Laboratory Resource Guide
to accompany
Essentials of Biology Laboratory Manual
Fourth Edition
Sylvia S. Mader
1
Scientific Method
Unit I The Cell
2
Measuring with Metric
3
Microscopy
4
Cell Structure and Function
5
Enzymes
6
Photosynthesis
Unit II Genetics
7
Cellular Reproduction
8
Sexual Reproduction
9
Patterns of Inheritance
10
DNA Biology and Technology
11
Genetic Counseling
Unit III Evolution and Diversity of Life
12
Evidences of Evolution
13
Microbiology
14
Plant Evolution
15
Plant Anatomy and Growth
16
Animal Evolution
Unit IV Animal Structure and Function
17
Basic Mammalian Anatomy I
18
Chemical Aspects of Digestion
19
Energy Requirements and Ideal Weight
20
Basic Mammalian Anatomy II
21
Nervous System and Senses
Unit V Ecology
22
Effects of Pollution on Ecosystems
1
Laboratory
1
Scientific Method
(LM pages 1–8)
Fourth Edition
This lab has been much improved in a number of ways. In Section 1.1, examples are
provided for each step of the scientific method and in Section 1.3 and 1.4, students use plus
and minus signs to hypothesize and then record a pillbug's reaction to test substances. Table
1.4 has an improved design for collecting class data.
New/Revised Figure. 1.1 Pillbugs on leaf
New/Revised Tables. 1.1 Pillbug Speed; 1.2 Hypotheses About Pillbug's Response to
Potential Foods; 1.3 Pillbug's Response to Potential Foods; 1.4 Pillbug's Response to
Potential Foods: Class Results
MATERIALS AND PREPARATIONS
Instructions are grouped by procedure. Some materials may be used in more than one
procedure.
Special Requirements
Living material. Live pillbugs, Armadillidium vulgare, for all sections of lab
Earthworm alternative. Refer to the section titled “Earthworm Alternative” at the end of
this laboratory if you wish to use earthworms instead of pillbugs.
Fresh material. Substances for instructor to feed pillbugs and substances for students to
test pillbug behavior are listed in Section 1.4.
1.2
Observing a Pillbug (LM pages 4–5)
_____ pillbugs, Armadillidium vulgare, live (Carolina 14-3082)
_____ pen, white (or correction fluid, white) or taped tags
_____ magnifying lenses or stereomicroscopes
_____ small glass or plastic dishes, such as disposable Petri dishes
_____ graduated cylinders or small beakers for observing pillbug movement
_____ rulers, metric, 30 cm plastic
_____ stopwatch
Pillbugs. If ordering, ask for 50 pillbugs for a class of 20 to 35 or more students. Order
pillbugs so that they arrive as close as possible to the date of use. Follow care and feeding
instructions provided with the pillbug order and/or see the following.
2
Collecting pillbugs (LM pages 1, 4–7). Pillbugs like moisture and avoid sunlight. They can
be found next to brick buildings along the grass line or next to sidewalks, or under logs and
planks of wood. They are attracted to wet grass covered with a cardboard box or plastic tarp.
Encourage students to collect their own pillbugs and give them lab participation points.
Collect pillbugs in the spring, summer, and fall as they are hard to find in the winter. After
collecting, pillbugs can be easily maintained in a terrarium to keep a fresh supply all year
long. They feed primarily on decaying organic matter; they like moisture and avoid sunlight.
They like carrots and cucumbers. Change the food daily to prevent mold growth.
Preparation for lab. Withdraw food 1–2 days prior to the experiment. Use white correction
fluid or tape tabs to number the pillbugs for identification.
1.4
Performing the Experiment and Coming to a Conclusion (LM pages 6–7)
_____ pillbugs, Armadillidium vulgare, live (Carolina 14-3082)
_____ small beakers, 35-mm film cans, watch glasses, or small Petri dishes for
distributing test substances
_____ Petri dishes, preferably 150 mm (or else 100 mm) for testing the pillbugs
_____ small plastic bottle for spritzing
_____ distilled water
_____ cotton balls
Suggested test substances:
_____ flour
_____ cornstarch or brand flakes
_____ coffee creamer
_____ baking soda
_____ fine sand (control)
_____ milk
_____ orange juice or apple juice
_____ ketchup
_____ applesauce
_____ carbonated beverage
_____ water (control)
Do not use salt, vinegar, or honey, as these substances are harmful to pillbugs.
Plain water is used as a control for liquids. Fine sand is used as a control for powders.
Experimental design (LM pages 6–7). These methods are recommended: For a dry
substance, make a circle of the test substance in a Petri dish and put the pillbug in the center
of the circle. For a liquid, put a cotton ball soaked with the test substance in the pillbug's
path. Rinse pillbugs between testing procedures by spritzing with distilled water and then
placing them on a paper towel to dry.
Suggestions. The experiment goes well and clean up is easier if there is a limited number of
test substances and each student chooses only two dry and two liquid test substances.
3
Substances can be distributed to several stations in small beakers, 35-mm film cans, watch
glasses, or small Petri dishes. Testing pillbugs in 150 mm Petri dishes works well.
EXERCISE QUESTIONS
1.1
Using the Scientific Method (LM pages 2–3)
Why does the scientific method begin with observations? To study the natural world,
scientists have to observe natural phenomena.
What is the benefit of formulating a hypothesis? The hypothesis tells what is to be tested
by experiment or further observations.
Why must a scientist keep a complete record of an experiment? So others can repeat the
experiment and can check that the data are valid.
What is the purpose of the conclusion? The conclusion tells whether the hypothesis was
supported or not.
How is a scientific theory different from a conclusion? Each experiment has a conclusion.
A scientific theory is based on many conclusions from various experiments in related fields.
1.2
Observing a Pillbug (LM pages 4–5)
Observation: Pillbug’s External Anatomy (LM page 4)
1.
 How can you recognize the head end of a pillbug? The head bears antennae and
eyes.
 How many segments and pair of walking legs are in the thorax? There are 7
segments and 7 pairs of legs.
Observation: Pillbug’s Motion (LM page 5)
1.
a. Describe the action of the feet and any other motion you see. The seven pairs of
legs move with the front pair leading, and each pair moves in succession thereafter.
b. Allow a pillbug to crawl on your hand. Describe how it feels. It tickles the skin as
it moves.
c. Does a pillbug have the ability to move directly forward? yes
d. Do you see evidence of mouthparts on the underside of the pillbug? A pillbug has
four pairs of mouthparts.
2. As you watch the pillbug, identify
a. the anatomical parts that allow a pillbug to identify and take in food. antennae,
eyes, and mouthparts
b. behaviors that will allow a pillbug to acquire food. For example, is the ability of
a pillbug to move directly forward a help in acquiring food. Explain. Yes,
because it is the most efficient way to reach food.
What other behaviors allow a pillbug to acquire food? A pillbug has appropriate
mouth parts for taking in and eating food.
c. a behavior that helps a pillbug avoid dangerous situations. The pillbug rolls into a
ball when it is threatened.
4
Table 1.1
Pillbug
1
2
3
Pillbug Speed*
Millimeters (mm) Traveled Time (sec) Speed (mm/sec)
71
30
2.36
122
60
2.20
64
30
2.12
Average speed: 2.23 mm/sec
*Answers will vary. The answers provided here are examples.
1.3
Formulating Hypotheses (LM page 6)
1-3. See Table 1.2 showing three possible student hypotheses regarding flour. Students uses
"0" for no response, "—" for moves away from the substance, and "+" for moving toward
the substance and eating it.
Table 1.2
Hypotheses about Pillbug’s Response to Potential Foods
Substance Hypothesis About
Reason for Hypothesis
Pillbug’s Response to Potential Foods
Flour
0
Flour is a bland substance.
Flour
—
Flour is a dry substance.
Flour
+
Flour is a food substance.
1.4
Performing the Experiment and Coming to a Conclusion (LM pages 6–7)
Experimental Procedure: Pillbug’s Response to Potential Foods (LM pages 6–7)
Table 1.3
Pillbug’s Response to Potential Foods
Substance
Pillbug’s Response Hypothesis supported?
Flour
+
Depends on hypotheses
Cornstarch
+
Coffee creamer
+
Baking soda
—
Fine sand
0*
Milk
+
Orange juice
—
Ketchup
—
Applesauce
+
Carbonated beverage +
Water
0*
*Pillbugs may move toward these substances but do not eat them.
5. Do your results support your hypotheses? depends on results
6. Are there any hypotheses that were not supported by the experimental results
(data)? How do such data give you more insight into pillbug behavior? Explain.
Answer depends on student hypotheses.
5
Table 1.4
Pillbug's Response to Potential Foods: Class Results
Answers will vary depending on class data.
8. On the basis of the class data do you need to revise your conclusion for any
particular pillbug response? depends on class data
Why is this the best methodology? The more trials, the more likely the results are valid.
9. Did the billbugs respond as expected to the controls, i.e., did not eat them. depends
on student results If they did not respond as expected, what can you conclude about
your experimental results? The results may be invalid.
LABORATORY REVIEW 1 (LM page 8)
1. What are the essential steps of the scientific method? making observations,
formulating a hypothesis, testing the hypothesis, coming to a conclusion
2. What is a hypothesis? tentative explanation of observed phenomena
3. Is it sufficient to do a single experiment to test a hypothesis? No, because reliability
increases the number of times the experiment is repeated and the results remain the
same.
4. What do you call a sample that goes through all the steps of an experiment and does
not contain the factor being tested? control
5. What part of a pillbug is for protection and what does a pillbug do to protect itself?
exoskeleton. A pillbug rolls into a ball to protect itself.
6. State the type of data you used to formulate your hypotheses regarding pillbug
reactions toward various substances. Observational data.
7. Why is it important to use one substance at a time when testing a pillbug’s reaction?
only then can you be certain of the pillbug’s reaction to that particular substance
Indicate whether statements 8 -10 are hypotheses, conclusions, or scientific theories:
8. The data show that vaccines protect people from disease. conclusion
9. All living things are made of cells. theory
10. The breastbone of a chicken is proportionately larger than that of any other bird.
hypothesis
Earthworm Alternative
Earthworms can be used instead of pillbugs for all of the exercises in this laboratory.
Place earthworms in large rectangular plastic storage containers and let them roam around for
approximately 15 min. These containers can also be used to keep earthworms between
6
experiments. Plexiglass is also needed to place test substances on while holding earthworms
above to see behavior towards substances.
Earthworms want to move rapidly to escape. They are inclined to move away from light,
move under things, and seem to want to move downward. They are expected to move away
from a heat source. They also move toward each other and pile up on each other. They can
move up and down on glass at a 45 degree angle.
With regard to what students already know about earthworm activity, they might predict
certain behaviors. Earthworms live (or hide) in the soil, so they would move down and
through soil. Soil prevents desiccation and keeps them cool and moist. By moving under
things, they could stay cooler, stay moist, and stay hidden in the dark. Perhaps light bothers
them also.
Earthworms can move backward and forward from both ends. When they are investigating a
substance, they make a long, skinny point out of the end they are investigating with, and if
they are repelled by a substance, they pull back and the end becomes thick and round.
When testing with liquids, if an earthworm gets even close to the substance, the substance
will be pulled along the earthworm’s body without the earthworm doing anything. Capillary
action or cohesion tension? To prevent this, hold the earthworm above the substance, in case
the substance (especially lemon juice) might harm the earthworm. Just let the worm move its
pointed end into or near the substance. You can tell when it is repelled as it will pull away.
Rinse the earthworm right away if it touches a substance (especially lemon juice).
WHEN FINISHED WITH EARTHWORMS, mix damp potting soil with some oatmeal,
potato peels, lettuce, or other organic matter from the test—not too much, just enough to give
the earthworms something to eat. Add earthworms. Cover container with newspaper. Keep
soil damp. When completely finished, release earthworms into garden or greenhouse soil.
7
Laboratory
2
Measuring With Metric
(LM pages 9-18)
Fourth Edition
This is a new lab that teaches the metric system.
MATERIALS AND PREPARATIONS
Instructions are grouped by exercise. Some materials may be used in more than one exercise.
2.1
Length (LM pages 10–12)
_____ meter stick, metric and English
_____ long bones from disarticulated human skeleton
_____ cardboard (10 cm  30 cm), two pieces
_____ rulers, plastic millimeter
2.2
Weight (LM pages 13–14)
_____ sturdy balance scale
_____ wooden block, small enough to hold in hand
_____ object, such as a piece of granite, or a trilobite fossil, small enough to fit
through the opening of a small graduated cylinder
_____ triple beam balance
_____ objects to weigh: penny, paper clip, quarter
2.3
Volume (LM pages 14–15)
_____ wooden block and object from above
_____ graduated cylinders, 50 ml or 100 ml
_____ test tubes (large enough to hold 20 ml of water)
_____ dropper bottles containing water
_____ index card, blank white (20 cm × 30 cm)
_____ beaker, 50 ml
_____ graduated pipette (for demonstration)
2.4
Temperature (LM page 16)
_____ thermometer, Celsius
_____ cold water, hot water, ice water
8
EXERCISE QUESTIONS
2.1
Length (LM pages 10–12)
You will want to know what these abbreviations stand for, so write them out here:
m = meter
µm = micrometer
cm = centimeter
nm = nanometer
mm = millimeter
How many cm are in a meter? 100
How many mm are in a centimeter? 10
How many µm are in a millimeter? 1,000 How many nm are in a micrometer? 1,000
Meter, Centimeter, and Millimeter (LM pages 10–11)
Observation: A Meterstick (LM pages 10–11)
2. How many centimeters are in a meter? 100 For example, The prefix centi- means
100. For example, how many cents are in a dollar? 100
3. How many millimeters are in a centimeter? 10 But the prefix milli- means a
thousand. How many millimeters are in a meter? 1,000 Obtain a penny and measure
its width in terms of mm. 18 mm Why does it seem preferable to measure a penny in
terms of millimeters? to use whole numbers
4. For example, if the bone measures from the 22 cm mark to the 50 cm mark, the
length of the bone is 28 cm. If the bone measures from the 22 cm marks to midway
between the 50 cm and 51 cm marks, its length is 27.5 cm = 285 mm.
5. Record the length of two bones. Recorded lengths will vary.
Millimeter, Micrometers, and Nanometer (LM pages 11–12)
Observation: Small Metric Ruler (LM page 12)
1. Use the ruler to measure the diameter of the circle shown to the nearest mm.
Diameter of circle in mm: 38 mm
2. Do you expect the answer to be a smaller number or larger number? larger
Size of the circle in µm: 38,000 µm
3. Diameter of circle in nm: 38,000,000 nm
4. You have shown that the diameter of the circle is 38 mm = 38,000 µm = 38,000,000
nm.
2.2
Weight (LM pages 13–14)
Using a revision of the formula on page 12 if necessary, do these conversions: 2g = 2,000
mg and 0.2 g = 200 mg.
Experimental Procedure: Weight (LM pages 13–14)
1. The weight of the wooden block is (Answers will vary.).
2. Measure the weight of an item small enough to fit inside the opening of a 50 ml
graduated cylinder. Answers will vary.
