.
At the end of a reaction, the reactants are released by the enzyme.
e.
Enzymes slow down chemical reactions.
f.
If an enzyme does not have its correct shape, it cannot perform the reaction
needed.
B. Use the drawing below to answer the questions:
a. What does each shape above represent?
b. What does the arrow represent?
c. The above picture is a representation of a reaction at the
level
a) atomic
b) molecular
c) cellular
d) organism
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Enzyme Lab Problem: Chemical Production
You work for Bimbco, an international corporation. Chemical prospectors working for Bimbco
in the rainforests of South America recently discovered a bright yellow colored fruit the locals
say has medicinal properties. The scientists noticed that the fruit is often eaten by Jaguars
and further research has shown that jaguars that eat this fruit are immune to feline leukemia
virus. Back in Bimbco Labs, research has shown that it is the yellow colored chemical in the
skins of the fruit that is protecting the cats from FeLV, and that this chemical is produced in
the fruit by an enzyme that converts a chemical called ONPG that naturally occurs in the fruit
into the yellow colored chemical, which they have called O-nitrophenol. The scientists at
Bimbco Labs have purified the enzyme needed to produce the O-nitrophenol and, luckily, you
can buy ONPG in bulk from a chemical supplier. So, now you are ready to use the purified
enzyme to create large quantities of O-nitrophenol from ONPG in an industrial size
bioreactor.
The reaction that is catalyzed by the plant enzyme can be written as follows:
ONPG
Purified plant enzyme
O-nitrophenol + galactose
(Yellow colored)
You have carefully considered many options for the location of your production plant
(including labor costs, tax breaks and business incentives, and the infrastructure in the area)
and you have narrowed your choice down to three possible locations:
1) Hibbing, Northern Minnesota (the land is cheap and government is enticing new
businesses with many incentives)
The average temperature is 4°C
2) Orlando, Florida (there is a skilled workforce and lots of local support for the
bioindustry)
The climate is warm and the average temperature is 37°C
3) Yuma, Arizona (the land is also cheap here, and there is easy access to all the
infrastructure in California)
It’s very hot and temperatures inside the reactor could reach 60°C for much of the
year.
In addition, to save costs, the management of Bimbco have suggested that you set up the
reactors to do the enzymatic reaction (the conversion of ONPG to
O-nitrophenol) in milk instead of water. This way you will save money on a preparation step
for the product and you’ll be able to sell the milk with added
O-nitrophenol directly to millions of cat-loving Americans as “Anti-leukemia Milk” that they will
feed their cats daily, in place of regular milk.
One pair at your table will investigate the appropriate location for the production plant
(directions start on page 104), while the other pair will test the viability of doing the reaction in
a milk based solution (directions start on page 106). Do the experiments as outlined below,
then apply your results to decide what you will recommend to the management of
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Bimbco regarding the location of the new plant and whether they should set up their reactors
to contain milk.
Testing the effect of temperature on Enzyme activity:
Three water baths will be used for this experiment. Two are set up for you on the side bench
- one is set at 37°C (Florida conditions) and one is set at 60 °C (Yuma conditions). You need
to set up a 4°C (Minnesota) waterbath yourself, by filling a beaker ¾ full with ice, then adding
water to make an icy slush in which you will stand your tube.
You are going to set up 4 reactions. Three of them will be identical –that is they will contain
identical ingredients, but they will be incubated at three different temperatures 4°C, 37°C and
60°C. The fourth tube will be incubated at 37°C but will not contain any enzyme.
When you set up the 3 identical reactions, you are actually going to pre-incubate the
ingredients in separate tubes to make sure everything is already at the correct temperature
before you mix them and the reaction starts.
1.
Obtain 8 regular test tubes. These are the large test tubes at your bench. DO NOT
use the smaller tubes that have a white vertical line close to the top for this part of the
experiment – these are for the spectrophotometer.
2.
Label the tubes 1a, 1b, 2a, 2b etc.. Although you are setting up 4 reactions you will
need 8 tubes because you will be incubating the ingredients for each reaction separately until
they reach the desired temperature.
3.
Using a graduated cylinder, fill the tubes as shown in the table below. Remember to
rinse the cylinder well when you switch from measuring one ingredient to another.
Reaction 1
Reaction 2
Reaction 3
Reaction 4
(4°C)
(37°C)
(60°C)
(37°C, no enzyme)
tube a
2 mL ONPG
2 mL ONPG
2 mL ONPG
2 mL ONPG
tube b
2mL purified plant enzyme
2mL purified plant enzyme
2mL purified plant enzyme
2mL Water
4.
Cover all of the tubes with parafilm and place both the tubes for reaction 1 in the 4°C
waterbath, both the tubes for reaction 2 in the 37°C water bath, both the tubes for
reaction 3 in the 60°C water bath and both the tubes for reaction 4 in the 37°C
waterbath. DO NOT MIX THE CONTENTS OF THE TUBES YET!
5.
Allow the contents of all the tubes to equilibrate to the temperatures of their respective
waterbaths for 10 minutes. Do NOT cut the time short. Keep an eye on the ice water
bath and add extra ice periodically to keep it slushy.
6.
After the 10 minute period is up, combine the contents of the pairs of tubes by pouring
the ONPG into the purified enzyme solution. Basically you are combining tube 1a and
1b, 2a and 2b etc. Cover each tube with parafilm, mix well and immediately RETURN
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THE TUBES TO THEIR WATERBATHS. Note the time at which the enzymes and
substrates (tubes a and b) were mixed.
7.
Allow the tubes to incubate in the waterbaths for 30 minutes. Remember to keep
adding ice as necessary to keep the 4°C bath cold. At the end of the 30 minutes note
the color of the solution in each tube. Record the intensity of yellow in table 1.
8.
Carefully pour the contents of each tube into a Spec 20 tube - these are the smaller
tubes with a vertical white line near their top. Do not label the Spec-20 tubes
themselves, but label their positions in the rack.
9.
Using the control tube to zero the spectrophotometer, follow the directions on page 121
to obtain the Absorbance at 420nm for each of the tubes.
10. Record the values in table 1.
11. Use the information at the top of page 108 to determine the µm of O-nitrophenol
produced per minute. Record the values in table 1.
12. Using the white board provided, make a rough line graph showing the relationship
between temperature and micromoles of O-nitrophenol produced per minute.
