INTRODUCTION TO
BOTANY
LABORATORY MANUAL
1
Exercise 1
THE COMPOUND MICROSCOPE
I.
Introduction
The microscope is an indispensable tool in Botany as in all biological sciences. It enables us to see
plant structures too small to be seen by the unaided eye. To date, the light and electron microscopes are in
use. Since the invention of the light microscope in the 1590s, there have been numerous improvements and
modifications on the performance of the instrument itself. However, part of this improvement depends on
the development of new techniques for specimen preparation. Regardless of the kind of microscope being
used, three elements are needed to form an image: a source of illumination, the specimen, to be observed,
and a system of lenses to focus the illumination on the specimen and to form the image. The compound
light microscope is a microscope that uses natural or artificial light and a series of lenses: the condenser,
the objectives. The intermediate lenses, and the ocular lens. The objective lens magnifies and projects
virtual image into the body tube while the ocular lens magnifies that image further and projects the enlarged
image into the eye.
II. Objectives
1. To identify the different parts and functions of the microscope.
2. To develop basic skills in the operation of the microscope.
3. To become familiar with measuring microscopic objects.
4. To determine magnification/reduction in the size of objects viewed under the microscope.
5. To become acquainted with preparation of wet or fresh mounts for microscopic examination.
III. Materials
Equipment/Apparatus:
Compound microscopes
Ocular and stage micrometers
Plant Specimens
Leaves
Rice grains
Hydrilla verticillata (digman)
Glassware:
Slides and cover slips
Other materials:
Small letter “e” (newsprint)
Lens paper or cotton with alcohol
Pair of scissors
IV. Procedure
A. Familiarization with Parts of the Microscope
The three basic parts of the microscope are: (a) mechanical parts; (b) magnifying parts, and (c)
illuminating parts.
a. Mechanical Parts (i.e. those parts concerned with the support and adjustment of the optical parts)
Parts
1. Base
2. Pillar
Functions
Stand that supports the microscope.
A short piece of metal that attaches to one end of the base: also supports the
microscope.
2
3. Handle or Arm
4. Inclination Screw
5. Body Tube
6. Ocular Tube or
Draw Tube
7. Revolving
Nosepiece
8. Dust Shield
9. Adjustment Screws
a. Coarse
Adjustment
Screw
b. Fine Adjustment
Screw
10. Stage
11. Mirror Rack
Curved metallic part arising from the pillar used for holding the microscope.
Found at the junction of the pillar and the handle used for tilting the
microscope.
Cylindrical structure vertically arising from the handle; holds the dust shield
and nosepiece.
Upper smaller end of the body tube bearing the eyepiece or ocular lens.
Circular structure where the objectives are attached that permits the shifting
of objectives.
Circular structure above the nosepiece used to protect the lenses of the
objectives.
Two pairs of wheels attached to either side of the body tube. These are:
• Used to adjust the low power objective in focusing.
• Used for delicate focusing in connection with the high power and oil
immersion objectives
A square or round platform with an opening at the center where the slide is
placed.
Found below the stage and attached to the pillar; holds the mirror in place.
b. The Magnifying Parts (i.e. those parts concerned with image enlargement of the specimen)
Parts
1. Ocular or Eyepiece
2. Objectives:
a. Scanner
b. Low Power Objective (LPO)
c. High Power Objective (HPO)
d. Oil Immersion Objective
Functions
Found on the draw tube through which the operator peeps during
actual focusing; usually carries magnification of 10X. A thin,
black line that appears to cut halfway across the field of view
which the student or instructor can use to point out regions of the
specimen under observation is called a pointer.
Tube-like structures attached to the revolving nosepiece.
These are:
- Carries a magnification of 4-5X
- Carries a magnification of 10-12X
- Carries a magnification of 40-60X
- Carries a magnification of 97-100X; requires the use of cedar
oil
c. The Illuminating Parts (i.e. those parts concerned with light provision and regulation to the
specimen.)
Parts
1. Mirror
2. Diaphragm
3. Condenser
Functions
Found below the stage near the base used to collect and direct light
to the specimen.