3. If so directed by your instructor, use a triple beam balance to take the weight of one
or more of these objects to a tenth of a gram:
Penny
2.5 g
Paper clip
1.4 g
Quarter
4.7 g
9
2.3
Volume (LM pages 14–15)
Experimental Procedure: Volume (LM pages 14–15)
1. For example, use a millimeter ruler to measure the wooden block used in the
previous Experimental Procedure to get its length, width, and depth. Answers will
vary according to the size of the block used. Computations of volume will also vary.
3. Hypothesize how you could find the total volume of the test tube. Fill the test tube
with water, and pour the water into the graduated cylinder. Read the volume in
milliliters.
What is the test tube’s total volume? Answers will vary.
4. Hypothesize how you could use this setup to calculate the volume of the small object
you weighed previously (see step 2, p. 13). Fill the cylinder with water to the 20 ml
mark. Drop the object into the cylinder, and read the new elevated volume. The
difference between the two readings is the volume of the object alone.
Now perform the operation you suggested. Answers will vary.
5. Hypothesize how you could determine how many drops from the pipette of the
dropper bottle equal 1 ml. Using a 10 ml graduated cylinder, count the number of drops
it takes to get to 1 ml.
How many drops from the pipette of the dropper bottle equal 1 ml? approximately
10 (Answers will vary with student’s technique and with the type of pipette.)
6. Are pipettes customarily used to measure large or small volumes? small
2.4
Temperature (LM page 16)
Experimental Procedure: Temperature (LM page 16)
1.
a. Water freezes at 32°F = 0°C.
b. Water boils at 212°F =100°C.
2. Human body temperature of 98°F is what temperature on the Celsius scale? 37°C
3. Record any two of the following temperatures in your lab environment. Answers will
vary.
2.5
Summary (LM page 17)
In Table 2.2. except for centimeter, the units are all 10X larger than the next unit. So,
when you convert a gram to milligram, you multiple the gram by 1000. To
determine what portion (milligrams to gram) you divide by 1000.
LABORATORY REVIEW 2 (LM pages 18)
1. What type of measurement is signified by kg? weight ml? volume cm? length
2.
3.
4.
5.
degrees? temperature μm? length
What type of measurement would utilize a meterstick? length a graduated
cylinder? volume a balance scale? weight
If a triple beam balance shows a weight of 100 g plus 10 g plus 1 g, what is the
weight of the object? 111 g
An object is added to a graduated cylinder that holds 250 ml, and the water
rises to 300 ml. What is the volume of the object in ml? 50 ml in cm3 ? 50 cm3
Name two units of measurement you expect to use in the next laboratory,
10
which concerns microscopy? micrometer (μm) and nanometer (nm).
6. How many micrometers are in a millimeter?
1,000
Convert 1.1 mm to μm. 1,100 μm
7. How many milliliters are in a liter?
1,000
Convert 500 ml to liters. 0.5 0 l
8. How many milligrams are in a gram?
Convert 5 g to mg.
1,000
5,000 mg
9. Convert 1.5 cm to μm. Show your work. 1.5 cm = 15 mm= 15,000 μm
10. A student looking for a shortcut drops an object in a graduated cylinder that
contained water to find its weight. What’s wrong?
and not weight. To measure weight use a scale.
This will measure volume
11
Laboratory
3
Microscopy
(LM pages 19-30)
Fourth Edition
This laboratory now contains only microscopic study as the metric system is now laboratory
2.
MATERIALS AND PREPARATIONS
Instructions are grouped by exercise. Some materials may be used in more than one exercise.
Special Requirements
Living material. Euglena. Order two weeks before laboratory.
Notes
Microscope supplies. Set aside an area in the lab for storage of clean microscope slides,
coverslips, and lens paper. Post a notice in this area, outlining the established procedures for
handling dirty slides. Possible procedures include:
1. Wash, rinse, and dry all slides, and return them to their boxes or place them in the drying
rack.
2. To wash, place dirty slides in the detergent solution provided; discard plastic coverslips.
Glass coverslips should be placed in detergent solution in a beaker.
3. Some laboratories prefer that the laboratory assistant wash all slides in an ultrasonic
cleaner, rinse the slides in distilled water, and allow the slides to drain dry.
3.2
Stereomicroscope (Binocular Dissecting Microscope) (LM pages 22–23)
_____ microscope, stereomicroscope with illuminator
_____ lens paper
_____ an assortment of objects for viewing (e.g., coins, plastomount)
3.3
Use of the Compound Light Microscope (LM pages 24–27)
_____ microscopes, compound light
_____ lens paper
_____ slide, prepared: letter e or newspaper, scissors, slides, and coverslips
_____ rulers, clear plastic millimeter
_____ slide, prepared: colored threads or to prepare your own, you will need slides
and coverslips, three or four colors of sewing thread (or hairs), scissors, and a
dropping bottle of water
3.4
Microscopic Observations (LM pages 27–29)
_____ microscope slides (glass or plastic)
_____ covers slips
12
_____
_____
_____
_____
_____
_____
_____
_____
_____
_____
_____
lens paper
microscopes, compound light
toothpicks, prepackaged flat
biohazard waste container for toothpicks
ethyl alcohol (ethanol), 70%; or alcohol swabs (if toothpicks are not
prepackaged)
Euglena
onion
optional prepared slide: human stratified squamous epithelium, cheek
methylene blue solution, or iodine-potassium-iodide (IKI) solution (premade))
dropping bottles, or bottles with droppers
Protoslo® or methyl cellulose solution
Methylene blue solution (LM page 28). Make up a 1.5% stock solution, using 1.5 g
methylene blue stain in 100 ml of 8.5% ethyl alcohol (ethanol, Carolina 86-1281). Dilute one
part stock solution with nine parts water for laboratory use, or use iodine (IKI) solution.
Methylene blue staining solution can also be purchased premade.
Iodine (IKI) solution (LM page 28). Iodine-potassium-iodide (IKI) solution can be
purchased premade, or the ingredients can be purchased separately as potassium iodide (KI)
and iodine (I). These dry ingredients have a long shelf life and can be mixed as needed,
according to the following recipe:
To make a liter of stock solution, add 20 g of potassium iodide (KI) to 1 liter of
distilled water, and stir to dissolve. Then add 4 g of iodine crystals, and stir on a stir plate;
dissolution will take a few hours or more. Keep the stock reagent in dark, stoppered bottles.
For student use, place in dropping bottles. Label as “iodine (IKI) solution.”
Iodine solution stored in clear bottles loses potency over time. If the solution lightens
significantly, replace it. Small dropper bottles can be stored for about a month, and they are
used in other exercises. A screw-capped, brown bottle of stock iodine can be stored for about
six months. Dispose of it if the solution turns light in color.
Human epithelium cheek slide (LM page 28). To eliminate the possibility of contact with
pathogens, this exercise can be done as a demonstration using a flexscope or videoscope for
students to view from their seats. Otherwise, because of the hazards connected with human
tissue samples and body fluids, you should take special precautions if students are preparing
their own epithelium slides. Use a biohazardous waste container for toothpick disposal, and
wash slides and coverslips in a 10% bleach solution. Microscopes should also be wiped with
a disinfecting solution.
Dropping bottles. Various styles of dropping bottles are available—for example, dropper
vials, glass screw-cap (Carolina 71-6438, -6434) with attached droppers; Barnes dropping
bottles (Carolina 71-6525); and plastic dropping bottles (Carolina 71-6550). See also
Carolina’s Laboratory Equipment and Supplies section.
13
Protoslo® (or methyl cellulose solution) (LM page 29). You can also use glycerol (Carolina
86-5530) and water as a substitute for Protoslo®. Note: Thickened Protoslo® can be
reconstituted with distilled water.
EXERCISE QUESTIONS
3.1
Light Microscopes Versus Electron Microscopes (LM pages 20-21)
Answer These Questions (LM page 21)
 Which two types of microscopes view the surface of an object? stereomicroscope
and scanning electron microscope
 Which two types of microscopes view objects that have been sliced and treated to
improve contrast? compound light microscope and transmission electron
microscope
 Of the microscopes just mentioned, which one resolves the greater amount of
detail? transmission electron microscope
3.2
Stereomicroscope (Binocular Dissecting Microscope) (LM pages 22–23)
Identifying the Parts (LM pages 22)
2. What is the magnification of your eyepieces? 10
5. Locate each of these parts on your stereomicroscope, and label them on Figure 3.3.
Figure 3.4 (left, top to bottom): eyepiece lenses, binocular head; (right, top to
bottom): magnification changing knob, illuminator, focusing knob
Focusing the Stereomicroscope (LM pages 23)
4. Does your microscope have an independent focusing eyepiece? yes, most likely Is the
image inverted? no
5. What kind of mechanism is on your microscope? Answers will vary.
6. and 7. The object will vary but only a portion of the object will be circled at the highest
magnification.
3.3
Use of the Compound Light Microscope (LM pages 24–29)
Identifying the Parts (LM pages 24–25)
Identify the following parts on your microscope, and label them in Figure 3.4.
Figure 3.4 (left, top to bottom): eyepiece(s) (ocular lens or lenses); body tube; nosepiece;
objective lens or lenses; stage or mechanical stage (optional); diaphragm/diaphragm control
lever; condenser
Figure 3.4 (right, top to bottom): arm; stage clips; coarse-adjustment knob; fineadjustment knob; light source; base
1. What is the magnifying power of the ocular lenses on your microscope? The
magnifying power of the ocular lens is marked on the lens barrel (usually 10).
5.
a. What is the magnifying power of the scanning lens on your microscope? usually
4
14
b. What is the magnifying power of the low-power objective lens on your
microscope? The magnifying power of the low-power objective lens is marked on the
lens barrel (usually 10).
c. What is the magnifying power of the high-power objective lens on your
microscope? The magnifying power of the high-power objective lens is marked on
the lens barrel (usually 40).
d. Does your microscope have an oil immersion objective? depends on microscope
14. Does your microscope have a mechanical stage? depends on microscope
Inversion (LM page 26)
Observation: Inversion (LM page 26)
1. Draw the letter e as it appears on the slide (with the unaided eye, not looking
through the eyepiece). The letter should be in the normal position.
2. Draw the letter e as it appears when you look through the eyepiece. The letter should
be upside down and reversed.
3. What differences do you notice? The letter is inverted—that is, it appears to be upside
down and backward compared to its appearance when viewed by the unaided eye.
4. Which way does the image appear to move? When moved to the right, the object
appears to move to the left.
5. Which way did the image move. opposite direction
Focusing the Microscope—Higher Powers (LM page 26)
4. On a drawing of the letter e, draw a circle around the portion of the letter that you
are now seeing with high-power magnification. Portion will vary but a smaller portion
is in view.
Total Magnification (LM page 27)
Observation: Total Magnification (LM page 27)
Table 3.2
Total Magnification*
Objective
Ocular Lens Objective Lens
Scanning power (if present) 10
4
Low power
10
10
High power
10
40
Oil immersion (if present)
10x
95-100x
Total Magnification
40
100
400
O
950-1000x
*Answers may vary with equipment.
3.4
Microscopic Observations (LM pages 27–30)
Human Epithelial Cells (LM page 28)
Observation: Human Epithelial Cells
3. Label Figure 3.6. 1. plasma membrane; 2. nucleus; 3. cytoplasm
Onion Epidermal Cells (LM page 28)
15
Observation: Onion Epidermal Cells
4. Label Figure 3.7. 1. nucleus; 2. cell wall
Table 3.3
Differences
Differences Between Human Epithelial and Onion Epidermal Cells
Shape
Orientation
Human Epithelial Cells
(Cheek)
Flattened, rounded
Random orientation
Boundary
Thin
Onion Epidermal Cells
Square or rectangular
Oriented end to end and in
lines/rows
Thick
Euglena (LM page 29)
Observation: Euglena (LM page 29)
5. How do your specimens compare with Figure 3.8? Answers will vary.
LABORATORY REVIEW 3 (LM page 30)
1. Of the three types of microscopes studied, which one best shows the surface of an
object? scanning electron microscope
2. Explain the designation “compound light” microscope:
a. compound There are two sets of lenses—objective and ocular.
b. light Light is used to view the object.
3. What function is performed by the diaphragm of a microscope? The diaphragm
regulates the amount of light for viewing the object.
4. Briefly describe the necessary steps for observing a slide at low-power under the
compound light microscope. Center the slide on the stage. Looking from the side, decrease
the distance between the slide and the objective lens until the lens comes to a stop. Looking
through the ocular lens(es), use the coarse-adjustment knob to increase the distance between
the slide and the lens until the object comes into view. Adjust the light, and fine-adjust the
focus.
5. Why is it helpful for a microscope to be parfocal? Little, if any, adjustment is needed
when switching from low to high power.
6. Why is locating an object more difficult if you start with the high-power objective
than with the low-power objective. The amount of the object you can see is smaller in high
power than in low power.
7. How much larger than normal does an object appear with a low-power objective
lens? 10X (ocular lens) X 10X objective lens = 100X
8. A virus is 50 nm in size. Would you recommend using a stereomicroscope, compound
light microscope, or an electron microscope to see it? electron microscope Why? Only
16
an electron microscope has the capability of observing an object this small because it
magnifies more and has greater resolving power.
9. What type of microscope, aside from the compound light microscope, might you use
to observe the organisms found in pond water? stereomicroscope
10. State two differences between onion epidermal cells and human epithelial cells.
Human epithelial cells are flat, round, and have a random orientation. Onion cells are
square and oriented end to end in rows.
17
Laboratory
4
Cell Structure and Function
(LM pages 31–44)
Fourth Edition
The prokaryotic cell was removed from this laboratory. Otherwise, this lab is essentially the
same as it was in the previous edition.
Special Requirements
Living material. Elodea, living, for Section 4.1and 4.3; whole sheep blood for Section 4.3
Fresh material. Potato for Section 4.3.
MATERIALS AND PREPARATIONS
Instructions are grouped by procedure. Some materials may be used in more than one
procedure.
4.1
Animal Cell and Plant Cell Structure (LM pages 32-35)
______ Elodea, living
______ forceps, dissecting fine point, stainless steel
______ dropping bottles
______ microscopes, compound light
______ lens paper
______ slides
______ coverslips
Elodea (LM page 35,40). Live Elodea can be purchased locally at aquarium stores or a
supply house. Place Elodea in distilled water in an aquarium with a continuous air supply
from an aquarium air stone and pump. Place in indirect window light or under artificial
illumination.
4.2
Diffusion (LM pages 36–37)
Diffusion Through a Semisolid (LM page 36)
______ petri dish
______ gelatin powder or agar powder for 1.5% solution
______ potassium permanganate (KMnO4) crystals
______ wax pencils
______ rulers, plastic millimeter (preferably transparent)
Diffusion demonstration through gelatin or agar (LM page 36). (Note: Agar allows faster
diffusion than gelatin.) Prepare one dish per student group. At least a day ahead, prepare a
18
1.5% gelatin solution in a beaker or flask by dissolving 1.5 g of gelatin powder or agar in 100
ml of boiling water; stir thoroughly until dissolved. Allow to cool until the glassware can be
handled with a hot mitt. Fill a petri dish 3 to 5 mm deep with gelatin solution. Put a lid on
dish until cool. After cooling, store the dish in a refrigerator. After gelling, make a small
depression in the center of the dish. Using forceps, drop a crystal of potassium permanganate
into the depression.