13. To test whether the inactivity of the enzymes at certain temperatures is reversible, put
the tubes that are not very yellow after 30 minutes incubation into the 37°C water bath.
Allow the tubes to sit for another 10 minutes and observe any color change. Record
your results in terms of the intensity of the yellow color in table 2. The Tubes will not be
reread using the Spectrophotometer.
14. Wash all the tubes (test tubes and Spec 20 tubes) and leave them inverted to drain in
the rack
15. Use the white board to draw diagrams to explain what is going on in your test tubes at a
molecular level at each of the three temperatures.
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Testing the effects of lactose on enzyme activity
1.
Obtain 4 regular test tubes. These are the large test tubes at your bench. DO NOT use
the smaller tubes that have a white vertical line close to the top for this part of the
experiment. Label the tubes A, B, C and D.
2.
Use a graduated cylinder to add 1.0 mL of ONPG into each of the four tubes.
3.
Rinse the cylinder well and then add buffer to the tubes as follows:
Tube A – 3.0 mL
Tube B – 2.0 mL
Tube C – 1.0 mL
Tube D – 2.0 mL
4.
Rinse the cylinder well and then add 10% lactose solution (milk sugar) to the tubes as
follows:
Tube A – 0.0 mL
Tube B – 1.0 mL
Tube C – 2.0 mL
Tube D – 2.0 mL
5.
Rinse the cylinder well and when you are prepared to start timing the reaction for 30
minutes, add 1.0mL of the purified plant enzyme solution to tubes A, B and C. Do not
add enzyme to Tube D. Tube D will be the control for this experiment.
Summary of tube contents:
Tube
A
B
C
D
ONPG
solution
1mL
1mL
1mL
1mL
Buffer
10% lactose
3.0mL
2.0mL
1.0mL
2.0mL
0mL
1.0mL
2.0mL
2.0mL
Purified plant
enzyme
1.0mL
1.0mL
1.0mL
0mL
Total
Volume
5.0mL
5.0mL
5.0mL
5.0mL
6.
Cover each tube with parafilm and shake to mix the contents.
7.
Incubate the tubes in a 37°C waterbath for 30 minutes
8.
After exactly 30 minutes remove the tubes and stop the reactions by adding 0.5mL of
1M Na2CO3 to each tube. (Na2CO3 is as base which will alter the pH of the solution
sufficiently to inactivate the enzyme and stop the reaction.)
9.
Note the intensity of yellow in Tubes A, B, C and D. Record your observations in Table
3.
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10. CAREFULLY pour the contents of the tubes into Spec20 tubes - these are the smaller
tubes with a vertical white line near their top. Do NOT label the Spec 20 tubes
themselves, but label their positions in the test tube rack.
11. Using the contents of Tube D to calibrate the spectrophotometer, follow the directions
on page 121 to determine the Absorbance at 420nm for each of the tubes. Ab420 will
be a measure of the concentration of O-nitrophenol molecules produced in each of the
tubes. Record the data in Table 3
12. Use the information at the top of page 109 to calculate the number of µM of Onitrophenol produced per minute. Record these values in Table 3.
13. Using the white board provided, make a rough line graph showing the relationship
between the amount of lactose present and micromoles of O-nitrophenol produced by
the enzyme per minute.
14. Use the white board and draw diagrams to explain at a molecular level what is going on
in each of your test tubes.
15. Wash all the tubes (test tubes and Spec 20 tubes) and leave them inverted to drain in
the rack
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Results – Temperature Experiment
Each of micromole (µM) of O-nitrophenol produces an absorbance of 0.004 at 420nm.
Divide the Ab420 reading you get for each sample by 0.004 to determine the number of
micromoles of O-nitrophenol produced in each experiment.
Table 1: Effect of temperature on enzyme activity
Tube
Temp.
°C
A
4°C
B
37°C
C
60°C
Intensity of
yellow after
30 minutes
Ab420
µM O-Nitrophenol
produced in 30
minutes
µM O-Nitrophenol
produced / minute
Table 2: Can altering incubation temperatures restore enzyme activity?
Tube
Initial
incubation
temperature °C
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Color after initial 30
minute incubation
Color after additional 10
minute incubation at 37°C
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Results – Lactose Experiment
Each micromole (µM) of O-nitrophenol produces an absorbance of 0.004 at 420nm. Divide
the Ab420 reading you get for each sample by 0.004 to determine the number of micromoles
of O-nitrophenol produced in each experiment.
Table 3: Effect of lactose on enzyme activity
Tube
Amount of
10%
lactose
added
Enzyme
present?
A
0 mL
YES
B
0.5 mL
YES
C
1.0 mL
YES
D
1.0 mL
NO
Intensity of
yellow after
30 mins
Ab420
µM O-Nitrophenol
produced in 30
minutes
µM ONitrophenol
produced /
minute
Mini Presentations
You are going to use your molecular diagrams and graph to explain your results to the other
pair at your table. You will do this in the form of a mini presentation that the other pair will
grade you on. The following page (page 110) gives you guidelines on what to include in
your presentation. Use pages 112 and 113 as appropriate to score the other pair, then, as a
group of four, complete the memo on page 114. As a group of four, you will staple together
your score sheets and memo and turn the packet in today.
In order to answer the questions that follow on pages 116 – 118 you will need to do
the mini-presentations with the other pair at your table. The questions will not be
graded, but these questions and more will be used on your lab quiz next week. MAKE
SURE YOU UNDERSTAND THE RESULTS OF TODAY’S EXPERIMENT AND CAN
ANSWER THESE QUESTIONS!!
***In addition, you have a Lab Report due next week***
For directions for the report see pages 119 and 120.
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Questions for the Temperature Team
These are the questions / material you need to answer and present to the other team:
1. What were the substrate(s) and product(s) of the reaction?
2. Show what happened in your experiment using your rough graph.
3. Explain what took place at the molecular level in each of your test tubes by
using your diagrams.
4. Explain why the reaction in the tube at 0oC would work if put at 37oC
5. Your recommendation to Bimbco management using your data to explain your
decision.
Questions for the Lactose Team
These are the questions / material you need to answer and present to the other team
1. What were the substrate(s) and product(s) of the reaction?
2. Show what happened in your experiment using your rough graph.
3. Explain what took place at the molecular level in each test tube by using your
diagrams.
4. Explain which type of inhibition this is (competitive vs noncompetitive), and what
the difference is between these two types of inhibition.