Found below the stage used to regulate the entry of light onto the
specimen. There are different types of diaphragm: iris, plate, or fan.
Lens found immediately beneath the hole of the stage used to
concentrate light rays on the specimen.
3
B.
Getting to know the parts of the microscope
1. Get the microscope assigned to you. Use both hands to carry the microscope and hold it in an
upright position. With one hand, secure the microscope by its arm and support the base of the
microscope with your other hand.
2. Set the microscope down in a side view position to better locate the other parts.
3. Identify the parts of your microscope and label the drawing in worksheet #1.
C. Use of the microscope
After familiarizing yourself with the parts of your microscope, you are now ready to use the
microscope.
1. Set the microscope down with the arm towards you and the stage away from you.
2. Rotate the nosepiece until the lower the objective clicks into position.
3. Slowly raise the stage or lower the objective by rotating the coarse adjustment until the low power
objective is about 1cm from the stage. When trying to locate the specimen, the low power objective
is always used first before shifting to the high power objective for a detailed study of the specimen.
4. Always relax and keep both eyes open when using the microscope to prevent eye strain.
5. While looking through the ocular, orient the mirror toward a light source so that it reflects the light
up through the open diaphragm, condenser, opening in the stage, and body tube. If this is done
properly, a bright, evenly distributed circle of light called the microscope field will be visible. If the
field is too bright, close the diaphragm to reduce the amount of light reflected from the mirror or if
it is too dim, open the diaphragm.
6. Cut a small letter “e” from the newspaper clippings available in the laboratory. Place the letter “e”
on a glass slide. Cover with a cover slip. Place the slide on the center of the stage.
7. When proper illumination of the field has been achieved, slowly raise the body tube (or lower the
stage) by means of the coarse adjustment knob until the letter come into focus. It is helpful to move
the slide slowly back and forth while doing this in order to help locate the materials being studied.
8. Observe the letter under low power. Make a sketch of what you observe on worksheet.
D. Measurement of microscopic objects using a micrometer eyepiece
Sometimes the biologist needs to know the dimensions of the objects being observed under the
microscope. If you want to estimate the size of microscopic objects more accurately, a microscope
equipped with an ocular micrometer can be used.
An ocular micrometer is a small glass disc etched with uniformly spaced lines (Fig. xx).
When an object is examined under a microscope with an ocular micrometer, the micrometer act as a
ruler that is superimposed on the object being measured. However, the units of distance between the
spaces of the ocular micrometer is arbitrary. Therefore, before an ocular micrometer can be used for
actual measurements of microscopic objects, it must first be calibrated against a standard. You need a
4
stage micrometer for calibration. A stage micrometer is a glass with scribed lines that are exactly
0.01mm (or 10 micrometers) apart (Fig. xx).
Figure xx. Ocular Micrometer
Figure xx. Stage micrometer
Calibration of an ocular micrometer
1.
2.
3.
4.
5.
6.
7.
8.
Get an ocular and a stage micrometer. These are delicate instruments and must be protected from dust,
grease, fumes, and friction. These micrometers must be handled at the edges.
How to attach an ocular micrometer to the microscope:
• Detach the ocular tube from the draw tube
• Unscrew the eyepiece lens (the top lens for some microscopes but for Zeiss microscopes, it is
lower part of the eyepiece) of the ocular tube. Very carefully insert the ocular micrometer.
• Put back the eyepiece lens and insert the ocular tube back into the draw tube.
Mount the stage micrometer. Center its scale and focus with the low power objective.
While looking through the low power objective, rotate the ocular and adjust the stage micrometer until
the lines of the ocular micrometer are parallel to those of the stage micrometer.
Match the lines at the left edges of the two micrometers by moving the stage micrometer.
Observe another point where the lines of the two micrometers coincide again.
Now count the number of spaces of the stage micrometer included within a given number of spaces in
the ocular micrometer.
Since the smallest spaces on the stage micrometer are known to be 0.01mm apart, compute the distance
of 1 ocular division or calibration constant as follows:
1 ocular division =
!"#$%& () *+,-% #* . /./1 ##
!"#$%& () (2"3,& #*
where: ms = micrometer spaces
What is the calibration constant of the microscope you are using? Show your computation on the
worksheet.