Potassium permanganate (LM page 36). Only 1 to 2 crystals are needed per student group.
While wearing gloves, dispense several crystals of potassium permanganate into a shallow,
wide-mouth, screw-top container appropriately labeled. (Note: Potassium permanganate
diffuses very quickly.)
Diffusion Across the Plasma Membrane (LM pages 36–37)
______ dialysis tubing, approximately 15 cm per setup
______ plastic droppers or Pasteur pipettes
______ rubber bands to close off the top of dialysis tubing
______ rubber bands that fit snugly around brim of 250 ml beaker
______ 1% glucose solution
______ 1–2% starch solution
______ beakers , 250 ml
______ water, distilled
______ iodine (IKI) solution
______ test tubes
______ test tube rack
______ wax pencils
______ Benedict’s reagent or glucose test strips, optional
______ boiling water bath:
______ hot plate (See Carolina’s Apparatus: Laboratory Equipment and
Supplies section.)
______ large beaker
______ pair beaker tongs
______ test tube clamps
______ boiling chips, pumice
______ thermometer, Celsius 50–150°C range (See Carolina’s Apparatus:
Thermometers section.)
1% glucose solution (LM page 37). This makes enough for all procedures for 20 student
groups. Place 1 g of glucose in 50 mL of distilled water. Stir to dissolve, and bring the
volume up to 100 mL.
1–2% starch solution (LM page 37). 20 mL per student group should be sufficient (using
standard test tubes for all procedures). Care must be taken in preparing this solution. To
make a 1% solution, dissolve 1 g of starch in a small amount of cold water to form a pourable
paste. Add this to 100 mL of boiling distilled water, while stirring, and
mix a few minutes. Cool. Add a pinch of sodium chloride (NaCl). If refrigerated, this
solution will last for several weeks; otherwise, a fresh supply should be prepared each day.
19
Iodine (IKI) solution (LM page 37). Use one dropper bottle per student group. Pre-made
iodine-potassium-iodide solution can be purchased, or the ingredients can be purchased
separately as potassium iodide (KI) and iodine (I). These dry ingredients have a long shelf
life
Benedict’s reagent (LM page 37). 50 mL per student group is sufficient. Benedict’s reagent
can be purchased as a powder to make 1 liter. Or to make 1 liter, mix 173 g of sodium citrate
and 100 g of sodium carbonate, anhydrous (Na2CO3) (Carolina 88-8770) with 800 mL of
distilled water. Warm this mixture to dissolve; then cool and filter it. Add distilled water to
make 850 mL. Then dissolve 17.3 g of copper sulfate (cupric sulfate, pentahydrate) in 100
mL of distilled water, and stir slowly into the first solution. Add distilled water to make 1
liter. When testing, Benedict’s reagent should be boiled approximately 5 minutes or longer.
Glucose test strips can be used in place of Benedict’s reagent to test for glucose in bag and
beaker.
Boiling water bath (LM page 37). Place a large beaker of water on a hot plate. Adjust the
dial on the hot plate so that the water is maintained at a gentle rolling boil during the
experiment. Thermometers are optional since students should know that boiling water is
100°C.
4.3
Osmosis: Diffusion of Water Across Plasma Membrane (LM pages 38–41)
Experimental Procedure: Osmosis (LM pages 38–39)
______ Osmosis Demonstration Unit
______ 50% corn syrup solution
______ plastic syringe for filling thistle tube
Osmosis Demonstration Alternative
______ dialysis tubing
______ beaker
______ 10–20% sucrose solution
______ rubber bands to close off the bottom of dialysis tubing
______ plastic clamps to close off the top of dialysis tubing
Osmosis demonstration (LM page 38). The Osmosis Demonstration Unit is particularly
easy to fill and empty. Partially fill the thistle tube with 50% corn syrup (or similar) solution.
Place the apparatus in a beaker containing distilled water. Other osmometers can be found in
Carolina’s Osmosis and Diffusion: Physiology section.
Osmosis demonstration alternative (LM page 38). This demonstration can also be done
using dialysis tubing and a beaker. See Experimental Procedure: Solute Diffusion Across the
Plasma Membrane for setup. Tie off one end of the tubing, then fill with
10–20% sucrose solution. Clamp or tie it off at the open end. Pat the bag dry and weigh.
Place the bag in a beaker of water for 45 minutes to 1 hour. Remove, pat dry, weigh
immediately.
Experimental Procedure: Demonstration of Tonicity in Red Blood Cells
(LM pages 39-40)
______ test tubes, Pyrex 16 mm X 150 mm with stoppers
______ stoppers, rubber laboratory, solid, size 1
20
______
______
______
______
______
______
sheep blood, pooled, citrated
water, distilled
0.8.% and 10% sodium chloride (NaCl) solutions
dropping bottles, or bottles with droppers
whole blood demonstration slides (optional)
microscopes, compound light
Whole blood (LM page 39). Blood should not be human blood. Use any available animal
blood, other than human, to remove the risk of transmission of the HIV virus. Use caution
with any animal blood as it may contain pathogens. Blood is shipped in iced, insulated
containers and should be stored in the refrigerator. If kept refrigerated, sheep blood may be
stored for up to 2 weeks.Prepare the test tubes as follows:
Tube 1:
5 ml 0.8.% NaCl plus three drops of sheep blood
Tube 2:
5 ml 10% NaCl plus three drops of sheep blood
Tube 3:
5 ml 0.8.% NaCl plus distilled water and three drops of sheep blood.
Stopper the tubes.
To prepare the NaCl solutions:
0.8.% NaCl: Add 8. g of NaCl to 1 liter of distilled water. Smaller volumes may be
prepared.
10% NaCl: Add 100 g of NaCl to 1 liter of distilled water. Smaller volumes may be
prepared.
Slides of whole blood (optional). Prepare a demonstration slide of the 0.8.% sheep blood
solution (Tube 1) and the 10% sheep blood solution (Tube 2) for student observation.
Experimental Procedure: Tonicity in Elodea Cells (LM pages 40)
______ See materials listed previously in Section 4.2.
______ 10% NaCl from the whole blood demonstrationExperimental Procedure:
Experimental Procedure: Tonicity in Potato Strips (LM page 41)
______ potato, fresh
______ rulers, plastic millimeter
______ razor blades, single-edged
______ wax pencils
______ cutting board for potato
______ 10% sodium chloride (NaCl) in wash bottles
______ test tubes and racks
______ water
______ paper towels
4.4
pH and Cells (LM pages 42–43)
Experimental Procedure: pH and Cells (LM page 42-43)
______ test tubes (3 per group)
______ test tube rack ______
pH 7 buffer (inorganic) solution
______ protein solution, buffered (e.g., albumin)
21
______
______
______
______
______
______
pH paper (range pH 0–14)
stirring rods, glass
0.1 N hydrochloric acid (HCl) (see Carolina Chemicals, Hydrochloric Acid)
11.1 M conc. in plastic-coated safety bottle
beakers, plastic 50 ml (two for each group)
droppers
water, distilled
pH 7 buffer (LM page 42). 50 ml per student group is sufficient. If you wish to make it yourself,
combine 50 ml 0.1 M potassium dihydrogen phosphate (KH2PO4), (1.36 g per 100 ml distilled
water) with 28.1 ml 0.1 M NaOH (0.4 g per 100 ml distilled water). Dilute this mixture to 100 ml
with distilled water.
Buffered “cytoplasm” (e.g., albumin solution) (LM page 42). 50 ml per student group
should be sufficient. Mix 1 g of albumin with 100 ml of pH 7.0 buffer. (Buffer may be
purchased.)
0.1 N HCl solution (LM page 42). Mix 0.83 ml concentrated HCl with 100 ml distilled
water. Place in dropper bottles.
Experimental Procedure: Effectiveness of Antacids (LM page 43)
______ mortar and pestle
______ antacids: Alka-Seltzer, Rolaids, Tums, or other antacid tablet
______ 0.04% phenol red solution
______ beakers, plastic 250 ml
______ 0.1 N hydrochloric acid (HCl) (see Carolina Chemicals, Hydrochloric Acid)
______ rods, glass stirring
______ dropper
EXERCISE QUESTIONS
4.1 Animal Cell and Plant Cell Structure (LM pages 32–43)
Study Table 4.1 to determine structures that are unique to plant cells and unique to
animal cells, and write them below the examples given:
Plant Cells
Animal Cells
1. Large central vacuole
1. Small vacuoles
2. Cell wall
2. Centriole
3. Chloroplast
Animal Cell Structure (LM page 33)
Label Figure 4.1 Answers follow. See Table 4.1 for a function of each labeled structure.
22
a. Nucleus
i. Mitochondria
b. Nuclear membrane
j. Ribosome
c. Nucleolus
k. Smooth ER
d. Cytoskeleton
l. Lysosome
e. Vesicle forming
m. Cytoplasm
f. Vesicle
n. Plasma membrane
g. Centrioles
o. Golgi apparatus
h. Rough ER
Plant Cell Structure (LM page 34)
Label Figure 4.2. See Table 4.1 for a function of these structures found in plant cells.
a. Chloroplast
h. Golgi apparatus
b. Nucleus
i. Mitochondrion
c. Nucleolus
j. Cytoplasm
d. Rough ER
k. Plasma membrane
e. Ribosome
l. Cell wall of adjacent cell
f. Smooth ER
m. Central vacuole
g. Cytoskeleton
n. Centrosome
Observation: Plant Cell Structure (LM page 35)
6. Can you locate the cell nucleus? Answers will vary, but usually yes.
7. Why can't you see the other organelles featured in Figure 4.2. A light microscope
does not permit seeing them; an electron microscope would be needed.
8. Can you detect the movement of chloroplasts in this cell or any other cell? Answers
will vary, but usually yes.
4.2 Diffusion (LM pages 36–37)
Experimental Procedure: Diffusion Through a Semisolid (LM page 36)
2. Length of time has been variable, dependent on when experiment began.
3. Measure (in mm) the movement of color from the center of the depression outward
in one direction: Answers will vary.
4. Calculate the speed of diffusion mm/60min. Answers will vary.
Calculate the speed of diffusion Answers will vary.
23
Table 4.2
Solute Diffusion Across Plasma Membrane
At Start of Experiment
Contents
Color
Bag
Glucose Starch
No color
At End of Experiment
Color
Benedict’s
Test
Blue-black
____
Beaker
Water Iodine
Yellowish
Less yellow
Positive (+)
Conclusion
Iodine
diffused into
bag.
Glucose
diffused into
bag.
Conclusions: Solute Diffusion Across the Plasma Membrane (LM page 37)
• Which solute did not diffuse across the dialysis membrane from the bag to the
beaker? starch How do you know. The solution in the beaker did not turn blue-black.
4.3 Osmosis: Diffusion of Water Across Plasma Membrane (LM pages 38-39)
Experimental Procedure: Osmosis (LM pages 38-39)
1. Note the level of liquid in the thistle tube, and measure how far it travels in 10
minutes: 1 mm
2. Calculate the speed of osmosis under these conditions: 6 mm/hr
Conclusions: Osmosis (LM page 38)
• In which direction was there a net movement of water? from beaker to thistle tube
Explain what is meant by “net movement” after examining the arrows in Figure
4.6b. Water moves in and out of thistle tube, but more water moves in than moves out of
tube.
• If the starch molecules in corn syrup moved from the thistle tube to the beaker,
would there have been a net movement of water into the thistle tube? no Why
wouldn’t large starch molecules be able to move across the membrane from the
thistle tube to the beaker? They are too large to cross a membrane.
• Explain why the water level in the thistle tube rose: In terms of solvent
concentration, water moved from the area of higher water concentration to the area
of lower water concentration across a differentially permeable membrane.
Tonicity in Cells (LM pages 39–40)
Animal Cells (Red Blood Cells) page 39
Experimental Procedure: Demonstration of Tonicity in Red Blood Cells (LM pages 39–40)
Table 4.3
Effect of Tonicity on Red Blood Cells
Tube Tonicity
Effect on Cells
Print Visibility
1
Isotonic
No effect
No
2
Hypertonic Cells lose water
No
3
Hypotonic
Cells gain water
Yes
Explanation
Cells are intact
Cells are intact
Cell have burst
24
Plant Cells (LM pages 40-41)
Experimental Procedure: Tonicity in Elodea Cells (LM page 40)
Table 4.4
Tonicity
Hypotonic
Hypertonic
Effect of Tonicity in Elodea Cells
Appearance of Cells
Normal
Shriveled center
Due to (scientific term)
Turgor pressure
Plasmolysis
Experimental Procedure: Tonicity in Potato Strips (LM page 41)
5. Which tube has the limp potato strip? tube 2 Use tonicity to explain why water
diffused out of the potato strip in this tube.? The solution in tube 2 was hypertonic.
Which tube has the stiff potato strip? tube 1 Use tonicity to explain why water
diffused into the potato strip in this tube? The solution in tube 1 was hypotonic.
6. Use this space to create a table to display your results.
Table
Effect of Tonicity on Potato Strip
Tube number Content
Tonicity
Results Explanation
1.
water
hypotonic
stiff potato strip
potato strip
2.
salt solution hypertonic
limp potato strip
potato strip
water diffused into
potato strip
water diffused out of
potato strip
Conclusion: Tonicity (page 41)
 In a hypotonic solution, animal cells swell to bursting. In red blood cells this is called
hemolysis. In a hypertonic solution, animals cells shrivel. In red blood cells this is
called crenation.
 In a hypotonic solution, the central vacole of Elodea cells exerts turgor pressure, and
chloroplasts are seen next to the cell wall. In a hypertonic solution, the central vacuole
loses water and plasmlysis occurs. The cytoplasm plus the chloroplast are seen in the
center of the cell.
 In a hypotonic solution, potato strips gain water; in a hypertonic solution, potato strips
lose water and become limp.
4.4 pH and Cells (LM pages 42–43)
Why are cells and organisms buffered? to maintain pH of the cells
Experimental Procedure: pH and Cells
Table 4.5
Tube
1
2
3
*
pH and Cells*
Contents
Water
Buffer
Cytoplasm
pH Before Acid
6–6.5
7
7
pH After Acid
2–3
7
7
Explanation
Not buffered
Buffered
Buffered
These results are based on 1 ml of test solution.
25
Conclusions: pH and Cells (LM page 43)
• Why would you expect cytoplasm to be as effective as the buffer in maintaining pH?
Living things are buffered.
Experimental Procedure: Effectiveness of Antacids (LM page 43)
Table 4.6
Effectiveness of Antacids
Data will depend on the antacids used.
Conclusions: Effectiveness of Antacids (LM page 43)
• Did dosage in mg have any effect on the results? depends on antacid used
• Which of the substances on the label could be a buffer? depends on antacid
LABORATORY REVIEW 4 (LM page 44)
1. What characteristics do all eukaryotic cells have in common? the presence of a nucleus
and membrane-bounded organelles
2. Which organelle digests macromolecules and plant parts? lysosome
3. Why would you predict that an animal cell, but not a plant cell, might burst when
placed in a hypotonic solution? Animal cells do not have cell walls.