5. Your recommendation to Bimbco management using your data to explain your
decision.
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SCORING FOR TEMPERATURE TEAM
First, score each item as follows: 2 (taught very well, very clear).
1 (taught OK)
0 (did not teach)
Don’t coach the presenters, let them present.
When they are done, provide feedback: point out items that were not done or not very clear.
Also, tell them about what they did well.
Have them re-present and give them a second score on the items they needed to re-present.
Question or item
First score
After
remediation
1.What were the substrate(s) and product(s) of the
reaction?
2. You can easily tell what happened in their
experiment from their rough graph.
3. You can easily understand what took place at the
molecular level in each test tube by looking at their
diagrams.
4. They explained why the tube at 0°C would work if
put at 37°C
5. They defended their recommendation to Bimbco
management using their data to back up their ideas
Presenters:
Scorers:
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SCORING FOR THE LACTOSE TEAM
First, score each item as follows: 2 (taught very well, very clear).
1 (taught OK)
0 (did not teach)
Don’t coach the presenters, let them present.
When they are done, provide feedback: point out items that were not done or not very clear.
Also, tell them about what they did well.
Have them re-present and give them a second score on the items they needed to re-present.
Question or item
First score
After
remediation
1. What were the substrate(s) and product(s) of the
reaction?
2. You can easily tell what happened in their
experiment from their rough graph.
3. You can easily understand what took place at the
molecular level in each test tube by looking at their
diagrams.
4. They explained which type of inhibition this is
(competitive vs noncompetitive).
5. They defended their recommendation to Bimbco
management using their data to back up their ideas
Presenters:
Scorers:
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EACH GROUP OF 4: TURN IN TODAY BEFORE YOU LEAVE
What are your group’s recommendations to Bimbco management concerning the location of
the new industrial plant and whether they should set up their reactors to contain milk?
Include an explanation of your recommendations.
Memorandum
To:
Bimbco Management
Re:
Anti-Leukemia Milk
Production
From: (names of the 4 people at your table):
Date:
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These questions and more will be used on your quiz next week
1. Which compound was responsible for the yellow color? ________________________
2. Why did each tube gradually turn yellow, rather than an immediate color change?
_______________________________________________________________________
_______________________________________________________________________
3. At what temperature was the plant enzyme most active? _______________________
4a. What effect does low temperature have on enzyme activity?
________________________________________________________________________
4b. What is your evidence?
__________________________________________________________________________
______________________________________________________________________
4c. Why does low temperature have this effect?
__________________________________________________________________________
______________________________________________________________________
4d. Is this a temporary or permanent effect?_____________________________________
4e. What is your evidence?
__________________________________________________________________________
______________________________________________________________________
5a. What effect does increasing temperature up to 37°C have on enzyme activity?
________________________________________________________________________
5b. What is your evidence?
__________________________________________________________________________
______________________________________________________________________
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5c. Why does increasing temperature have this effect?
__________________________________________________________________________
______________________________________________________________________
6a. What effect does extremely high temperature have on enzyme activity?
________________________________________________________________________
6b. What is your evidence?
__________________________________________________________________________
______________________________________________________________________
6c. Explain at a molecular level how high temperatures have this effect
__________________________________________________________________________
______________________________________________________________________
6d. Is this a temporary or permanent effect? _____________________________________
6e. What is your evidence?
__________________________________________________________________________
______________________________________________________________________
7. From your data, what is the optimal temperature for this enzyme’s activity?
________________________________________________________________________
8. In which concentration of lactose was the most O-nitrophenol produced?
________________________________________________________________________
9. In which concentration of lactose was the least O-nitrophenol produced?
________________________________________________________________________
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10. What effect did lactose have on the production of O-nitrophenol?
__________________________________________________________________________
______________________________________________________________________
11. How can you explain the effect of the lactose?
__________________________________________________________________________
______________________________________________________________________
12. What were the controls for these experiments?
________________________________________________________________________
________________________________________________________________________
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Enzyme Experiment Lab Report
(40 Points)
Your report must be double spaced, in 12 point font, with normal 1 inch margins. Remember,
it’s not the length that counts but the quality. IT MUST BE NEAT.
Your lab report must have the following:
1) AN INTRODUCTION (20 points).
The introduction should be 1 – 2 pages long and about enzymes and the reaction you
did in your experiments. DO NOT INCLUDE THE PROCEDURE – do not tell me the
procedure you did in lab.
Things to Include:
a) What are enzymes? What are they made of?
• What are the monomers that make enzymes AND HOW ARE THEY
IMPORTANT TO THE FOLLOWING:
• the shape or conformation of an enzyme – explain levels of conformation
• does every enzyme have the same shape? WHY OR WHY NOT?
• the ability to bind a specific substrate – HOW does an enzyme do this?
• the ability to carry out a reaction – HOW does an enzyme do this?
b) Why do cells need enzymes?
c) The reaction in this experiment:
• what is the major reaction you studied in this lab,
• what are the substrates and products,
• why is this reaction used in this lab? That is, what made it easy to follow the
progress of this particular reaction? How did we measure the progress of the
reaction?
2) EXPERIMENTAL RESULTS (20 points). Depending on which experiment you did:
TEMPERATURE GROUP
a) A diagram showing what happened AT THE MOLECULAR LEVEL during the
reaction
b) A discussion of your results containing NOT ONE number, that is, you need to tell
me what happened, don’t just put your data table into sentence form. This will
probably be only a couple of sentences.
c) GRAPH of results: incubation temperature vs micromoles of ONP produced/min.
You should know by now the expectations for a graph. There is a sheet of graph
paper on page 122
OR
Turn over…
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LACTOSE / ENZYME INHIBITION GROUP
a) A diagram showing what happened AT THE MOLECULAR LEVEL during the
reaction
b) A discussion of your results containing NOT ONE number, that is, you need to tell
me what happened, don’t just put your data table into sentence form. This will
probably be only a couple of sentences.
c) GRAPH of results: quantity of lactose vs micromoles of ONP produced/min.
You should know by now the expectations for a graph. There is a sheet of graph
paper on page 122
The answers to the questions on pages 116 – 118 will help you with your introduction
and discussion
DO NOT PUT IN DATA IN YOUR DISCUSSIONS, NOR
THE PROCEDURES USED FOR THE LABS
NUMBERS
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How to use a Spec-20
1. Turn on the Spec20 and wait for 15 minutes.
2. Set the Wavelength using the dial.
3. Insert the Spec tube containing your control reaction into the machine, making sure
the tube is properly aligned. The white stripe on the Spec tube should be aligned with
the ridge on the Spec.