10. You may now remove the stage micrometer and replace it with the slide containing the specimen that
you want to measure.
9.
5
11. The calibration constant is used to determine the length or width of the microscopic specimen that you
are measuring. Multiply the number of divisions of the micrometer eyepiece covered by the specimen by
the calibration constant.
Length or width = number of ocular ms x calibration constant
12. Prepare a wet mount of a Hydrilla leaf and measure the diameter of a tooth cell.
E.
Preparation of a wet mount
In Botany, you will be asked to make a wet or a fresh mount to facilitate viewing some plant parts.
1.
2.
3.
4.
Use forceps or tweezers to pick off a young, leaf from a portion of the digman stem.
Place the leaf on the center of a clean glass slide.
With a dropper, put a drop of water over the leaf.
Get a cover slip and put one edge of the cover slip down on one side of the drop of water. Then
with a dissecting needle, slowly bring the cover slip down the water. If air bubbles form, gently tap
the cover glass. Bubbles interfere with viewing of the specimen.
5. Focus under the LPO of the microscope. Measure the diameter of a tooth cell which is located along
the sides of the leaf. What is the diameter of the tooth cell? Show you computation on the worksheet.
If you are asked to make wet mounts of roots, stems, or fruits, take a sharp blade and make thin
sections of the specimen. Place these sections on a glass slide and proceed as above in number 4.
F.
Computation of magnification
Magnification may be defined as the number of times an object is enlarged by the magnifying lens
or the number of times a drawing is enlarged or reduced from the original size of the object.
1. To determine the total magnification of the specimen as seen through the microscope, multiply the
magnification of the ocular by the magnification of the objective.
Total Magnification = magnification of eyepiece x magnification of objective
2. To determine the magnification in a drawing made, divide the size of drawing, by the actual size of
object or specimen.
Magnification of drawing =
456% () 7&,859:2+",3 *56% () ($;%2+
3. Compute for the magnification of the length of a leaf, length of rice grain and diameter of tooth cell
drawn 5 cm long.
4. Show your computation on the worksheet.
6
Name _______________________________________
Course/Year/Section ___________________________
Date Performed ___________________
Date Submitted ___________________
WORKSHEET NO. 1.
THE COMPUND MICROSCOPE
A. Label the parts of the microscope below:
7
B. Using the microscope.
In the box provided below, sketch the letter e using the LPO. Sketch the letter e using the HPO.
1. Is the position of the letter e the same as when viewed with naked eye or is it inverted?
_______________________________________________________________________________
2. When you move the slide to the left, in which direction does the letter image appear to move?
_______________________________________________________________________________
3. When you move the slide toward you, in which direction does the letter image appear to move?
_______________________________________________________________________________
4. Explain your observations.
5. When you shift to HPO, what happens to the letter image and field of view?
_______________________________________________________________________________
C. Measurement of microscopic objects using a micrometer eyepiece.
Calibration constant (computation) Diameter of the tooth cell: ______________________________
8
D. Magnification
Specimen
Actual Size
(Show computation)
Hydrilla tooth cell (diameter)
Length of a leaf
Rice Grain (length)
9
Magnification in a drawing made
Exercise 2
THE CHEMICAL COMPONENTS OF A PLANT CELL
I.
Introduction
One of the unifying characteristics of living things is that of organization. All living things are
organized and structured from molecules to individuals and communities. What are plant cells made
of? Plant cells are made up of inorganic and organic compounds. In general, molecules that contain
carbon are referred to as organic compounds while pure carbon components is lacking hydrogen are
classified with non-carbon compounds as inorganic chemicals. Plants as well as other organisms
consist of organic compounds such as carbohydrates, proteins, lipids, and nucleic acids. The
inorganic components of plants include a variety of elements such as carbon, hydrogen, and oxygen.
These can be classified into three groups: monosaccharides, disaccharides, and polysaccharides.
Starch, which is a polysaccharide, is regarded as the major food reserve in plants.
Proteins are made up of basic units called amino acids. The composition and size of the protein
molecule depends upon the kind and number of its amino acid subunits.