4. Which of the cellular organelles would be included in the following categories:
a. Membranous canals and vacuoles endoplasmic reticulum, Golgi apparatus, vesicles,
vacuoles, lysosomes
b. Energy-related organelles mitochondria and chloroplasts
5. How do you distinguish between rough endoplasmic reticulum and smooth
endoplasmic reticulum?
a. Structure Rough endoplasmic reticulum has ribosomes; smooth endoplasmic reticulum
does not.
b. Function Rough endoplasmic reticulum is the site of protein synthesis; smooth
endoplasmic reticulum produces lipids.
6. If a dialysis bag filled with water is placed in a molasses solution, what do you predict
will happen to the weight of the bag over time? The bag will lose weight. Why? Water
would diffuse out of the bag and enter the molasses solution.
7. What is the relationship between plant cell structure and the ability of plants to stand
upright? Strong cell walls and water-filled vacuoles that maintain turgor pressure help
plants to stand upright.8. The police are trying to determine if material removed from
the scene of a crime was plant matter. What would you suggest they look for? To
determine if it was plant matter, the police should microscopically look for cell walls and
chloroplasts, and they should test for starch.
26
9. A test tube contains red blood cells and a salt solution. When the tube is held up to a
page, you cannot see the print. With reference to a concentration of 0.8.% sodium
chloride (NaCl), how concentrated is the salt solution? The solution has a lower
concentration than 0.9% NaCl since it is hypotonic to the cells and has caused them to burst.
10. Predict the microscopic appearance of cells in the leaf tissue of a wilted plant. The
vacuole has pulled away from the cell wall, and the chloroplasts have moved to the center of
the cell.
27
Laboratory
5
Enzymes
(LM pages 45–52)
Fourth Edition
New to this edition, each section asks students to hypothesize the outcome of the
experimental procedure before the experiment is done. In this way students can get a sense
of the scientific process.
MATERIALS AND PREPARATIONS
Instructions are grouped by procedure. Some materials may be used in more than one
procedure.
Special Requirements
Fresh material. A potato is needed per lab to prepare catalase for Sections 5.1-5.4.
Equipment. Incubator (or water bath) and refrigerator (or ice bath) for 5.2 Effect of
Temperature on Enzyme Activity. 15 minute incubation required.
All Exercises
_____ water, distilled
_____ test tubes and racks
_____ beakers
_____ graduated transfer pipets
5.1
Catalase Activity (LM Pages 46–47)
_____ catalase, buffered
_____ hydrogen peroxide, purchased locally
_____ 5% sucrose solution
_____ potassium phosphate, dibasic
_____ potassium phosphate, monobasic
_____ pre-mixed buffer, pH7
Order solutions/agents or prepare your own.
Buffered catalase (LM page 47). Make potato catalase fresh for each lab by grinding one
small potato or half of a large potato with 50 mL water in a blender. Strain the potato mixture
through a sieve to remove any large pieces of potato. Sections 5.1-5.3 requires buffered
catalase. Section 5.4 requires nonbuffered catalase. You may put the buffered/nonbuffered
catalase in a beaker on the supply bench, and students can use transfer pipettes to dispense
the enzyme into test tubes.
28
Phosphate buffer (LM page 47). Add 7.70 g potassium phosphate, dibasic, K2HPO4, and
6.80 g potassium phosphate monobasic, KH2PO4, to one liter distilled water. Mix, check pH,
and use to dilute catalase as needed. Premixed buffer may be used, as well.
Hydrogen peroxide (LM page 47). The hydrogen peroxide used in this experiment can be
purchased from a local store.
5% sucrose solution (LM page 47). Dissolve 5 g sucrose in 100 mL distilled water.
Dispense from a beaker with dropper pipettes.
Note: Caution the students that they should swirl the enzyme and substrate to mix, then allow
the tube to sit for 20 seconds before measuring the height of the bubble column. The bubbles
produced by the reaction are very small, and resemble shaving cream foam. If the
catalase/sucrose mixture is swirled for 20 seconds, the catalase will produce large bubbles,
which some students confuse for the enzyme reaction.
5.2
Effect of Temperature on Enzyme Activity (LM pages 47-48)
_____ catalase (see section 5.1)
_____ hydrogen peroxide (purchased locally)
_____ incubator
_____ refrigerator or ice bath
_____ boiling water bath:
_____ hot plate
_____ large beakers
_____ beaker tongs
_____ thermometer
_____ test tube holders
5.3
Effect of Concentration on Enzyme Activity (LM page 49)
_____ catalase (see section 5.1)
_____ hydrogen peroxide (purchased locally)
5.4
Effect of pH on Enzyme Activity (LM pages 50–51)
_____ catalase, nonbuffered (see section 5.1)
_____ 5 M HCl
_____ hydrogen peroxide (purchased locally)
_____ 5 M NaOH
Order solutions/agents or prepare your own.
5 M HCl. CAUTION—This solution will get HOT (LM page 50). Add 400 mL distilled
water to a 1-liter graduated beaker. Place beaker with magnetic spinbar on a stirring plate.
While stirring, slowly pour in 416 mL concentrated HCl. Add distilled water to bring the
volume up to 1,000 mL.
5 M NaOH. CAUTION—This solution will get very HOT (LM page 50). In a 1-liter
beaker with a magnetic spinbar, gradually add a total of 200 grams NaOH pellets to
750 mL distilled water, allowing the heat to dissipate between additions of NaOH. After the
solution cools, add distilled water to bring the volume up to 1,000 mL.
29
EXERCISE QUESTIONS
5.1 Catalase Activity (LM pages 46-47)
What is the reactant in this reaction? H2O2 What is the substrate for catalase? H2O2
What are the products in this reaction? H2O and O2 Bubbling occurs as the reaction
proceeds. Why? O2 production
Hypothesize which tube 1, 2, or 3 in Table 5.1 will have the greater bubble column
height. Include an explanation in your hypothesis. Tube 1 because only this tube contains
both catalase and its substrate.
Experimental Procedure: Catalase Activity (LM page 47)
Table 5.1 Catalase Activity
Tube Contents
Bubble Column Height
1
Catalase
20 mm
Hydrogen peroxide
2
Water
0 mm
Hydrogen peroxide
3
Catalase
0 mm
Sucrose solution
Explanation
Substrate and enzyme are
both present.
Tube lacks enzyme.
Tube lacks correct substrate.
Conclusions: Catalase Activity (LM page 47)
•
Which tube showed the bubbling you expected? tube 1.
•
Which tube is a negative control? tube 2 If this tube showed bubbling, what
could you conclude about your procedure? Results are not due to catalase; therefore,
experiment is invalid.
•
Enzymes are specific. Which tube exemplifies this characteristic of an enzyme?
tube 3.
5.2 Effect of Temperature on Enzyme Activity (LM pages 47-48)
With this information in mind, examine Table 5.2 and hypothesize which tube (1, 2, or
3) will have more product per unit time as judged by bubble height. tube 2 Include a
complete explanation in your hypothesis. The enzyme exposed to normal body temperature
will perform best because the molecules will be moving about but the temperature is not high
enough to denature the enzyme.
Experimental Procedure: Effect of Temperature (LM pages 48)
5. Record your results in Table 5.2. Plot your results in Figure 5.3.
Table 5.2 Effect of Temperature
Tube
Temperature (°C)
1 Refrigerator
2 Incubator
5°C
37°C
Bubble Column
Height (mm)
8. mm
23 mm
3 Boiling water
100°C
0 mm
Explanation
Temperature too hot.
Denaturation occurred.
Temperature below optimum.
Optimum temperature
Conclusions: Effect of Temperature (LM page 48)
30
•
Was your hypothesis supported. Depends on hypothesis.
•
What is your conclusion concerning the effect of temperature on enzyme
activity? A warm temperature speeds an enzymatic reaction, but a hot temperature
denatures an enzyme.
5.3 Effect of Concentration on Enzyme Activity (LM page 49)
With this in mind, examine Table 5.3 and hypothesize which tube (1, 2, or 3) will have
more product per unit time as judged by bubble column height. tube 3 Explain your
answer. The more enzyme molecules, the more active sites per substrate, and the more
product within a limited time frame.
Experimental Procedure: Effect of Enzyme Concentration (LM page 49)
Table 5.3 Effect of Enzyme Concentration
Tube
Amount of
Bubble Column
Explanation
Enzyme
Height (mm)
1
none
20 mm
Explanation for all tubes: The
2
1 cm
30 mm
greater the enzyme concentration,
3
3 cm
40 mm
the more O2 during the allotted time
Conclusions: Effect of Concentration (LM page 49)
•
Was your hypothesis supported. Depends on hypothesis.
•
If unlimited time was allotted, would the final results be the same in all tubes?
Yes. Explain why or why not. All tubes have the same amount of substrate and enzymes can
be used over and over again.
•
Would you expect similar results if the substrate concentration were varied in
the same manner as the enzyme concentration? yes Why or why not? It would take less
time for the substrate to encounter an active site.
•
What is your conclusion concerning the effect of concentration on enzyme
activity? Increased amount of enzyme or substrate will increase the rate of enzyme activity.
5.4 Effect of pH on Enzyme Activity (LM page 50-51)
With this information about catalase in mind, examine Table 5.4 and hypothesize which
tube (1, 2, or 3) will have more product per unit time as judged by bubble height. tube 2
Include a complete explanation in your hypothesis. pH 7 is optimum pH for catalase
Experimental Procedure: Effect of pH
1-3. Measure the height of the bubble column and record your results in Table 5.4. Plot
your results from Table 5.4 here (Fig. 5.4).
Table 5.4 Effect of pH
Tube pH
Bubble Column Height (mm)
1
3
17 mm
2
7
35 mm
3
11
12 mm
Explanation
pH too acidic for catalase
optimum pH for catalase
pH too basic for catalase
Conclusions: Effect of pH (LM page 51)
31
•
Was your hypothesis supported. depends on hypothesis.
•
What is your conclusion concerning the effect of pH on enzyme activity? Any pH
other than the optimum pH will decrease the activity of an enzyme.
5.5 Factors that Affect Enzyme Activity (LM page 51)
Table 5.5 Factors that Affect Enzyme Activity
Factors
Promote Enzyme Activity
Enzyme Specificity
Active site available
Temperature
Intermediate
Enzyme or substrate
High
concentration
pH
Optimum pH
Inhibit Enzyme Activity
Active site not available
Extreme
Low
Too acidic or basic for the enzyme
Conclusions: Factors that Affect Enzyme Activity (LM page 51)
•
Why does enzyme specificity promote enzyme activity? The active site of an
enzyme has a shape that is complementary to the shape of its substrate. In this way the
enzyme brings together the substrates so that the reaction occurs.
•
Why does a warm temperature promote enzyme activity? It increases the motion
of molecules and therefore the number of times substrates find the active site.
•
Why does increasing enzyme concentration promote enzyme activity? It
increases the number of active sites available.
•
Why does optimum pH promote enzyme activity? Optimum pH is required to
maintain the shape of the active site of an enzyme.
LABORATORY REVIEW 5
(LM page 52)
1. What happens at the active site of an enzyme? Substrates are oriented to bring about
the reaction.2. On the basis of the active site, explain why the following conditions
speed a chemical reaction:
a. More enzyme There are more active sites available for substrates.
b. More substrate It is more likely that a substrate molecule will encounter an active site.
3. Name two other conditions (other than the ones mentioned in question 2) that
maximize enzymatic reactions. a. optimum temperature
b. optimum pH
4. Explain the necessity for each of the two conditions you listed in question 3.
a. Movement of molecules increase as temperature rises
b. Enzyme shape is maintained
5. Lipase is a digestive enzyme that digests fat droplets in the basic conditions
(NaHCO3 is present) of the small intestine. Indicate which of the following test
tubes would show digestion following incubation at 37°C, and explain why the
32
others would not.
Tube 1: Water, fat droplets No enzyme
Tube 2: Water, fat droplets, lipase Wrong pH
Tube 3: Water, fat droplets, lipase, NaHCO3 Digestion occurs
Tube 4: Water, lipase, NaHCO3 No substrate
6. Fats are digested to fatty acids and glycerol. As the reaction described in question
proceeds, the solution will become what type pH? acidic Why? Fatty acids are
present.
7. Given the following reaction:
catalase
2 H2O2
H2O + O2
hydrogen
water oxygen
peroxide
a. Which substance is the substrate? hydrogen peroxide
b. Which substance is the enzyme? catalase
c. Which substances are the end products? water and oxygen
d. Is this a synthetic or degradative reaction? degradative
How do you know? The larger molecule on the left becomes the two smaller molecules on
the right.
33
Laboratory
6
Photosynthesis
(LM pages 53–62)
Fourth Edition
This lab remains the same as it was in the previous edition.
MATERIALS AND PREPARATIONS
Instructions are grouped by exercise. Some materials may be used in more than one exercise.
Special Requirements
Fresh material. Fresh or frozen spinach, depending on preparation alternative chosen, for
6.1 Photosynthetic Pigments
Living material. Duckweed or Elodea (Anacharis) for 6.2 Solar Energy and 6.3 Carbon
Dioxide Uptake
Equipment, preassembly required. Volumeter for 6.2 Solar Energy and 6.3 Carbon
Dioxide Uptake
Fume hood for 6.1 Photosynthetic Pigments
All Exercises
_____ safety goggles (See Carolina’s Safety: Face Protection Section)
_____ latex gloves and/or nonlatex gloves (See Carolina’s Safety: Hand Protection
Section)
_____ lab coats (See Carolina’s Safety: Body Protection Section) or other clothing
protection
_____ distilled water
_____ wax pencils
_____ rulers, plastic centimeter
6.1
Plant Pigments (LM pages 54–55)
_____ fresh spinach pigment extract:
_____ spinach, fresh
_____ blender, glass or stainless steel
_____ cheesecloth
_____ polypropylene utility funnel, 4 ¼”, or Buchner funnel
_____ filter paper
_____ acetone
_____ frozen spinach pigment extract alternative:
_____ spinach, frozen, 40 g
_____ blender, glass or stainless steel
_____ acetone
34
_____
_____
_____
_____
_____
_____
_____
_____
_____
_____
_____ ethanol
_____ sodium chloride (Crystal)
_____ filter, paper
_____ amber bottle(s)
test tubes, rimless, large culture 25 × 150 mm
cork with paper-clip hook for large test tubes
chromatography paper, Whatman #1
glass capillary tube
paper towels
scissors
fume hood
chromatography solution:
_____ petroleum ether
_____ acetone
_____ jar, wide-mouth, screw-cap
test tube rack, 25 mm holes
pencils
Whatman #1 chromatography paper (LM page 54). Use sheets of 12 × 12 cm Whatman
#1 chromatography paper. Cut the sheets to fit the chromatography apparatus, rounding or
pointing one end.
Fume hood and cautions (LM page 54). For the chromatography exercise, direct students’
attention to the fume hood and ether cautions in the Lab Manual.