4. Close the lid.
5. Calibrate the Spec20 using the dial:
Set %Transmittance to 100 and Absorbance to 0.00
(use the switch next to the readout to toggle between Transmittance and Absorbance).
6. Remove the control and insert your sample with the white trip on the Spec tube
properly aligned.
7. Close the lid.
8. Read and record the %Transmittance or Absorbance of your sample.
9. When you change the wavelength setting of the Spec20 you must repeat steps 2
through 6.
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Brain Warm Up: Photosynthesis and Cellular Respiration
INTRODUCTION
A. Answer the following True/False questions – we will go over them in class.
a.
The goal of respiration is to build a molecule of glucose
b.
Plants do cellular respiration
c.
The goal of photosynthesis is to build a molecule of glucose
d.
ATP Synthase is a protein molecule
e.
ATP is a protein molecule
f.
A Hydrogen ion gradient is used to power ATP synthase to make ATP, and this
occurs in both photosynthesis and cellular respiration.
B. Use the equations below to answer the questions:
A)
6CO2 + 6H2O
C6H12O6 + 6O2
B)
C6H12O6 + 6O2
6CO2 + 6H2O
a. Circle the product(s) in the first reaction
b. Circle the reactant(s) in the second reaction
c. If reaction B was slowed down or stopped would CO2 levels increase or decrease?
d. Which reaction summarizes cellular respiration? How do you know?
e. Which reaction summarizes photosynthesis? How do you know?
f. The production of ATP is the overall goal of which of these processes, A or B?
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Photosynthesis and Cellular Respiration Review Lab
Part I: Vernier Lab Pro
You will be collecting data on variations in the amount of carbon dioxide gas in a sealed
chamber containing spinach leaves. You will be measuring the amount of carbon dioxide gas
in the chamber and tracking the changes in real time.
Procedure: The equipment MAY be set up already for you OR, do the following:
1. Turn on the computer. For the Mac Laptops the username is “Student” and there is no
password.
2. Start the program “Logger Pro” (Remember: the icons to launch the programs are
hidden at the bottom of the screen).
3. Once you’re in the logger pro program, choose:
file>open>biology with computers>experiment 31B
A window will open asking you to check the correct sensors are plugged in.
4. Plug the blue plastic data collector (the data logger Lab Pro) into a power outlet
5. Hook the data logger up to the computer using the USB cable
6. Plug the CO2 sensor into Channel 1 on the left side of the data logger
7. Get 5 – 10 DRY spinach leaves and carefully put them in the clear plastic bottle.
8. Connect all the pieces as shown in the diagram below.
10. Now, on the computer screen you should see a blank graph and a set of readings for
the level of CO2 in the chamber displayed at the lower left of the screen.
11. After you place the spinach leaves in the bottle, cover the bottle to block light from the
spinach leaves and let it sit for 5 mins.
CO2 sensor
Carefully place
5 - 10 DRY
spinach leaves
in the bottle.
Lab Pro
Channel 1
USB
connection to
computer
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Cable to
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While you are waiting for your set up to equilibrate:
1a. In which organelle(s) does photosynthesis take place?
1b. In which organelle(s) does respiration take place?
1c. Which of these organelles do plant cells contain?
1d. Which of these reactions are the plant cells in the dark chamber currently doing?
6CO2 + 6H2O
C6H12O6 + 6O2
C6H12O6 + 6O2
6CO2 + 6H2O
1e. Sketch the graphs for how you think the amounts of O2 and CO2 will change in the
chamber while the leaves remain sealed in there in the dark. Don’t worry about exact
numbers just sketch the trends. The lines have been started for you...
Amount
of O2
Amount
of CO2
Time
Time
Data collection:
1. After your setup has equilibrated for 5 minutes, you are ready to start collecting data.
Press “Collect Data” (the green button above the top graph) to have the computer start
collecting data. The computer will automatically collect readings from the probe every 5
seconds and start to plot graphs of the CO2 concentration in the chamber.
2. Collect data for 10 minutes. You may need to adjust the Y axis in order to more easily see
the data being collected on the graph.
3. Once 10 minutes have passed, uncover the set-up and turn on the light. Make sure the
spinach leaves are well illuminated and collect data for a further 10 minutes. You may have
to restart the data collection by hitting the green button above the graph, and then choose
the option “Append to Latest” to continue the line on the graph.
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1f.
EMCC Spring 2011
Which of these reactions are the plant cells in the illuminated chamber doing?
6CO2 + 6H2O
C6H12O6 + 6O2
C6H12O6 + 6O2
6CO2 + 6H2O
1g. How will this effect your graphs. That is, what will happen to the amounts of CO2 and O2
in the chamber once the light is turned on?
1h. Look at the graph of results on the computer - has CO2 been added or removed from
the chamber since the light was turned on?
1j
Is this what you expected? Why/Why not?
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Brain Warm Up: DNA Structure
INTRODUCTION
A. Answer the following True/False questions – we will go over them in class.
a.
DNA is made of proteins and carbohydrates.
b.
DNA is the molecule that chromosomes are made of.
c.
DNA contains information to build proteins.
d.
RNA and DNA are the exact same kind of molecules. There really is no reason
why they have two different names.
e.
Enzymes synthesize DNA and RNA.
f.
Nucleotides have 4 parts or components.
B. Use the drawing below to answer the questions
What is the name of this shape?
What do the lines inside represent?
(just hydrogen bonds is NOT a good answer)
How many base pairs can you see?
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DNA and RNA Structure
Part I: DNA Structure
FIG 1
Here is a picture of DNA - you can see how stringy it
is. Get a computer and go to the following site:
http://www.pbs.org/wgbh/nova/genome/dna.html#
Click on “Journey into DNA” – flash version
Click on the positive magnifying lens to answer the following questions:
1.
How many cells does this animation state you have in your body?
2.
Explain what it means when cells need to differentiate or specialize as your body
develops from a single fertilized egg cell.
3.
What do you think the term “human genome” means?
4.
How many chromosomes are in each cell of your body?
5.
What is the percentage of your genome that DOES code for proteins?
6. The number of genes is incorrect in the animation. The latest data indicates that humans
have about half of this number. So, what is the latest estimate as to how many genes we
have?