Lipids of fats are a heterogeneous collection of substances which are characterized by their low
solubility in water but higher solubility in organic solvents such as acetone and ether.
II.
Objectives
By the end of the exercise you should be able to know how to test for the presence of carbohydrates,
proteins, and lipids.
III.
Procedure
A.
Qualitative Test for Carbohydrates
A.1.
Benedict’s Test for Reducing Sugars
Carbohydrates are molecules made of C, H, and O in a ratio of 1:2:1 (e.g., the chemical formula for
glucose (C6H12O6). Carbohydrates are made of monosaccharides, or simple sugar. Paired
monosaccharides form disaccharides. Sucrose is a disaccharide of glucose linked to fructose.
Similarly, linking three or more monosaccharides form a polysaccharide, such as starch, glycogen,
or cellulose.
Many monosaccharides such as glucose are reducing sugars, meaning that they possess free aldehyde
– CHO or ketone -C=O groups that reduce weak oxidizing agents such as the copper in Benedict’s
reagent. Benedict’s reagent contains cupric (copper) ion complexed with citrate in alkaline solution.
Benedict’s test identifies reducing sugars based on their ability to reduce cupric (Cu) ions to cuprous
oxide at basic (high) pH. Cuprous oxide is green to reddish orange. A green solution indicates a small
amount of reducing sugars, and reddish orange indicates an abundance of reducing sugars. Nonreducing sugars such as sucrose produce no change in color (i.e. the solution remains blue).
10
Procedure:
1.
2.
3.
4.
Obtain 7 test tubes and number them 1 to 7.
Add to each of the first three test tubes the materials to be tested: (onions, potato cubes, ripe
fruit), to the 4th test tube, glucose, to the 5th test tube, sucrose, to the 6th test tube, starch solution,
and to the 7th distilled water. Add 2 ml Benedict’s solution to each tube.
Place all of the test tubes in a boiling water bath for 3 minutes and observe color changes
during this time.
After 3 minutes, remove the tubes from the water bath and let them cool to room temperature.
Record your observations (i.e. the color of their contents) in worksheet #2.
A.2. Iodine Test for Starch
Staining by iodine (iodine-potassium iodide, IKI) distinguishes starch from monosaccharides,
disaccharides and other polysaccharides. The basis for the test is that starch is a coiled polymer
of glucose, and iodine interacts with these coiled molecules and becomes bluish black. Iodine
does not react with carbohydrates that are not coiled and remains yellowish brown. Therefore
a bluish black color is a positive test for starch, and a yellowish brown color is a negative test
for starch. Notably, glycogen a common polysaccharide in animals, has a slightly different
structure than does starch and produces only an intermediate color reaction.
Procedure:
1.
2.
3.
4.
B.
Obtain 7 test tubes and number them 1 to 7.
Add to each test tube the same materials as in A.1.
Add 3-5 drops of iodine to each tube.
Record your observations (i.e. the color of the tubes’ content) in worksheet #2.
Qualitative test for Proteins
Proteins are made of amino acids, each of which has an amino acid (-NH) and carboxyl (-COOH)
group. The bond between the two groups found on adjacent amino acids in a protein is a peptide bond
and is identified by a Biurest test. Specifically, peptide bonds in a protein complex with the Cu in
Biuret reagent and produce a violet color. A Cu must combine with four to six peptide bonds to
produce a color, therefore, free amino acids do not react positively. Long chain polypeptides
(proteins) have many peptide bonds and produce a positive reaction. A violet color is a positive
test for the presence of protein; the intensity of color relates to the number of peptide bonds reacting.
B.1 Biuret Test
Procedure:
1.
2.
3.
4.
Obtain five test tubes and number them 1 to 5. To the first test tube add egg albumin, to the
second test tube a slice of fruit, to the fourth, sliced bean seeds and to the fifth, distilled water.
Add 2 ml of 2.5% sodium hydroxide (NaOH) to each tube.
Add 3 drops of Biuret reagent to each tube and mix.
Record your observations (i.e. the color of the tubes’ content) in worksheet #2.