Fresh spinach pigment extract (LM page 55). If a fume hood is available, prepare the
extract there. Wash and thoroughly drain the spinach. Cut the veins and petioles from the
leaves. Put the spinach in a glass or stainless steel blender, add acetone, and blend to form a
thick slurry. Extract should be filtered, using a cheesecloth plug in a funnel or a small
Buchner funnel with aspiration. Refrigerate the slurry in a tightly stoppered container labeled
“Pigment Extract.” Extract exposed to light and room temperature begins decomposing
within an hour, while refrigerated extract may last overnight.
An alternate method involves drying spinach leaves slowly in a dry oven and then
pulverizing them in a blender or with a mortar and pestle. Leaf powder is useful for weeks if
stored in a sealed container and placed in a cool, dark area. Pulverization reduces leaf volume
considerably. The dry leaf powder can be added to a small amount of acetone to form a thick
slurry.
Frozen spinach pigment extract alternative (LM page 55). Partially defrost and divide a
package of frozen spinach into 40 g portions. Combine 40 g frozen spinach with 200 ml
acetone in a blender. Blend 2 to 3 minutes on high. Let stand 3 minutes. Decant supernatant,
save as 1. Add 100 ml ethanol to solids remaining in blender. Blend 2 to 3 minutes on high.
Decant supernatant, save as 2. Combine 1 and 2, and filter to remove any remaining solids.
Add a pinch of sodium chloride. Refrigerate in amber jar.
Chromatography solution (LM page 55). 100 ml is sufficient for five student groups.
Combine forty-five parts petroleum ether with five parts acetone, and store in a screw-capped
35
container. Label as “Chromatography Solution.” Keep the container tightly closed, since this
solution is volatile and extremely flammable. (If a fume hood is available, prepare the
solution there.) Have a wide-mouth, screw-capped jar, labeled “Used Chromatography
Solution,’’ available in which to place used solution. Keep the jar tightly closed.
Disposal (LM page 55) Organic solvents should be recycled or disposed of according to
local procedures and regulations.
6.2
Solar Energy (LM pages 56–59)
_____ Duckweed or Elodea, fresh
_____ aeration equipment for Elodea, aquarium air pump 10-20 gal tank
_____ razor blades, single-edged
_____ volumeter:
_____ test tubes, large culture, 25 × 150 mm rimless
_____ rubber stoppers, #5, single-holed
_____ glycerol
_____ pipet, graduated
_____ sodium bicarbonate powder 3% (NaHCO3) solution
_____ water, distilled
_____ aquarium aerator for sodium bicarbonate
_____ test tube rack for 25 mm tubes
_____ beaker, 1,000 ml (plastic, glass)
_____ lamp, 150 watt or aquarium light (full-spectrum bulb)
Volumeter (LM page 57). Prepare one volumeter per student group ahead of time. Insert a
graduated pipette into a single-holed rubber stopper that fits into a large culture test tube, as
shown in Figure 6.4. When the rubber stopper is in place during the experiments, a
continuous column of liquid will form between the test tube and the pipette. Adjust the
placement of the leading edge of the liquid by applying pressure to the stopper. The oxygen
emitted by the Elodea will displace the liquid in the test tube, thus moving the edge of the
liquid in the pipette. The student will read the change in millimeters.
3% sodium bicarbonate (NaHCO3) solution (LM page 56) Prepare 125 ml per student
group. Dissolve 30 g of NaHCO3 in 1,000 ml of distilled water. Aerate the solution with an
aquarium aerator for 30 minutes before the laboratory exercise to saturate with carbon
dioxide. Discard the solution after use.
Elodea (LM page 56) Use fresh duckweed or Elodea (one healthy sprig per student group is
sufficient) that has been maintained in continuously aerated distilled water. Change the water at
least every two days.
6.3
Carbon Dioxide Uptake (LM page 60)
_____ 0.04% phenol red solution
_____ straws, individually packaged
_____ Elodea (or duckweed),
_____ Volumeter as described in Section 7.2.
36
0.04% phenol red solution (LM page 60). Prepare 100 ml per student group. Dissolve 0.04
g of phenol red in 100 ml of distilled water. Have students use caution when blowing through
the straw into the test tube of phenol red. Overzealous students may blow the phenol red out
of the tubes and onto themselves. Students need only blow on the surface of the liquid to get
a color change.
EXERCISE QUESTIONS
6.1 Plant Pigments (LM pages 54–55)
Experimental Procedure: Plant Pigments (LM page 54–55)
9.
Which pigment is the most nonpolar (that is, has the greatest affinity for the
nonpolar solvent)? beta-carotene
10.
Calculate the Rf (ratio-factor) values for each pigment. Beta-carotenes will have
the largest values, which will be less than one, and chlorophyll b will have the smallest.
11.
Do your results suggest that the chemical characteristics of these pigments might
differ? Yes Why? They must differ, otherwise all the pigments would migrate the same
distance.
6.2 Solar Energy (LM pages 56–59)
Verify that photosynthesis releases oxygen by writing the equation for photosynthesis.
solar energy
CO2 + H2O —————————————> (CH2O)n + O2
Role of White Light
Experimental Procedure: White Light (LM pages 56–58)
4.Measure in millimeters the distance the edge moved. Why did the edge move
forward? The edge moved in response to oxygen production, which forced the liquid
outward in the tubing.
5.Record the length of time it takes for the edge of the solution in the tubing to recede
1mm. Answers cab vary. Why does cellular respiration, which occurs in a plant all the
time, cause the edge to recede? Oxygen, which was produced during photosynthesis, was
being used by the plant during cellular respiration. As the volume of oxygen decreased
(because photosynthesis is not occurring when the tube is wrapped by foil), less water was
forced into the tubing, and the edge receded.
6.
If the Elodea had not been respiring in step 4, how far would the edge have
moved? Add the distance the edge receded to the distance the edge moved forward during
the initial experiment with the white light.
7.
Calculate the rate of photosynthesis. 201 mm/hr (Rates will vary with plant
condition, distance from the lamp, and room temperature.)
Table 6.2 Rate of Photosynthesis (White Light)
Net photosynthesis (white light)
Cellular respiration (no light)
Gross photosynthesis (net + cellular respiration)
Rate of photosynthesis
Data
32/10 min
1.5/10 min
33.5 (mm/10 min)
201 (mm/hr)
37
Role of Green Light
According to Fig. 6.5, what color light do the chlorophylls absorb best? violet, blue, and
red Least? green, the reflected color
What color light do the carotenoids (carotenes and xanthophylls) absorb best? greenyellow Least? yellow, orange, the reflected colors
Does photosynthesis use green light? not extensively
Experimental Procedure: Green Light (LM pages 58–59)
8.
Based on your data in Table 6.3, this percentage = 37%
Based on class data in Table 6.3, this percentage = Compute from class data.
Table 6.3 Rate of Photosynthesis (Green Light)
Your Data
Class Data
Gross Photosynthesis (mm/10 min)
White (from Table 6.2)
33.5 mm/10 min
Green
11.5 mm/10 min
Rate of Photosynthesis (mm/hr)
White (from Table 6.2)
201 mm/hr
Green
75 mm/hr
Note: The results presented in this table are sample data. Actual results will vary.
Conclusions (LM page 59)
•
Explain why the rate of photosynthesis with green light is only a portion of the
rate of photosynthesis with white light. Photosynthesis does not use green light extensively.
•
How does the percentage based on your data differ from that based on class
data? Explanation will vary according to particular student.
6.3 Carbon Dioxide Uptake (LM page 60)
Experimental Procedure: Carbon Dioxide Uptake (LM page 64)
2.
Blowing onto the solution adds what gas to the test tube? primarily carbon
dioxide When carbon dioxide combines with water, it forms carbonic acid. What causes
the color change? Carbonic acid releases hydrogen ions. As the pH decreases, the color of
the indicator changes from red to yellow.
6.
Hypothesize why the solution in the test tube eventually turned red. The plant
uses carbon dioxide in photosynthesis. As carbon dioxide is absorbed, carbonic acid is
reconverted to carbon dioxide and water. When the plant has taken up all the blown-in
carbon dioxide, the amount of hydrogen ions and, therefore, the pH of the solution, returns to
the previous level. Therefore, the phenol red returns to its initial color.
Use of a Control (LM page 60)
•
Considering the test sample in Table 6.4, suggest a possible control sample for
this experiment: a sample that does not contain Elodea but that contains phenol red with
carbon dioxide blown in to produce the same yellow color
•
Why should all experiments have a control? In a control sample, the variable
38
being tested is missing. Therefore, if a control sample gives positive results, the experiment is
invalid—the reagents may be contaminated or the procedure may need improvement.
Table 6.4 Carbon Dioxide Uptake
Tube
Time for Color Change
Test sample: Elodea + phenol red solution + CO2 30–40 minutes
Control sample: CO2 + phenol red solution
No change
6.4 Carbon Cycle (LM page 61)
1.
Which organelle in plants carries out the reaction in the previous equation in the
reverse (right-to-left) direction? chloroplast
2.
Pertaining to photosynthesis, the energy in the equation is provided by solar
energy.
3.
Which organelle in plants and animals is involved in carrying out the reaction in
this equation in the forward direction? mitochondria
4.
Pertaining to cellular respiration, the energy in the equation becomes chemical
bond energy in what molecule? ATP
5.
Would it be correct to say that solar energy eventually becomes the chemical
bond energy in ATP? yes Why? Because solar energy becomes chemical bond energy of
carbohydrates and chemical bond energy of carbohydrates becomes that of ATP molecules.
6.
Considering that both plants and animals carry on cellular respiration, revise
Figure 6.6 to improve its accuracy. The corn plants should be placed next to the cow also.
LABORATORY REVIEW 6 (LM page 62)
1. Where In do the light reactions of photosynthesis take place? In thylakoid
membranes of chloroplasts.
2. What procedure did you use to separate plant pigments? chromatography
3. What determines the speed with which a pigment moves up the chromatography
paper? solubility in a solvent
4. Where do plants ordinarily get the energy they need to carry on photosynthesis?
white light of solar energy
5. Blue, red, and green light are all present in what color of light? white
6. Why do blue and red light, but not green promote photosynthesis. Chlorophyll is
able to absorb blue and red light but not green light.
7.
Does Elodea respire in the light or in the dark? Both the light and the dark
8. Phenol red turns what color when carbon dioxide is added? yellow
9. What happens to carbon dioxide during photosynthesis? It is converted to
carbohydrate.
39
10. Some plants are colorless. Do you predict that they carry on photosynthesis. No, a
plant requires a pigment to absorb solar energy and photosynthesize.
40
7
Cellular Reproduction
(LM pages 63-72)
Fourth Edition
This edition has a separate lab for mitosis and another for meiosis. The mitosis lab gives
students an exercise to do as they learn the phases of mitosis.
MATERIALS AND PREPARATIONS
Instructions are grouped by exercise. Some materials may be used in more than one exercise.
All Exercises
________microscopes, compound light
________lens paper
7.2
Animal Cell Mitosis and Cytokinesis
Animal Mitosis (LM page 66-68)
______ mitosis models, animal
______ slide, prepared: whitefish mitosis
7.3
Plant Cell Mitosis and Cytokinesis
Plant Mitosis (LM page 68-71)
______ mitosis models, plant (
______ slide, prepared: onion (Allium) root tip
Note: For the experimental procedure regarding time span of phases, it would be best to have
students pool their data; a minimum of 40 observed cells should work well.
EXERCISE QUESTIONS
7.1 The Cell Cycle (LM pages 64–65)
State the event of each stage on the line provided.
G1
Organelles begin to double in number.
S
Replication of DNA
G2
Synthesis of proteins
M
Mitosis
Explain why the entire process is called the “cell cycle.” In dividing cells, the stages
repeat.
The S Stage of the Cell Cycle (LM page 65)
Label the sister chromatids, and centromere in the drawing of a duplicated chromosome in
Figure 7.3 1. sister chromatids; 2. centromere
41
Each daughter cell will have 18 chromosomes.
The M Stage of the Cell Cycle (LM page 65)
Consult Figure 7.2 and write the phases of mitosis here: prophase, metaphase, anaphase,
telophase
7.2 Animal Cell Mitosis and Cytokinesis (LM page 68)
Observation: Animal Cell Mitosis (LM page 66)
1. What is the number of chromosomes in the parent cell and in the daughter cells in
this model series? Answer may vary, depending on what model is being used.
3. Do these models show the spindle, which is illustrated in Figure 7.4? Most likely yes.
4. What is the shape of animal cells? Blastula cells are round. What is the appearance
of the spindle pole? An aster is present.
Whitefish Blastula Slide (LM page 67)
3. Match these statements to the correct phase of animal cell mitosis to Figure 7.5 and
write the correct statements on the lines provided.
Prophase: Duplicated chromosomes have no particular arrangement in the cell
Metaphase: Duplicated chromosomes are aligned in the equator of the spindle
Anaphase: Daughter chromosomes are moving to the poles of the spindle
Telophase: Two daughter cells are now forming
4. Explain the different appearance of the chromosomes. In the prophase cell, the
chromosomes are duplicated and in the telophase cell, the chromosomes are single.
Cytokinesis in Animal Cells (LM page 68)
Are any of the cells in your whitefish blastula slide undergoing cytokinesis? Most likely
yes.
Do you see any cleavage furrows? Most likely yes.
7.3 Plant Cell Mitosis and Cytokinesis (LM page 68-71)
Observation: Plant Cell Mitosis (LM page 69)
3. What is the number of chromosomes in each of the cells in this model series? Answer
may vary, depending on what model is being used.
Onion Root Tip Slide (LM page 69)
Time Span for Phases of the Cell Cycle in the Onion Root Tip (LM p. 69-70)
Hypothesize how many minutes the cell spends during each of these phases of the cell
cycle. Hypotheses will most likely vary.
Experimental Procedure Time Span for Phases of the Cell Cycle in the Onion Root Tip (LM
p. 70)
3. Calculate the percentage of cells in each phase of the cell cycle and record in Table
7.2.
4. Calculate the time span for each phase of the cell cycle and record in Table 7.2.
42
Table 7.2 Time Span for Phases of the Cell Cycle in the Onion Root Tip
Phase
Number Seen
% of Total
Time Span (min)
Interphase 30
75
1080
Prophase
4
10
144
Metaphase 1
2.5
36
Anaphase
2
5
72
Telophase
3
7.5
108
Total
40
100
1440
Conclusions: Time Span for Phases of the Cell Cycle in the Onion Root Tip (LM 70)
 Were your hypotheses supported or not supported by your observation of onion
root tip cells undergoing the cell cycle? Answers will vary.
 Suggest a possible explanation for the length of time a cell spends on different
phases of the cell cycle. During interphase a cell is carrying on normal activities;
During prophase, a cell gets ready for metaphase because the chromosomes must be
positioned correctly and this allows the daughter chromosomes to quickly separate
during anaphase. Telophase takes longer because the cells reorganize into daughter
cells.
Cytokinesis in Plant Cells (LM page 71)
Were any of the cells undergoing cytokinesis as shown in Figure 7.7 during telophase?
Most likely, yes.
How do you know? Cell plate is present.