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7. What do banding patterns on chromosomes indicate?
8. What is the range of the number of bases for a gene?
9. How many feet of DNA are in the nucleus of each cell in your body?
Yuck!! “FEET”??? This is supposed to be a science website!!
10. Use your answers from #1 and #9 to determine the total length of DNA in your entire
body. YOUR FINAL ANSWER NEEDS TO BE IN METERS: (1 meter = 3.3 feet)
FYI: 1 mile = 1,609 meters
The length you just calculated would stretch to the moon and back about 6 million times!
10. The term chromatin actually refers to DNA + proteins – what do they say these proteins
do?
11. Histones have a positive charge. DNA has a negative charge. For each of these
molecules – which parts of the molecules would give them these charges? Be specific.
12. What is taking place between the bases on the DNA molecule that you cannot see?
13. The base on the last slide is a
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PYRIMIDINE
or
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PURINE
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14. OK, now does the picture at the beginning make sense? Why does DNA look so
stringy?
15. Figure 2 is showing a molecule of RNA –
What is different between this and DNA?
•
FIG 2
•
•
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16. Open up the DNA puzzle kit and set to the side all of the brown and tan pieces, you will
not be using these. For the rest of the pieces, identify the color of the following:
ribose
adenine
deoxyribose
guanine
cytosine
thymine
uracil
phosphate
17. Put a deoxyribose in front of you. It should look like the figure on the left:
O
H
It is necessary for you to learn about the
numbering of the carbons in the deoxyribose.
CH2
Use the figure at the right as a key and
number the carbons on the figure at the left.
That is, you are going to identify all 5 carbon
atoms in the deoxyribose from the puzzle.
The numbers are actually written as primes:
1’, 2’, 3’, 4’, 5’
18. Now, use the pieces to make a nucleotide of DNA using the base adenine – make sure
the phosphate is on the 5’ carbon.
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Draw the nucleotide exactly as you see it with the puzzle pieces; label the
carbons with their prime numbers.
19. On the 3’ carbon, there is a line indicating a bond to attach to the next nucleotide. If this
nucleotide was at the end of a molecule of DNA, it would not be a bond but a functional
group which is a _______________ group.
Use figure 3 to figure this out.
FIG 3
20. Using figure 3, build this piece of DNA with your puzzle kit. DO IT THE CORRECT
WAY: PUT TOGETHER YOUR 8 NUCLEOTIDES FIRST, THEN HOOK THEM
TOGETHER. Then, label all of the 5’ and 3’ carbons on both strands of Fig 3, don’t
worry about 1’, 2’ and 4’ carbons.
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Keep your puzzle out and go to the following site on your computer. Go to:
http://www.johnkyrk.com/DNAanatomy.html
Click on the green triangle to continue.
21. Where do phosphodiester bonds form, that is between which functional groups on
adjacent nucleotides do these bonds form? How many of these bonds do you see in
Figure 3?
22. Click to the next animation. What are they showing you here?
23. Click through the next couple of frames and finish the animation series. You will
probably need to step back and forth to answer the last question:
They show a huge mistake on how DNA is put together. You actually did it correctly
using the puzzle – what is the problem? Concentrate on how the DNA strand is
assembled…
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Brain Warm Up: DNA replication
INTRODUCTION
A. Answer the following True/False questions – we will go over them in class.
a.
DNA polymerase III synthesizes a new DNA strand in a 5’ – 3’ direction
b.
Helicase unwinds DNA to get it ready for replication.
c.
Primase always makes primers first before DNA polymerase III can start to work.
d.
Primers are made of RNA and are made in a 5’ to 3’ direction.
e.
DNA polymerase I seals two new DNA fragments together.
f.
Single strands of DNA are held apart by SSBPs.
B. Use the picture below to answer the questions
a.
Label the 5’ and 3’ ends of all the DNA strands shown.
b.
Put an X where the origin of replication is located.
c.
Which Okazaki fragment is the last one to have its primer replaced?
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DNA Replication - PUZZLE
Use the DNA puzzle to answer the questions.
1.
Open up the DNA puzzle kit and set to the side all of the brown and tan pieces, you will
not be using these again.
2.
Put together six individual DNA nucleotides. DO NOT JOIN THEM TOGETHER to
make a strand yet. Make two nucleotides with the base C, two with the base A and one
each using the bases G and T.
3.
Now hook these nucleotides together to make a single strand of DNA - You can make
any sequence (order of bases) you like. Make sure you have a 5’ phosphate end and
the appropriate 3’ hydroxyl end.
4.
Now, pretend you are primase and you are going to make a primer to this piece of DNA.
Q1.
Which end of the DNA template strand will you start to pair the RNA
nucleotides to? _____
5.
Now make six individual RNA nucleotides, AWAY from the DNA strand. DO NOT JOIN
THEM TOGETHER to make a strand yet. Make two nucleotides with the base G, two
with the base U and one each using the bases C and A.
6.
Now you are going to use some of these RNA nucleotides to build a primer for your
DNA strand. Starting at the correct end of the DNA strand (that you identified in #4.)
pick the correct RNA nucleotide, then move it in to the DNA strand. Repeat this
procedure moving along your strand of DNA as if you are primase until you have joined
3 RNA nucleotides together to make a short primer. SHOW ME WHEN YOU ARE
DONE.
7.
When you were moving along the strand of DNA, you had to move some proteins out of
the way (OK, you have to use your imagination!)
Q2.
What were these proteins called?
____
Q3.
Why were they in your way? That is, what were they doing? _______
______
_____________________________________ ___ _____ __________
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Draw your piece of DNA/RNA like the example below. The lines for the sugar
phosphate backbone have been drawn in for you already. You need to fill in the bases
with their hydrogen bonds AND you will need to somehow show there is a difference
between the DNA and RNA strands than just the base uracil (just think of a simple way
to do this) ALSO label the 3’ and 5’ ends of each strand.
A
U
C
G
Fill in your molecule
here:
Example molecule
9.
Now, you are going to be the enzyme DNA Polymerase III. Continue to elongate the
new strand by adding DNA nucleotides onto the 3’ end of the new strand. TO DO THIS
AS ACCURATELY AS POSSIBLE: FIRST MAKE A SET OF 3 DNA NUCLEOTIDES,
KEEP THEM SEPARATE. THEN, ADD THEM ONE BY ONE ONTO THE GROWING
STRAND. You can stop when all the bases on the template strand are paired up.