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B.2. Xanthoproteic Test
The Xanthoproteic test can be used in the absence of Biuret Reagent. To each test tube add carefully
1 ml of concentrated nitric acid and place in boiling water bath for 5 minutes. A white precipitate
forms which upon heating turns yellow and finally dissolves, giving rise to a yellow solution. Cool
the solution and add 8-10 ml of 10% NaOH to make the solution definitely alkaline. The yellow color
deepens to orange. Record results in worksheet #2.
C.
Qualitative Tests for Lipids
Lipids include a variety of molecules that dissolve in non-polar solvents such as ether and acetone
but not in polar solvent such as water. Triglycerides are abundant lipids that are made of glycerol and
three fatty acids. Tests for lipids are based on a lipid’s ability to selectively absorb pigments in fatsoluble dyes such as Sudan IV.
C.1
Sudan Test
Procedure:
1.
2.
3.
4.
5.
Obtain five test tubes and number them 1 to 5.
To the following test tubes place: albumin, honey, slices of avocado fruit, bean seeds, and
cooking oil.
Add 3 ml of water to each tube.
Add 5 drops of Sudan IV to each of the remaining tubes. Mix the contents of each tube.
Record the color of the tubes’ contents in worksheet #2.
C.2. Grease-spot Test
A simple test for lipids is based on their ability to produce translucent grease-marks on unglazed
paper.
Procedure:
1.
2.
3.
4.
5.
6.
IV.
Obtain one piece of writing paper.
Add a drop of cooking oil near one corner of the paper and a drop of water near the opposite
corner.
Let the fluids evaporate.
Look at the paper as you hold it up to the light.
Test in a similar way the other materials used in the Sudan IV test.
Record your observations in worksheet #2.
Chemicals/Materials:
10% NaOH, 2.5% NaOH
Concentrated nitric acid
Albumin
Sudan IV
Iodine-potassium iodide
Vegetable oil
Glucose solution
Starch solution
Sucrose solution
Honey
Potato
Onions
Bean seeds
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Name _______________________________________
Course/Year/Section ___________________________
Date Performed ___________________
Date Submitted ___________________
WORKSHEET NO. 2.
THE CHEMICAL COMPONENTS OF A PLANT CELL
Table 1. Some Qualitative Reactions of Carbohydrates
1.
2.
3.
4.
5.
6.
7.
Materials Tested
Onions
Potato cubes
Ripe banana
Glucose
Sucrose
Starch solution
Distilled water
1.
2.
3.
4.
5.
6.
7.
Onions
Potato cubes
Ripe banana
Glucose
Sucrose
Starch solution
Distilled water
Test
Observation
Benedict’s
Iodine
Questions:
1. What kind of carbohydrates exhibited the positive reaction to Benedict’s test? Iodine test?
_____________________________________________________________________________________
_____________________________________________________________________________________
_____________________________________________________________________________________
2. What reactions occur which comprise the basis of the positive color reactions involved in Benedict’s
test? Iodine test?
_____________________________________________________________________________________
_____________________________________________________________________________________
_____________________________________________________________________________________
3. An unknown sample gives a positive reaction to Benedict’s test but fails to give a positive reaction to
Iodine test. What possible explanation can you offer and how will you go about testing your hypotheses?
_____________________________________________________________________________________
_____________________________________________________________________________________
_____________________________________________________________________________________
_____________________________________________________________________________________
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4. Explain the significance of carbohydrates in plants.
_____________________________________________________________________________________
_____________________________________________________________________________________
_____________________________________________________________________________________
Table 2. Some Qualitative Reactions of Proteins
Materials Tested
1. Egg Albumin
2. Honey
3. Ripe banana
4. Sliced bean seeds
5. Distilled water
1. Egg Albumin
2. Honey
3. Ripe banana
4. Sliced bean seeds
5. Distilled water
Test
Observation
Biuret
Xanthoproteic
Questions:
1. Which of the substances tested gave a positive reaction to the Sudan IV test? Why?
_____________________________________________________________________________________
_____________________________________________________________________________________
_____________________________________________________________________________________
2. Which of the substances tested gave a positive reaction to the grease spot test? Why?
_____________________________________________________________________________________
_____________________________________________________________________________________
_____________________________________________________________________________________
3. Differentiate fatty acid, glycerol, and fat.
_____________________________________________________________________________________
_____________________________________________________________________________________
_____________________________________________________________________________________
4. Discuss the role of lipids in plants.
_____________________________________________________________________________________
_____________________________________________________________________________________
_____________________________________________________________________________________
_____________________________________________________________________________________
14
Exercise 3
THE PLANT CELL
I.