Offer an explanation for why Figure 7.8 is so detailed. It’s an electron micrograph.
Would you predict that the vesicles of the cell plate lay down the new cell wall inside or
outside the vesicles? Explain your answer. Inside, because this is where the components
to make the cell wall accumulate.
LABORATORY REVIEW 7 (LM page 72)
1. Divide the cell cycle into two main portions, and tell in general what is happening in
these two portions. Interphase (cells are contributing to the workings of the body) and
Mitosis (cells are dividing).
2. Most of the time the cell is in which of these portions of the cell cycle? Why is this
advantageous? Interphase because this is the time that cells go about their normal
activities.
3. Name and define the two events that take place when a cell divides. Mitosis (nuclear
division) and cytokinesis (division of cytoplasm) occur when a cell divides.
4. What is the function of the centromere during mitosis? The centromere provides a
place of attachment for spindle fibers.
43
5. Evolution of the spindle was of central importance to eukaryotic cells. What role is
played by the spindle during mitosis?
The chromosomes move because they are attached to spindle fibers that lengthen (push) and
shorten (pull) the chromosomes.
6. Do the chromosomes have sister chromatids during these phases of mitosis (yes or
no)?
a. Prophase Yes
b. Metaphase Yes
c. Anaphase No
d. Telophase No
7. Contrast the appearance of animal cells and plants during mitosis. Animal cells have
asters but plant cells do not have asters.
8. Explain how it is possible for each phase of mitosis and the daughter cells to have the
same number of chromosomes. Duplication of chromatids provides the daughter
nuclei for daughter cells. Counting the centromeres tells the number of chromosomes.
9. Contrast how cytokinesis occurs in animal cells with how it occurs in plant cells.
Cytokinesis in animal cells occurs by furrowing; in plant cells it occurs by formation of a cell
plate.
10. What would a tissue look like if cytokinesis did not occur? The tissue would have multiple
nuclei.
44
Laboratory
8
Sexual Reproduction
(LM pages 73–82)
Fourth Edition
This edition has a separate lab for mitosis and another for meiosis. Section 8.2 now
emphasizes how meiosis results in variation among the gametes. Text art was added to this
section as a guide to the hands-on experimental procedure in this section.
MATERIALS AND PREPARATIONS
Instructions are grouped by exercise. Some materials may be used in more than one exercise.
8.1
Meiosis: Reduction Division (LM pages 74-75)
Meiosis in Lily Anther (p. 74)
______ Microscope slide of meiosis in Lily anther
8.2
Production of Variation During Meiosis Building Chromosomes to Simulate
Meiosis (LM page 76-78)
______ Chromosome Simulation
______ scissors
______ Animal meiosis model74
8.3
Human Life Cycle
______ Paper and pencil
EXERCISE QUESTIONS
8.1 Meiosis: Reduction Division (LM pages 74-75)
Before proceeding
a. Distinguish between diploid (2n ) and haploid (n): Diploid equals the full number of
chromosomes when there are pairs of chromosomes. Haploid equals half the full number
of chromosomes when each type chromosome does not have a pair.
b. Distinguish between a homologue and a tetrad: Each member of a pair of chromosomes
is called a homologue. Tetrads occur when the homologues are paired and each one
consists of two chromotids; therefore four chromatids are in close proximity.
Observation: Meiosis in Lily Anther (LM pages (74-75)
Phases of Meiosis I (LM page 74-75)
45
2. Tell what is happening in each of these (meiosis I) phases. See descriptions in Figure
8.3 Meiosis I
3. In Figure 8.3, place a 2n or n beside each drawing. Cells in prophase I, metaphase I,
and anaphase I are 2n; each cell forming in Telophase I and daughter cells in interkinesis
are n.
Phases of Meiosis II (LM page 75)
2. Tell what is happening in each of these (meiosis II) phases. See descriptions in Figure
8.4 Meiosis II.
3. In Figure 8.4, place a 2n or n beside each drawing. All cells during meiosis II are n.
8.2 Production of Variation During Meiosis (LM 76-78)
Anaphase II (LM page 78)
10. Pull the two magnets of each duplicated chromosome apart. What does this action
represent? This action represents separation of centromeres and daughter chromosomes
moving to opposite poles.
Telophase II (LM page 78)
11. At the end of telophase, the daughter nuclei reform.
•
Therefore, how many nuclei are usually present when meiosis II is complete?
four
•
In this exercise, how many chromosomes were in the parent cell nucleus
undergoing meiosis II? two
•
How many chromosomes are in the daughter nuclei? two Explain. When the
chromatids of the chromosomes in the parental cell separate, they become daughter
chromosomes, which go into the daughter cells.
Summary of Production of Variation During Meiosis (LM page 78)
1. If the parent cell is 2n=4, the daughter cells are n = 2.
2. Why do the puppies born to these parents show variation?
a. This process is called crossing-over.
b. During metaphase I, align independently, and therefore differently. This means
that daughter cells following telophase I can have different combinations of
chromosomes.
c. During fertilization, variant sperm fertilize variant eggs helping to ensure that the
new individual inherits a different combination of chromosomes than the parent had.
8.3 Human Life Cycle (LM pages 79-81)
As you read the following text, fill in boxes in Figure 8.6 with the terms “mitosis” or
“meiosis.” Left to right: meiosis, mitosis, mitosis.
Summary of Human Life Cycle (LM page 79)
Fill in the blanks to ensure your understanding of the role of meiosis and mitosis in
humans.
1. Name of organ that produces gametes in males testes; in females ovaries
46
2. Name of process that produces gametes in males spermatogenesis; in females
oogenesis
3. Type of cell division involved in process in males meiosis; in females meiosis
4. Name of gamete in males sperm; in females egg
5. Number of chromosomes in gamete in males n; in females n
6. Results of fertilization zygote
7. Number of chromosomes provided 2n
Mitosis Versus Meiosis (LM pages 80)
Table 8.1
Differences Between Mitosis and Meiosis
1. Number of divisions
2. Chromosome number in daughter cells
3. Number of daughter cells
Table 8.2
Mitosis
One
Same as the
parent cell
Two
Meiosis
Two
One-half of the
parent cell
Four
Mitosis Compared with Meiosis I
Mitosis
Prophase: No pairing of chromosomes
Metaphase: Duplicated chromosomes at metaphase plate
Anaphase: Sister chromatids separate
Telophase: Chromosomes have one chromatid
Meiosis I
Prophase I: Pairing of
homologues
Metaphase I: Duplicated
homologues at equator
Anaphase I: Homologues
separate
Telophase I: Chromosomes
have two chromatids.
Provide the correct term for each definition. (LM page 80)
1. Type of cell division that keeps the chromosome number the same and occurs during
growth and repair Mitosis
2. Type of cell division that reduces the chromosome number and occurs during gamete
formation Meiosis
3. Half the diploid number of chromosomes Haploid (n) number
4. Male gamete with the n number of chromosomes Sperm
5. Female gamete with the n number of chromosomes Egg
LABORATORY REVIEW 8 (LM page 82)
1. Where would you expect to find meiosis taking place in human males? testes
females? ovaries
47
2. If there are 13 pairs of homologues in a parent cell, how many chromosomes
are there in a sperm? 13 Explain how you arrived at this number. Pairs separate
during meiosis.
3. Account for why each of your body cells contains two of each kind of
chromosome. Prior to mitosis, DNA replicated and the chromosomes duplicated.
Separation of chromatids provides the 2n number for each cell.
4. Does metaphase of mitosis, meiosis I, or meiosis II have the haploid number of
chromosomes at the equator of the spindle?
meiosis II
5. List four differences when comparing mitosis with meiosis.
Figure 8.7
See Table 8.2 and
6. If the cells of an organism have 12 chromosomes, what is the number of
chromosomes at the equator during metaphase of mitosis? 12
during metaphase of meiosis II? 6
7. A student is simulating meiosis I with a pair of homologues that are red-long
and yellow-long. Why would you not expect to find both red-long and yellow-long
in one resulting daughter cell? Red-long and yellow-long are homologues and
homologues separate during meiosis.
8. With reference to a pair of homologues, describe the change in the two
participating nonsister chromatids following crossing-over. Each nonsister
chromatid will have genetic material from the other nonsister chromatid.
9. What would be the appearance of a cell that completes mitosis but does not
undergo cytokinesis? The cell would have two nuclei.
with a cell that completes meiosis but does not undergo cytokinesis? The cell would
have four nuclei.
10. In the life cycle of humans, when does mitosis occur? During growth and
development of zygote and during growth of individual.
48
Laboratory
9
Patterns of Inheritance
(LM pages 83–96)
Fourth Edition
This lab was rewritten to facilitate its use by the instructor because students observe only the
results of live genetic crosses rather than performing the crosses. More paper and pencil
practice is provided for students as they fill in Punnett squares and do additional genetics
problems.
New or Revised Figures: 9.2 What are the expected results of a cross?; 9.4 Monohybrid
Cross (in corn) ; FOIL method to determine gametes for two-trait problems (p. 89)
MATERIALS AND PREPARATIONS
Instructions are grouped by exercise. Some materials may be used in more than one exercise.
Special Requirements
Living material. Tobacco seedlings for 9.1 One-Trait Crosses. Alternative corn seedlings
may be used. Drosophila flies for 9.2 and 9.3
All sections
______stereomicroscope
______ hand lens
______ lens paper
9.1 One-Trait Crosses (LM pages 84-88)
Color of Tobacco Seedlings (LM page 85-86)
_____ tobacco seedlings (a Biokit®, is available from Carolina)
The kit contains seeds, growth chambers, and germination papers for a class of thirty
students. Sow seeds approximately ten days before use. The seedlings can be maintained for
about a week. (The albino ones will die shortly thereafter.)
Color of Corn Kernels (LM pages 86-87)
_____ Corn Genetics Biokit® (Carolina Biological Supply) The kit comes with
fifteen ears, marker pins, a teacher’s manual, and thirty student guides. A variety of genetic
corns and student guides are available.
9.2
Two-Trait Crosses (LM pages 88-93)
Color and Texture of Corn (LM pages 89-90)
49
_____ Corn Dihybrid Genetics Biokit® (Carolina 17 6380) This kit comes with
fifteen ears, marker pins, a teacher’s manual, and thirty student guides. A variety of other
genetic corns and student guides are available.
Wing Length and Body Color in Drosophila (LM pages 91-93)
_____ Results of the cross LlGg X llgg from Carolina Biological Supply (optional)
_____ card, white index
_____ lens paper
_____ brush, camel-hair
Drosophila Cross (LM page 91). If you wish students to count the flies do this: After
receiving the results of the cross LlGg X llgg from Carolina Biological Supply, transfer the
flies into a fresh vial that has no culture medium and freeze the flies overnight. Have students
put the flies back into the vial and return to you for use by another group. As the culture
bottle produces new flies, continue to freeze them for future labs.
9.3
X-Linked Crosses (LM pages 93-95)
Red/White Eye Color in Drosophila
_____ Results of the cross red-eyed male × heterozygous red-eyed female from
Carolina Biological Supply (optional)
_____ card, white index
_____ lens paper
_____ brush, camel-hair
Drosophila X-linked cross (LM 93) If you wish students to count the flies do this: After
receiving the results of the cross red-eyed male x heterozygous red-eyed female from
Carolina Biological Supply, transfer the flies into a fresh vial that has no culture medium and
freeze the flies overnight. Have students put the flies back into the vial and return to you for
use by another group. As the culture bottle produces new flies, continue to freeze them for
future labs.
EXERCISE QUESTIONS
9.1 One-Trait Crosses (LM pages 84–88)
Color of Tobacco Seedlings (LM pages 85-86)
Experimental Procedure: Color of Tobacco Seedlings
What is the expected phenotypic ratio? 3:1 = three green plants to one white plant
Table 9.1 Color of Tobacco Seedlings
Number of Offspring
Green Color
White Color Phenotypic Ratio
Plate #
3:1
Plate #
3:1
Plate #
3:1
Totals
85
30
3:1
Class Data
3:1
50
Conclusions: Color of Tobacco Seedlings (LM page 86)

Do your results differ from the expected ratio? yes Explain. Counting small numbers
of offspring is more likely to cause a variation from the expected ratio.

Was your class data closer to the expected ratio? yes
Color of Corn Kernels (LM, page 86-87)
Experimental Procedure: Color of Corn Kernels (LM, page 87)
What is the expected phenotypic ratio? 1:1=one purple kernel to one yellow kernel
Table 9.2 Color of Corn Kernels
Number of Kernels
Purple Color
Yellow Color Phenotypic Ratio
Plate #
1:1
Plate #
1:1
Plate #
1:1
Totals
60
60
1:1
Class Data
1:1
Conclusions: Color of Corn Kernels (LM page 87)

Do your results differ from the expected ratio? yes Explain. Counting small numbers
of offspring is more likely to cause a variation from the expected ratio.

Was your class data closer to the expected ratio? yes Explain. Counting a large
number of offspring is more likely to result in the expected ratio.
One-Trait Genetics Problems (LM page 88)
1. In pea plants purple flowers (P) is dominant and white flowers (p) is recessive. What
is the genotype of pure-breeding white plants? Pure-breeding means that they produce
plants with only one phenotype. pp If pure-breeding purple plants are crossed with
these white plants, what phenotype is expected? Purple plants
2. In pea plants, tall (T) is dominant and short (t) is recessive. A heterozygous tall
plant is crossed with short plant. What is the expected phenotypic ratio? 1:1
3. Unexpectedly to the farmer, two tall plants have some short offspring. What is the
genotype of the parent plants and the short offspring?: parents are Tt and offspring is tt
4. In horses, two trotters are mated to each other and produce only trotters and pacers
are mated to each other and produce only pacers. When one of these trotters is mated
to one of the pacers, all the horses are trotters. Create a key and show the cross. Key: T
= trotter, t = pacer Cross: TT X tt
5. A brown dog is crossed with two different black dogs. The first cross produces only
black dogs and the second cross produces equal numbers of black and brown dogs.
51
What is the genotype of brown dog? bb The first black dog? BB The second black dog?
Bb
6. In pea plants green pods (G) is dominant and yellow pods (g) is recessive. When two
pea plants with green pods are crossed, 25% of the offspring have yellow pods. What is
the genotype of all plants involved? parents are Gg ; 50% of offspring are Gg; 25% of
offspring are gg; 25% of offspring are GG.
7. A breeder wants to know if a dog is homozygous black or heterozygous black. If the
dog is heterozygous, which cross is more likely to produce a brown dog Bb X bb or Bb
X Bb? Explain The cross Bb X bb gives a 50% chance of a brown dog but Bb X Bb gives a
25% chance of a brown dog.
8. If the cross in #6 produces 220 plants, how many offspring have green pods and how
many have yellow pods? 55 have yellow bods and 165 have green pods If the cross in #2
produces 220 plants, how many offspring are tall and how many are short? 110 are tall
and 110 are short
9.2
Two-Trait Crosses (LM pages 88-93)
Color and Texture of Corn (LM pages 89–90)
Experimental Procedure: Color and Texture of Corn
1.