10. Now, you are a different enzyme that removes the RNA primer and replaces it with
complementary DNA. TO DO THIS AS ACCURATELY AS POSSIBLE, FIRST MAKE A
SET OF 3 DNA NUCLEOTIDES, KEEP THEM SEPARATE. THEN, REMOVE ONE
RNA NUCLEOTIDE AT THE CORRECT END AND REPLACE IT WITH THE
CORRECT DNA NUCLEOTIDE. MOVE ALONG THE STRAND ONE AT A TIME
Q4.
Which enzyme are you?
You actually need to have an additional protein to help seal the DNA together but the
puzzle doesn’t allow us to see this step separately.
Q5.
What is this protein’s name?
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11. Now draw your completed double-stranded DNA molecule like this:
A
T
C
G
Fill in your molecule
here:
Q6.
When you were working as enzymes with DNA and RNA, it was fairly easy for
you to put nucleotides in and out – WHY? What is it about DNA and RNA
nucleotides that makes them want to pair up/line up in this way? (Hint: in the
middle)
12. Now, close your books and notes:
Label everything on the diagram below, including the orientation of the strands.
See how much you can fill in without using the book or your notes.
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DNA replication - COMPUTER
1.
Get a computer and go to http://www.johnkyrk.com/DNAreplication.html
2.
Click one time.
3.
On the picture below, draw in helicase and SSBP’s. You must do it on both sides of the
bubble. In addition, draw a box around the origin of replication on EACH strand – not
like the dotted box you see on the screen.
Q1: Why does the origin of replication contain a lot of AT base pairs?
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4.
Click one more time, the animation goes fast.
5.
On the diagram above, put stripes on the tags showing the RNA primers.
Q2: Approximately how many nucleotides long are the primers?
6.
On the diagram, label the 5’ and 3’ ends on BOTH the old strands and the new
RNA/DNA strands.
Q3: The two strands that you can see being built in the snapshot shown above are:
a. both leading strands
b. both Okazaki fragments
c. a leading on the left, Okazaki on the right
d. an Okazaki on the left, leading on the right.
7.
Click again.
Q4: Actually, you know that the DNA polymerase that has exonuclease activity
(“exonuclease” means it is able to remove nucleotides from the chain) is called:
8.
Click again.
Q5: DNA replication occurs BEFORE which two types of cell division?
And
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The Cell Cycle
Name:
Each of the microscopes on the side bench are focused on onion root tip cells. The root tips
of plants are fast -growing, so cell division occurs often in this tissue type. These cells have
been stained to show the chromosomes, and each microscope is focused on a cell in a
different stage of mitosis.
Determine the order of the phases of the cell cycle that are presented in the microscopes:
Microscope #
Interphase
Prophase
Metaphase
Anaphase
Telophase
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Brain Warm Up: Gene Expression
INTRODUCTION
A. Answer the following True/False questions – we will go over them in class.
a.
Transcription takes place in the nucleus
b.
The enzyme used to do transcription is DNA polymerase III.
c.
Translation takes place on the RER or on cytoplasmic ribosomes.
d.
When a mutation occurs this means that the RNA sequence only is affected.
e.
tRNAs contain codons to anticodons.
f.
The process of making proteins using information encoded in DNA is called
transcription
B. Use the picture below to answer the questions
c.
a.
Which process is being shown?
b.
The codon on the mRNA for amino
acid threonine is ACC. What was the
original DNA sequence for this codon?
If the DNA sequence that corresponds to that codon was changed to TAG, what would
happen? That is, what would occur with the mRNA and what does that mean for the
tRNA trying to insert the amino acid threonine?
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Gene Expression – Team 1
You and your partner are going to make an interactive model of gene expression using the
materials provided in lab. You are going to make the model and then present it to a different
group at a different table. You are going to score each other’s model and turn the score
sheets in at the end of the lab. You can find the score sheet on page 151.
Here is your piece of DNA:
3’
T G C C T A C T A A C G G T C G T T T G T G A G A G A T C C T A T C G C T 5’
1.
First, underneath the sequence shown above, write the sequence of the piece of RNA
that would be transcribed from this DNA sequence.
2.
Underline the start codon, then for every three nucleotides draw a vertical line:
GCCCGA to show the reading frame for this gene.
3.
Now make a model based on the diagram handed out to you. You need to be able to
manipulate the model to show the following actions and detail to another group:
a) DNA in the nucleus opens up. You need to have the bases on the strand that you
use as the template to make mRNA visible – that is, the other group needs to see
T C G, etc.
b) Your mRNA is made from the open DNA – you can have the mRNA already made
since it is impossible to show single RNA nucleotides coming together. The mRNA
bases MUST BE LABELED: the A U G C’s need to be visible – even arrange them
in groups of three if possible.
c) The mRNA will already be partially processed – you don’t need to worry about
showing introns being removed – we will just imaging it happening!
d) Show a 5’ cap and 3’ poly A tail being added to your mRNA.
e) Show the mRNA leaving the nucleus – 5’ end comes out first.
f) Have the small ribosomal subunit bind first to the 5’ end.
g) Have the large ribosomal subunit bind next.
h) Position the mRNA to show the start codon and the appropriate tRNA. At this point
you need to make sure you have a pool of all the tRNA’s you are going to need.
AND your tRNA’s need to show the anticodon in addition to having the
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appropriately labeled amino acid at the end that can be detached and added to a
growing protein. In addition, each amino acid needs to be a different color.
i) Show the mRNA being translated: the correct tRNA comes in, the anticodon lines
up with the codon, the amino acid is dropped off and a peptide bond is formed
between the amino acids.
Do not worry about showing the EPA sites on the ribosome.
j) When you reach your stop codon, you should have a release factor made and
show how it binds to the stop codon.
k) Show how all of the molecules and the ribosome fall apart or disassemble after the
release factor interacts with the mRNA.
l) Your protein needs to be able to be folded – it will probably be impossible to show
the true secondary level (alpha helix and beta pleated sheet) but it needs to have
some sort of structure.
NOW, a mutation occurs!
Here is your mutated DNA - you need to figure out what type of mutation occurred.
3’
TGCCTACTAACGGTCGTTTGTGTAGAGATCCTATCGCT
5’
m) Show the group the mutation in your DNA – describe which type of mutation it is.
n) Have a second protein made to show the group what happens with your mutation –
the amino acid sequence may have changed, therefore, the shape may be
different. Clearly explain to the group all the possible impacts of this mutation on
this protein.