Do the Punnett square in order to state the expected phenotype ratio among the
offspring 9:3:3:1 (9 purple smooth to 3 purple rough to 3 yellow smooth to 1 yellow rough )
Table 9.3 Color and Texture of Corn
Purple
Smooth
162
78
51
291
Purple
Rough
52
16
16
97
Number of Kernels
Yellow
Smooth
55
20
18
93
Yellow
Rough
18
9
6
33
Phenotypic
Ratio
Sample # ____
9:3:3:1
Sample # ____
9:3:3:1
Sample # ____
9:3:3:1
Totals
9:3:3:1
Class Data
9:3:3:1
These data are possible, however individual and class data will vary. Questions can be
answered using these data if students do not do the experiment.
Conclusions: Color and Texture of Corn (LM pages 90)
Calculate the actual phenotypic ratios based on the data and record in Table 9.3. Do
your results differ from the expected ratio per individual data? most likely per class
data? closer to expected ratio Explain. The more offspring counted the greater the
probably of achieving the expected ratio.
Wing Length and Body Color in Drosophila (LM pages 91-92)
What is the expected phenotypic ratio for this cross? 1:1:1:1
Table 9.4 Wing Length and Body Color in Drosophila
52
Number of Offspring
Class data
Phenotypes
Long Long Short Short
Gray ebony Gray Black
28
32
25 30
128
120
120
120
Phenotypic
Ratio
1:1:1:1
1:1:1:1
Conclusions: Wing Length and Body Color in Drosophila (LM page 92)
Calculate the actual phenotypic ratio based on the data and record in Table 9.4. Do the
results differ from the expected ratio per individual data? yes, probably per class data
not as much. Explain. The more offspring that are counted, the greater is the probability of
achieving the expected ratio.
Two-Trait Genetics Problems (LM pages 92-93)
1. In tomatoes, tall is dominant and short is recessive. Red fruit is dominant and yellow
fruit is recessive. Choose a key for height: T= tall, t= short for color of fruit: R = red, r
= yellow What is the genotype of a plant heterozygous for both traits? TtRr What are
the possible gametes for this plant? TR, tR, tr, Tr
2. Using words, what are the likely parental genotypes if the results of a two-trait
problem are 1:1:1:1 among the offspring? heterozygous in both traits X homozygous
recessive in both traits
3. In horses, black (B) and trotting gait (T) are dominant while brown (b) and a pacing
gait (t) are recessive. If a black trotter (homozygous for both traits) is mated to a
brown pacer , what ratio is expected among the offspring? All black trotter
4. Two black trotters have a brown pacer offspring. What is the genotype of all horses
involved? black trotter parents: BbTt brown pacer offspring: bbtt
5. The phenotypic ratio among the offspring for two corn plants producing purple and
smooth kernels is 9:3:3:1. (See lab for the key) a. What is the genotype of the parental
plants? PpSs What is the phenotype of the 9 offspring? Purple smooth 3 of the offsping?
purple rough the other 3? yellow smooth and the 1 offspring? yellow rough
6. Which matings could produce at least some fruit flies heterozygous in both traits?
Write yes or no beside each. (You do not need a key)
ggLl X Ggll yes
GGLl X ggLl yes
GGLL x ggll yes
Explain. In each cross, it is possible to choose a GgLl combination for the offspring.
7. State two new crosses that could not produce fruit flies heterozygous in both traits?
GGLL X GGLL, GGLL X GgLL, GGLl X GGLl (Any combination in which the offspring
must receive two capital letters for one of the traits.)
ggll X ggll
53
8. Chimpanzees are not deaf if they inherit both an allele E and an allele G. A cross
between two deaf chimpanzees produces only chimpanzees that can hear. What are the
genotypes of of all chimpanzees involved? Parents: GGee X ggEE Offspring: GgEe
9.3
X-linked Crosses (LM pages 93-94)
Red/White Eye Color in Drosophila (LM page 93-94)
Complete this Punnett square and state the expected phenotypic results for this cross?
females all red eyes males 1:1
Experimental Procedure: Red/White Eye Color in Drosophila (LM pages 94)
Table 9.5 Red/White Eye Color in Drosophila
Number of Offspring
Your Data:
Red Eyes
White Eyes
Males
16
17
Females
63
0
Class Data:
Males
45
48
Females
215
0
Phenotypic Ratio
1:1
All red eyes
1:1
All red eyes
Conclusions: Red/White Eye Color in Drosophila (LM page 94)

Do your results differ from the expected ratio per individual data? yes per class
data? not as much Explain. The more offspring that are counted, the greater is the
probability of achieving the expected ratio.

In the space provided, do a Punnett square to calculate the expected phenotypic
results for the cross XRY × XrXr. females all red eyed; males all white-eyed
X-linked Genetics Problems (LM page 95)
1. State the genotypes and gametes for each of these fruit flies:
genotype
white-eyed male
Xr Y
white-eyed female
XrXr
red-eyed male
XRY
homozygous red-eyed female
XRXR
R
r
heterozygous red-eyed female
X X
gamete(s)
Xr , Y
Xr
XR, Y
XR
R
r
X ,X
2. What are the results if a white-eyed female is crossed with a red-eyed male?
Males All white-eyed males Females All red eyes
3. Regardless of any type cross, do white-eyed males inherit the allele for white eyes
from their father or mother? mother Explain Males receive a Y from their father
4. In sheep, horns are sex linked and H = horns and h = no horns. Using symbols, what
cross do you recommend if a farmer wants to produce hornless males? XHY X XhXh
54
5. In Drosophilia, bar eye (B) is dominant and no bar eye (b) is recessive. What are the
results of these crosses?
Bar eyed male X non-barred eyed female XBY X XrXr = XrY ;All no-barred males; 1:1 Bareyed: nonbarred eyed females
Bar eyed male X heterozygous female XBY X BXb= Males 1:1 and Females all bar-eyed;
non-bar eye male X heterozygous female
6. A female fruit fly has white eyes. What is the genotype of the father? XrY What could
be the genotype of the mother? XrXr or XRXr
7. In a cross between fruit flies, all the males have white eyes and the females are 1:1.
What is the genotype of the parents? female parent XrXr male parent XRY
8. In a cross between fruit flies, a white-eyed male and red-eyed female produce no
offspring that have white eyes. What is the genotype of the parents. Male parent XrY
Female parent XRXR
LABORATORY REVIEW 9 (LM page 96)
1.
If offspring exhibit a 3:1 phenotypic ratio, what are the genotypes of the
parents? Aa x Aa
2.
In fruit flies, which of the characteristics you studied was X-linked? red/white eye
color
3.
If offspring exhibit a 9:3:3:1 phenotypic ratio, what are the genotypes of the
parental generation? AaBb x AaBb
4.
If a cross results in 90 long-winged flies to 30 short-winged flies, what are the
phenotypes of the parents? long-winged
5.
Briefly describe the life cycle of Drosophila. The adults reproduce by laying eggs.
The eggs hatch into larvae that feed. The larvae form pupae, in which the tissues are
reorganized into an adult.
6.
In the cross AaBb X aaBB, what are the gametes for AaBb? AB, Ab, aB, ab For
aaBb? aB, ab What are the genotypic results for this cross? AaBB, AaBb (2), Aabb, aaBB,
aaBb (2), aabb
7.
What is the genotype of a white-eyed male fruit fly? XrY
8.
Suppose you counted 40 green tobacco seedlings and 2 white tobacco seedlings in
one agar plate. Do your results show that both parent plants were heterozygous for the
color allele? Yes, because only Aa X Aa can produce an offspring that is aa.
9.
Suppose you counted tobacco seedlings in six agar plates, and your data were as
55
follows: 125 green plants and 39 white plants. What is the phenotypic ratio? 3:1
10. Suppose that students in the laboratory periods before you removed some of the
purple and yellow corn kernels on the ears of corn as they were performing the
Experimental Procedure. What effect would this have on your results? An accurate
9:3:3:1 ratio could not be obtained due to incomplete data.
56
Laboratory
10
DNA Biology and Technology
(LM pages 97-110)
Fourth Edition
A new procedure is used for isolating DNA in a test tube.
MATERIALS AND PREPARATIONS
Instructions are grouped by exercise. Some materials may be used in more than one exercise.
10.1
DNA Structure and Replication (LM pages 98-100)
_____
model, DNA; model kit, DNA-RNA; or puzzle kit, DNA
10.2
RNA Structure (LM pages 101-02)
_____
puzzle kit, DNA
10.3
DNA and Protein Synthesis (LM 102 -105)
_____
model kit, DNA-RNA Protein Synthesis
Kits and models. For Sections 10.1 to 10.3, DNA kits are available from Carolina Biological
Supply and Lab Aids, from which students construct models. The kits vary in degree of
sophistication and in price. Descriptions and price information for the Carolina products can
be found in the “Genetics” section of the Carolina catalog. Alternatively, students can simply
use the figures in the lab manual to gain an understanding of the concepts.
10.4 Isolation of DNA and Biotechnology (LM page 106-107)
DNA isolation
_____
safety goggles (See Carolina’s Safety: Face Protection Section)
_____
latex gloves and/or nonlatex gloves (See Carolina’s Safety: Hand
Protection Section)
_____
lab coats (See Carolina’s Safety: Body Protection Section) or other
clothing protect
_____
_____
_____
_____
_____
_____
_____
_____
slice of fruit or vegetable
mortar and pestle
large, clean test tube on ice
0.9% NaCl
dishwasher detergent (e. g., Blue Dawn)
transfer pipets
ice-cold ethanol
small, clean test tube
57
_____
For laboratory:
_____
_____
_____
_____
_____
_____
phenol red
test tubes, large
test tube rack
ice-water bath
transfer pipets
0.9% NaCl solution
95% ethanol, ice cold (5 ml per student group)
Gel Electrophoresis (LM page 106-107)
Note: If desired, students can gain an understanding of the gel electrophoresis process by
using the description and figures in the Lab Manual, rather than performing the actual
procedure.
_____ horizontal gel electrophoresis apparatus:
_____ power supply
_____ cables
_____ electrophoresis chamber with gel
_____ Electrophoresis DNA Separation Kit
Horizontal gel electrophoresis apparatus.Biological suppliers have various types of
electrophoresis apparatuses for sale. Biostar Corporation (P.O. Box 5756, Lafayette, In
47903) has Quadracell units (QEC-100) and power supply (MAB-125), which allow four
gels of four lanes each per unit.
Electrophoresis DNA Separation Kit If a kit is not obtained, the following supplies will be
needed:
Electrophoresis buffer (optional). If you have purchased a kit, the electrophoresis
buffer will be included. Otherwise, make up a sterile 5% stock TBE buffer as follows: 54 g
of Tris base (Tris aminomethane buffer), 27.5 g of boric acid, 20 ml of 0.5 M EDTA
(disodium ethylene diamine tetraacetate 2H2O) (pH 8.0). Note: The wells also can be loaded
before adding the buffer. Then they will need to be sealed with agarose solution.
Agarose solution. Agarose powder can be purchased from biological suppliers. It also
comes as part of a molecular biology experiment package, along with instructions for making
the gel slab.
Gel slabs. Gel slabs can be used immediately, or they can be covered with plastic and
left overnight (or longer) in the refrigerator.
Micropipettes and micropipette tips. Either adjustable or fixed pipettes are
recommended. When using adjustable pipettes, you need only one (5–50 ml) per setup, with
one kind of tip. To pipette 100 ml, just use the 50 ml adjustment level twice. (VWR
Scientific, with offices in many major cities, is a good supplier of adjustable pipettes.)
58
The tip can be cleaned by rinsing three times, but when working with bacteria, using
a new/sterile tip each time is preferable. (Tips can be reused after rinsing and resterilization
in their dispenser boxes.)
10.5 Detecting Genetic Disorders
____ paper and pencil
EXERCISE QUESTIONS
10.1 DNA Structure and Replication (LM pages 98–100)
Observation: DNA Structure (LM page 99)
1. Label phosphate, base pair, and deoxyribose in your drawing.
See Figure 11.2B in text
Table 10.1 Base Colors
In Figure 10.1
Cytosine
red
Thymine
purple-blue
Adenine
gold
Guanine
green
In Your Kit
3. What type of molecules make up the backbone (uprights of ladder) of DNA
(Fig. 10.1)? sugar and phosphate molecules
4. Label a hydrogen bond in Figure 10.1. The label goes on the only write-on line
available. Dashes are used to represent hydrogen bonds in Figure 10.1 because
hydrogen bonds are weak.
5. Notice … that the base A is always paired with the base T, and the base C is always
paired with the base G.
6. In Figure 10.1, what molecules make up the rungs of the ladder? hydrogen-bonded
bases adenine pairs with thymine; cytosine pairs with guanine
7. Why is DNA also called a double helix (Fig. 10.1)? The two strands making up DNA’s
ladder configuration twist around one another in the form of a helix.
DNA Replication (LM pages 99-100)
Observation: DNA Replication
1. What bonds are broken in order to unzip the DNA strands? hydrogen bonds
4. Are your molecules identical? yes
5. Because of complementary base pairing, each new double helix is composed of an
old strand and a new strand. Write old or new in 1–10, Figure 10.2a, b, and c. 1. old;
2. old; 3. old; 4. new; 5. new; 6. old; 7. old; 8, new; 4. new; 10. old Why is DNA
replication called semiconservative? Because each new double helix is composed of an
old (parental) strand and a new (daughter) strand.
6. Does replication provide a means for passing DNA from cell to cell and organism to
organism? yes Explain. By replicating (making a copy of itself) daughter cells receive a
copy of the DNA.
59
Table 10.2 DNA Replication
Old strand G G G T T C C A T T A A A T T C C A G A A A T
C A T A
New strand C C C A A G G T A A T T T A A G G T C T T T A G T A T
10.2 RNA Structure (LM Pages 101-102)
1. Describe the backbone of an RNA molecule. RNA, like DNA, has a sugar phosphate
backbone.
2. Where are the bases located in an RNA molecule? to the side
Observation: RNA Structure (LM pages 101-102)
1. Label the ribose (the sugar in RNA), the phosphate, and the base in your drawing and
in 1–3, Figure 10.3. 1. phosphate 2. base 3. ribose;
Table 10.3 Base Colors
In Figure 10.3
Cytosine
red
Uracil
purple-blue
Adenine
gold
Guanine
green
In Your Kit
Table 10.4 DNA Structure Compared with RNA Structure
DNA
RNA
Sugar
Deoxyribose
Ribose
Bases
Adenine, guanine, thymine, cytosine
Adenine, guanine, uracil,
cytosine
Strands
Double stranded with base pairing
Single stranded
Helix
Yes
No
Complementary Base Pairing (LM, p. 102)
Complete Table 10.5 to show the complimentary DNA bases for the RNA bases
Table 10.5 DNA and RNA Bases
RNA Bases C
U
DNA Bases G
A
A
T
G
C
10.3 DNA and Protein Synthesis (LM pages 103-105)
What is the role of each of these participants in protein synthesis.