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Gene Expression – Team 2
You and your partner are going to make an interactive model of gene expression using the
materials provided in lab. You are going to make the model and then present it to a different
group at a different table. You are going to score each other’s model and turn the score
sheets in at the end of the lab. You can find the score sheet on page 151.
Here is your piece of DNA:
3’
T G C C T A C T A A C G G T C G T T T G T G A G A G A T C C T A T C G C T 5’
1.
First, underneath the sequence shown above, write the sequence of the piece of RNA
that would be transcribed from this DNA sequence.
2.
Underline the start codon, then for every three nucleotides draw a vertical line:
GCCCGA to show the reading frame for this gene.
3.
Now make a model based on the diagram handed out to you. You need to be able to
manipulate the model to show the following actions and detail to another group:
a) DNA in the nucleus opens up. You need to have the bases on the strand that you
use as the template to make mRNA visible – that is, the other group needs to see
T C G, etc.
b) Your mRNA is made from the open DNA – you can have the mRNA already made
since it is impossible to show single RNA nucleotides coming together. The mRNA
bases MUST BE LABELED: the A U G C’s need to be visible – even arrange them
in groups of three if possible.
c) The mRNA will already be partially processed – you don’t need to worry about
showing introns being removed – we will just imaging it happening!
d) Show a 5’ cap and 3’ poly A tail being added to your mRNA.
e) Show the mRNA leaving the nucleus – 5’ end comes out first.
f) Have the small ribosomal subunit bind first to the 5’ end.
g) Have the large ribosomal subunit bind next.
h) Position the mRNA to show the start codon and the appropriate tRNA. At this point
you need to make sure you have a pool of all the tRNA’s you are going to need.
AND your tRNA’s need to show the anticodon in addition to having the
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appropriately labeled amino acid at the end that can be detached and added to a
growing protein. In addition, each amino acid needs to be a different color.
i) Show the mRNA being translated: the correct tRNA comes in, the anticodon lines
up with the codon, the amino acid is dropped off and a peptide bond is formed
between the amino acids.
Do not worry about showing the EPA sites on the ribosome.
j) When you reach your stop codon, you should have a release factor made and
show how it binds to the stop codon.
k) Show how all of the molecules and the ribosome fall apart or disassemble after the
release factor interacts with the mRNA.
l) Your protein needs to be able to be folded – it will probably be impossible to show
the true secondary level (alpha helix and beta pleated sheet) but it needs to have
some sort of structure.
NOW, a mutation occurs!
Here is your mutated DNA - you need to figure out what type of mutation occurred.
3’
TGCCTACTAACGGTCGATTGTGAGAGATCCTATCGCT
5’
m) Show the group the mutation in your DNA – describe which type of mutation it is.
n) Have a second protein made to show the group what happens with your mutation –
the amino acid sequence may have changed, therefore, the shape may be
different. Clearly explain to the group all the possible impacts of this mutation on
this protein.
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Gene Expression Scoresheet
The other team is to present their gene expression model to you and then explain the impact
of the mutation they experienced. Score the team concerning their dynamic model AND their
presentation
Names of the group you are scoring:
Completely
done
Question or item
Partially
done
Not done
at all
Bases on the DNA clearly visible
Bases on the RNA clearly visible.
5’cap and poly A tail present on the mRNA.
RNA moves from nucleus to cytoplasm – 5’ end first
Small ribosomal subunit binds first, then the large
ribosomal subunit
Pool of tRNA’s present.
All tRNA’s have clearly visible anticodons.
All tRNA’s have appropriate amino acids and the amino
acids look different from each other.
The protein starts with the correct start codon, tRNA
and amino acid.
Clearly showed translation: the correct tRNA pairs with
the codon on the mRNA AND they indicate where a
peptide bond is formed..
Translation is taking place on the ribosome
A correct stop codon is used and a release factor is
clearly visible.
The protein is folded some how.
The mutation they have is appropriately explained, that
is, where did the mutation take place and what kind of
mutation is it?
The impact of the mutation is shown to you by a
different protein that was made using the mutated
gene.
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Recovering the Romanovs
1.
Name:
Go to http://www.dnai.org/ , then click on “Applications” in the left navigation bar on the
site. Select the tab labeled “Recovering the Romanovs” on the bottom of the screen.
Part 1
2.
Read through Part 1 “Romanov Family” until you get to the Pedigree of the Tsarina
(slide 5).
Q1: Not including Alexei, what was the average lifespan for the hemophiliac males?
3.
On the next panel are videoclips, you do not need to view them if you do not want to.
4.
Finish the rest of Part 1.
Q2: Who else was executed along with the royal Russian family?
Part 2
5.
Read through the part about Anna Anderson to be familiar with her story. You do not
need to do the activities in this section.
Part 3
6.
Start part 3 “Science Solves a mystery”
Q3: In which year did scientists investigate a burial site in Russia?
Q4: How many bodies were they expecting to find?
7.
You do not need to listen to the videoclips.
8.
Count the skeletons (slide 3)
Q5: How many are there?
9.
Analyze the skeletons (slides 4 and 5)
Q6: All of the skeletons were . . .
Q7: How did they determine which bones belonged to which person?
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10. Watch the nuclear DNA animation (slide 8).
Q8: Are the bases of the DNA labeled?
Q9: What is the only part of the DNA that is actually labeled?
11. Watch the mitochondrial DNA animation (slide 8). Mitochondria have their own separate
circular pieces of DNA. Scientists believe that mitochondria were originally bacteria (a
couple billion years ago) that ended up living inside larger cells. Bacteria are the same
size as our mitochondria and bacteria also have circular pieces of DNA.
12. Keep reading about mitochondrial DNA.
Q10: Why is it that your mitochondrial DNA only came from your mother?
13. Click on the Mitochondrial Inheritance animation (Slide 10)
Q11: As you notice, the red mitochondrion represents a sequence of DNA from the
mom’s mitochondrion. Why is it that the red mitochondrion is not carried down to the
right side of the pedigree?
14. Read the questions on slide 10 and then click to reveal the answers. The last answer
tells you how they can identify the Romanovs.
15. For the next two questions (slide 11), click on the Tsarina’s Pedigree tab. Remember,
this is following mitochondrial DNA, this is NOT the same thing as following the gene for
hemophilia.
Q12: How many generations are shown on this pedigree?
16. Hide the pedigree and click on the answers to the questions.
17. Read the next slide and continue to the next one, you don’t need to click on the server
link since the results are presented in slide 14. Read the answers to the questions.
18. Continue to the Tsar’s pedigree (slide 15).
Q13: How many generations are shown on this pedigree?
19. Proceed through the rest of the presentation to the end.
Q14: What happened to Alexi and Anastasia?
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Brain Warm Up: Meiosis
INTRODUCTION
A. Answer the following True/False questions – we will go over them in class.
During the production of sperm by meiosis, four haploid cells are produced from
one diploid cell
Meiosis occurs only in the gonads. It does not occur in any other tissue in the
human body
A cell with 2n=16 that goes through meiosis will produce two cells, each with four
chromosomes in them
a.
b.
c.
d.
Chromosomes are replicated before prophase I of meiosis begins
e.
When two gametes fuse, a zygote is produced
f.
Crossing over of homologous chromosomes occurs during prophase II
B. Below is a karyotype of a newly discovered type of deep sea octopus.
1 2 3 4
a)
b)
c)
d)
Draw an arrow pointing to a chromatid
Circle a replicated chromosome
Put a box around a homologous pair of chromosomes
Shade in two pieces of DNA that will have the same sequence
e) During Anaphase I what will be separated?
1 and 2
or
2 and 3
f) When will 3 and 4 be separated?
g) For this octopus, 2n=
h) How many chromosomes will be in one of this octopuses gametes?
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Meiosis - Reebops
1.
NAME:
Work in groups of four. Open the envelope containing the Reebop chromosomes. Two
of you will have the yellow (mom) set and two will have the blue (dad) set.
Each group of two will follow these instructions:
2.
Spread out your chromosomes and organize them.
Q1: As a reebop, what is your diploid number?
Q2: As a reebop, what is your haploid number?
Q3: What do the letters on your chromosomes represent?
Q4: Use the information from the “Genetic Database of Reebop Traits” to determine the
genotype and phenotype of your Reebop. Fill in the table below.
Genetic Database of Reebop Traits
AA = 2 antenna (pins)
Aa = 1 antenna
aa = no antenna
MM = 1 colored hump (any color small marshmallow)
Mm = 2 colored humps
mm = 3 colored humps
QQ = Red nose
Qq = orange nose
qq = yellow nose
DD or Dd = 3 body segments (in addition to the head)
dd = 2 body segments
Tt or TT = curly tail (pipe cleaners)
tt = straight tail
EE or Ee = 2 eyes (tacks)
ee = 3 eyes
LL or Ll = 4legs (push pins)
ll = 6 legs
My Reebop’s Genotype:
My Reebop’s Phenotype:
Antenna
Humps
Nose
Body Segments
Tail
Eyes
Legs
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Build your Reebop parent according to your phenotype using the marshmallows,etc.
Q5: Look at the other parent (your Reebop’s mate). Just look at the creature itself,
NOT the chromosomes the other pair has, and determine its genotype as much as
you can.
My Reebop’s mate’s Genotype:
Antenna
Humps
Nose
Body Segments
Tail
Eyes
Legs
4.
Now, back to your own reebop. You are now going to work only with the
chromosomes that carry the E, Q and L genes. Put all the other chromosomes back
in the envelope and pretend these six are the only chromosomes that Reebops have.
Q6: Draw your chromosomes as “sausages”, showing the alleles like this:
A
Q7: Now draw each chromosome as a replicated chromosome.
Make sure you show the alleles AND KEEP THE CHROMOSOMES THEIR
RESPECTIVE SIZES
A
A
Q8: Circle a single chromatid and label it.
Q9: Clearly show and label a pair of homologues.
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5.
You are going to draw the following phases of meiosis, showing what happens to the
three pairs of chromosomes you are focusing on. Make sure your drawing clearly
shows the different chromosomes and their alleles. It should be easy for me to follow
each chromosome/chromatid sequentially.
6.
Use the top circle on the next page to draw metaphase I. The dotted line shows the
metaphase plate, or equator of the cell
Q10: What separates in anaphase I?
7.
Draw metaphase II in the lower circles on the next page.
Q11: What separates in anaphase II?
8.
Draw your four gametes on page 158. At the end of meiosis a set of four haploid cells
(gametes) are produced. Choose one of your gametes and circle it. This will be your
egg cell or sperm cell that you will use in sexual reproduction.
9.
Get together with the other parent in your group and combine your chosen gametes to
simulate fertilization.
Q12: What did you just make when the sperm fertilized the egg? That is, what is the
name of this new cell you just created?
Q13: Is this new cell haploid or diploid?
Q14: What is the genotype of your offspring? (only the three chromosomes you used)
Q15: What is its phenotype?
Q16: Of the three characteristics you followed, which one(s) were just like mom? Dad?
Different?
Q17: Why are prophase I and metaphase I of meiosis important?
10. Disassemble your parent, put the tacks and pins into the beaker to soak. Throw away
the marshmallows and sticks.
11. Staple the drawings and papers of your group (four people) together.
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EMCC Spring 2011
Metaphase I
Metaphase II
TURN OVER
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EMCC Spring 2011
Four gametes produced:
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BIO181 Lab Manual
EMCC Spring 2011
Lab Final Study Guide
This is only a general list of topics we have covered and skills you should have practiced in
lab throughout BIO181. Any or all of these could be in the Lab Final.
•
General lab equipment (from Lab 1)
•
Substrates, products, what enzyme are made of and how they work. What effects
enzyme activity.
•
How to draw a graph (axes with reasonable and even scale, labels and units; a title,
the independent variable on the bottom axis, the dependant variable on the side axis)
•
Explaining or predicting the movement of water by osmosis. Using the correct terms
concerning tonicity for the environment or a cell.
•
Macromolecule tests - biuret for protein, benedicts for reducing sugars, IKI for starch.
What a positive and a negative test looks like for each.
•
Metric measurement conversions.
•
Estimating cell sizes using a microscope
•
Calculating magnifications
•
General features of animal and plant cells viewed using a compound microscope.
•
Identifying the stages of mitosis and meiosis using models of chromosomes.
•
Determining genotypes from phenotypes and vice versa
•
Determining flow of energy between objects. Identifying and describing the two laws of
thermodynamics.
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