DNA: stores information (i.e., proper sequence of amino acids)
mRNA: carries information to ribosomes
tRNA: brings amino acids to ribosomes
Transcription (LM page 103)
60
Label Figure 10.4. 1. RNA polymerase; 2. mRNA transcript
Observation: Transcription
Table 10.6 Transcription
DNA
T A C A C G A G C AA C T A A C A T
mRNA
A U GU G C U C G U U G AU U G U A
4. Locate the end of the strand that will move to the ribsosomes in the cytoplasm.
5’end
Translation (LM pages 104–105)
Label Figure 10.5. 1. amino acid; 2. tRNA; 3. anticodon
Observation: Translation (LM pages 104)
1. Using the mRNA sequence given in Table 10.7, number the tRNA–amino acid
complexes in the order they will come to the ribosome. Figure 10.6 : Top row : 1, 2, 5;
Bottom row : 4, 3, 6, 7
2. Complete Table 10.7. Why are the codons and anticodons in groups of three? The
genetic code is a triplet code.
Table 10.7 Translation
mRNA codons
AUG CCC GAG* GUU GAU UUG UCU
tRNA anticodons
UAC GGG CUC CAA CUA AAC AGA
Amino acid
Met Pro
Glu Val
Asp
Leu
Ser
*Both GAG and GAA code for Glu
10.4 Isolation of DNA and Biotechnology (LM page 106)
Experimental Procedure: Isolation of DNA
Answer the following questions(page 107)
1. What is biotechnology? Biotechnology is the manipulation of DNA for the benefit of
human beings and other organisms.
2. Speculate how the ability to isolate DNA and run gel electrophoresis of DNA related
to biotechnology. Isolating DNA and performing gel electrophoresis shows that
DNA is subject to laboratory procedures the same as any other molecule.
3. Name a biotechnology product someone you know is now using or taking as a
medicine. Answers will vary.
10.5 Detecting Genetic Disorders (LM pages 107–109)
Genetic Sequence for Sickle Cell Disease
1. In what three-DNA-base sequence does HbA differ from HbS?
HbA
CTC
HbS
CAC
2. What are the codons for these three bases?
HbA
GAG
HbS
GUG
3. What is the amino acid difference?
HbA
glu
HbS
val
Detection of Sickle-Cell Disease by Gel Electrophoresis (LM pages 109)
61
2. In Figure 10.10 which lane contains only HbS, signifying that the individual is
HbSHbS. Lane 2
3. Which lane contains only HbA, signifying that the individual is HbAHbA. Lane 1
4. Which lane that contains both HbS and HbA, signifying that the individual is HbAHbS.
Lane 3
Detection by Genomic Sequencing
What are the chances that this couple will have a child with sickle-cell disease? Both
partners are heterozygous; therefore the chance this couple will have a child with sickle-cell
disease is 25%.
Summary: Detecting Genetic Disorders (LM page 109)
•
What two methods of detecting genetic disorders were described in this section?
Gel electrophoresis; genomic sequencing
•
Which method is more direct and probably requires more expensive equipment
to do? Genomic sequencing
•
Which method probably preceded the other method as a means to detect sicklecell disease? Gel electrophoresis
LABORATORY REVIEW 10 (LM page 110)
1. Explain why DNA is said to have a structure that resembles a ladder. The paired
bases are the rungs of the ladder and the sugar-phosphate backbones are the supports.
2. Do the two DNA double helices following DNA replication have the same, or a
different, composition? Same
3. How is complementary base pairing different when pairing DNA to DNA than when
pairing DNA to mRNA? Uracil replaces thymine in RNA.
4. Explain why the genetic code is called a triplet code. Every three bases stands for one of
the twenty amino acids in DNA.
5. What role does each of the following molecules play in protein synthesis?
a. DNA Contains inherited genetic information.
b. mRNA Contains codons
c. tRNA Has a specific anticodon
d. Amino acids The unit molecules of a protein
6. Which of the molecules listed in question 5 are involved in transcription? DNA and
mRNA
7. Which of the molecules listed in question 5 are involved in translation? mRNA, tRNA,
and amino acids.
8. What is the purpose of gel electrophoresis? The separate DNA molecules or amino
acids.
62
9. Why does sickle-cell hemoglobin (HbS) migrate slower than normal hemoglobin
(HbA) during gel electrophoresis? HbS contains valine (no charge) instead of
glutamate (has charge).
10. Below is a sequence of bases associated with the template DNA strand:
TAC CCC GAG CTT
a. Identify the sequence of bases in the mRNA resulting from the transcription of
theabove DNA sequence. AUG GGG CUC GAA
b. Identify the sequence of bases in the tRNA anticodon that will bind with the first
codon on the mRNA identified above. UAC
63
Laboratory
11
Genetic Counseling
(LM pages 111–124)
Fourth Edition
This lab was rewritten to begin with genetic inheritance and provide students with many
genetic problems pertaining to human disorders. Multiple allele inheritance and the use of
blood type to determine paternity has been added to increase interest.
MATERIALS AND PREPARATIONS
11.2
Patters of Genetic Inheritance (LM pages 114-119)
Multiple Alleles (page 118)
_____
Kit to determine paternity available from Carolina Biological Supply
EXERCISE QUESTIONS
11.1 Determining the Genotype (LM pages 112-113)
Autosomal Dominant and Recessive Traits (LM page 112-113)
1. What is the homozygous dominant genotype for type of hairline? WW What is the
phenotype? widow's peak
2. What is the homozygous recessive genotype for finger length? ss What is the
phenotype? long fingers
3. Why does the heterozygous individual Ff have freckles? Freckles is dominant and they
have one dominant allele.
4. Maria and the members of her immediate family have attached earlobes. Her
maternal grandfather has unattached earlobes. What is the genotype of her maternal
grandfather? Ee Explain. Maria's mother has the genotype ee (results in the recessive
phenotype), therefore her maternal grandfather, who has unattached earlobes, must be Ee.
5. Moses does not have a bent little finger, but his parents do. Deduce the genotype of
his parents. Moses genotype is ll; therefore, his parents who have bent little fingers must be
Ll.
6. Manny is adopted. He has hair on the back of his hand. Could both of his birth
parents have had no hair on the back of the hand? No Explain. The presence of hair on
the back of the hand is a dominant characteristic; at least one parent had to have hair on the
back of the hand for Manny to have it.
7. Simona and her husband have widow peaks One child has a widow’s peak and the
other does not. Give the possible phenotype of all persons involved.
Isabella and her husband Ww X Ww
Child with straight hairline ww
Child with widow’s peak WW or Ww
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Experimental Procedure: Human Traits (page 113)
4. Are dominant phenotypes always the most common in a population? No Explain.
Phenotypes depend on inherited alleles and not on whether traits are dominant or recessive.
Table 11.1 Autosomal Human Traits
Answers may vary according to the class members. Students may not know whether they are
homozygous dominant or heterozygous. If so, they can use A? for their genotype, for
example.
11.2 Patterns of Genetic Inheritance (LM pages 114-119)
Inheritance of Genetic Disorders
In Figure 11.3a,
¼ offspring have the phenotype = 25% chance
¾ offspring have the phenotype = 75% chance
In Figure 11.3b,
½ offspring have the phenotype = 50% chance
1a. With reference to Figure 11.3a, if a genetic disorder is recessive and both parents
are heterozygous (Aa), what are the chances that an offspring will have the disorder?
1 in 4 (25%)
b. With reference to Figure 11.3a, if a genetic disorder is dominant and the parents are
heterozygous (Aa), what are the chances that an offspring will have the disorder? 3 out
of 4 (75%)
2a. With reference to Figure 11.3b, if parents are heterozygous (Aa) by homozygous
recessive (aa), and the genetic disorder is recessive, what are the chances that an
offspring will have the disorder? 50/50 (50%)
b. With reference to Figure 11.3b, if the parents are heterozygous (Aa) by homozygous
recessive (aa), and the genetic disorder is dominant, what are the chances that an
offspring will have the disorder? 50/50 (50%)
Autosomal Disorders (LM pages 115-116)
1. Neurofibromatosis is a dominant disorder. If a heterozygous (Aa) woman reproduces
with a homozygous (aa) normal man, what are the chances a child will have
neurofibromatosis? 50/50 (50% )
2. Cystic fibrosis is a recessive disorder. A carrier is an individual that appears to be
normal but carries a recessive allele for a genetic disorder. A man and a woman are
both carriers (Aa) for cystic fibrosis. What are the chances a child will have cystic
fibrosis? 1 in 4 (25%)
3. Huntington disease is a dominant disorder. Drina is 25 years old and as yet has no
signs of Huntington disease. Her mother does have Huntington disease (Aa), but her
father is free (aa) of the disorder. What are the chances that Drina will develop
Huntington disease? 50/50 (50%)
4. Phenylketonuria (PKU) is a recessive disease. Mr. and Mrs. Martinez appear to be
normal, but they have a child with PKU. What are the genotypes of Mr. and Mrs.
Martinez? Both are heterozygous (Aa) for the disease.
5. Tay-Sachs is an autosomal recessive disorder. Is it possible for two individuals who
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do not have Tay-Sachs to have a child with the disorder? Yes. Explain. If both parents
are heterozygous carriers (Aa) for the disease each child has a 25% chance of Tay-Sachs.
X-Linked Disorders (LM page 116-117)
Does a color-blind male give his son a recessive-bearing X or a Y that is blank for the
recessive allele? Y
1a. What is the genotype for a color-blind female? XbXb How many recessive alleles does
a female inherit to be color blind? two
b. What is the genotype for a color-blind male? XbY How many recessive alleles does a
male inherit to be color blind? One
2a. With reference to Figure 11.4a, if the mother is a carrier (XBXb) and the father has
normal vision (XBY), what are the chances that a daughter will be color blind? None
b. A daughter will be a carrier? 50/50 (50%) A son will be color blind? 50/50 (50%)
3a. With reference to Figure 11.4b, if the mother has normal vision (XBXB) and the
father is color blind (XbY) , what are the chances that a daughter will be color blind?
none b. A daughter will be a carrier? 100% A son will be color blind? none
X-Linked Genetics Problems (LM page 117)
1.A woman with normal color vision (XBXb) whose father was color blind (XbY), marries
a man with normal color vision (XBY) What genotypes could occur among their
offspring?. Their children could be XBXB, XBXb, XBY, or XbY. What genotypes could occur
if it was the normal-visioned man’s father who was color blind? This means his wife is
not a carrier and since both parents are normal, the children could be only XBX or XBY.
2. Antonio’s father is color blind (XbY) but his mother is not color blind (XBXb or XBXB).
Is Antonio necessarily color blind? no How so? Even if his mother is XBXb he could inherit
the XB. Could he be color blind? Yes How so? If his mother is XBXb he could inherit the Xb.
3. Make up a cross involving hemophilia that could be answered by a Punnett square,
as in Figure 11.4a or b. For example, A normal man reproduces with a carrier female. What
are the chances that a son will have hemophilia. What is the answer to your genetics
problem? For this cross, the answer is 50%.
Multiple Alleles (LM page 118-119)
Experimental Procedure: Using Blood Type to Help Determine Paternity
If a person is AB+, which wells would show agglutination? All three wells.
Table 11.2 Blood Types of Involved Persons
Mother*
Child*
Wanda
Sophia
#1
Blood type BAB+
A*Your instructor may have you confirm these results.
Father ?
#2
B-
#3
AB+
Conclusion (page 119)
1. Noting that only father #3 could have given Sophia the Rh antigen, from whom did
she receive the IB allele? mother From which parent did she receive the IA allele? father
#3. Is there any other possible interpretation to the results of blood typing? No.
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Blood Typing Problems (page 119)
1. A man with type A blood reproduces with a woman who has type B blood. Their
child has blood type O. Using IA, IB, and i give the genotype of all persons involved:
man IA i
woman IBi, and child ii.
2. If a child has type AB blood and the father has type B blood, what could the genotype
of the mother be? IAIA or IAi
3. If both mother and father have type AB blood, they cannot be the parents of a child
who has what blood type? Type O blood
4. What blood types are possible among the children if the parents are IAi X IBi . (Hint:
do a Punnett Square using the possible gametes for each parent.) Types A, B, AB and O.
11.3 Genetic Counseling (LM page 119-123)
Observation: Sex Chromosome Anomalies (LM pages 120)
Label each karyotype in Figure 11.5.
a. Poly X; 3 Xs are present b. Turner; one X is
present c. Klinefelter; X and Y are present d. Jacob; XYY are present
Determining the Pedigree (LM pages 121-–122)
Pedigree Analyses
1. a. Notice that neither of the original parents is affected but several children are
affected. This could only happen if the trait were autosomal recessive..
b. What is the genotype of the following individuals?
Generation I, individual 1: Aa This individual has to be heterozygous because some
of the children are affected.
Generation II, individual 1: aa This individual has to be homozygous recessive
because he is affected.
Generation III, individual 8: Aa This has to be the case because the mother is
homozygous recessive, and the individual has to inherit at least one of her recessive
alleles.
2. a. Notice that only males are affected. This could only happen if the trait were Xlinked recessive.
b. What is the genotype of the following individuals?
Generation I, individual 1: XAXa This female has to be a carrier because she has
an affected son.
Generation II, individual 8: XAX? Unable to determine whether this female is
a carrier or not because she had no children.
Generation III, individual 1: XAY This male is unaffected; therefore, he must have
received a dominant allele.
Construction of a Pedigree (LM page 123)
2. Choose a key for this trait? a = normal eyelashes; A = double row of eyelashes
4. Which pattern is correct? autosomal dominant
5. Use correct genotypes to show a cross between Henry and Isabella and calculate the
expected phenotypic ratio among the offspring:
Aa × aa; 1:1
6. What are the percentage chances of Henry and Isabella having a child with double
eyelashes? 50% Explain why each child has the same chance for double eyelashes.
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Because each child has a 50% chance of receiving either and A or a from their father
(Henry).
LABORATORY REVIEW 11 (LM page 124)
1. If an individual exhibits the dominant trait, do you know the genotype? Why or why not? The
individual has a dominant allele but may also have a recessive allele for the trait.
2. Isabella’s father does not have freckles, but Mary does. What genotypes could Mary’s
mother have? FF of Ff
3. What are the chances two individuals with an autosomal recessive trait will have a child with
this trait? 100%
4. Show a cross that would produce a phenotypic ratio of 1:1 among the offspring. Aa X aa
5. If the parents are heterozygous for cystic fibrosis, what are the chances of a child having
cystic fibrosis? 3:1
6. Tom has blood type AB. Show all possible genotypes for this type blood. IAIB
7. Mary has blood type A and Don has blood type B; can they be the parents of a child with type
O blood? Show why or why not. Yes because Mary could be IAi and Don could be IBi
8. What syndrome is inherited when an egg carrying two X chromosomes is fertilized by a
sperm carrying one Y chromosome? XXY Klinefelter syndrome
9. What is the inheritance pattern in a pedigree if the parents are not affected and a child is
affected? Give a genotype for all persons. Autosomal recessive. Parents are Aa and child is
aa
10. If only males are affected in a pedigree, what is the likely inheritance pattern for the trait?
X-linked recessive. Draw a three-generation pedigree showing the inheritance of the trait from
an affected grandfather to an affected grandson. (No spouses are affected).
Grandfather: XaY Daughter XAXa Grandson XaY
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