Introduction
The study of plants starts with the study of the basic unit – the plant cell. Plant cells compose the
plant body and the workings of these cells result in the complex functioning of a plant. How do
plant cells look like? Do all plants look similar? What parts of the cell can be seen under the
microscope? How can these parts contribute to the overall function of the cell? What laboratory
methods can be used to make the study of plant easier? These are some of the questions we will
answer in today’s exercise.
II.
Objectives
At the end of the exercise, you will be able to:
1. Identify a plant cell and distinguish the basic parts of the plant cell.
2. Develop skills in wet mount preparation and use stains to facilitate the study of living plant parts.
III.
Materials
Equipment/Instruments:
Microscopes
Reagents and other materials:
1. Glass slide and cover slip
2. Scalpel or blade
3. Droppers
4. IKI solution
Plant specimens:
1. Allium cepa (onion bulb)
2. Sanseviera zeylanica (buntot-tigre)
3. Hydrilla verticillata ( digman)
4. Ficus sp. (Indian rubber tree)
5. Solanum tuberosum (potato)
6. Rhoeo discolor (Bangka-bangkaan)
7. Portulaca oleracea (gulasiman)
8. Begonia sp. (begonia)
9. Musa paradisiaca (banana fruit)
10. Dieffenbachia sp. (dumbcane)
11. Coleus sp. (mayana stem)
12. Commelina sp. (alikbangon)
13. Ipomea aquatica (kangkong)
14. Ixora sp. (santan leaf)
15. Phaseolus vulgaris (mongo seed coat)
16. Luffa acutangula (patola)
17. Wood fragments
18. Cocos nucifera (coconut husk)
15
IV.
Procedure
A.
Examine a living non-green cell from the onion skin
1. Prepare a wet mount of Allium cepa (onion) skin or epidermis by peeling off a portion of the
epidermis from the concave surface of the onion bulb scale. Put the specimen on a glass slide,
add a drop of water, slowly put a cover slip, and examine under the microscope. Use first LPO
then shift to HPO. Draw what you see on the worksheet and answer the questions.
2. Stain the specimen by placing one or two drops of iodine solution at the edge of the cover slip.
Draw out the iodine solution under the cover slip by holding a piece of absorbent tissue at the
opposite edge of the cover slip. Label your drawing and answer the questions on your
worksheet.
B.
Examining the living green cell
Take a young green leaf from the tip of Hydrilla verticillata (digman). Put it on a slide and add
a drop of water. Place a cover slip. Focus under LPO on the side of the leaf (or the margins) where
it is not very thick. Here you can see a few layers of cells and some cellular details. Draw and
label what you see. Answer the questions on your worksheet.
C.
Examining other colored bodies in a cell
1. With a blade, place a small thin piece of the pulp (not the skin) of a ripe tomato on a slide with
a drop of water. Study your preparation at its thinnest part. Adjust the iris diaphragm of your
microscope to a darker phase. This will enable you to view the colored bodies. Draw what you
observe and answer the questions on your worksheet.
2. With a blade or scalpel, scrape some fragments from a cut potato onto a slide. Place a drop of
water and a cover slip. After observing under LPO and HPO, add a drop of iodine solution to
the slide of the preparation and draw out from the slide of the cover slip. Examine your
preparation. Draw what you observe and answer the questions on your worksheet.
D.
Examining a vacuole and its contents
Plant cells contain vacuoles which are fluid-filled spaces not occupied by the cytoplasm. In young
cells, these are often small and scattered; in older, they are often large and fill most of the cell.
Vacuoles contain substances which are used by plants.
1. Make a thin section of the violet portion of the leaf of Rhoeo spathacea (Bangka-bangkaan).
Add a drop of water and put a cover slip. Examine under LPO and focus on the violet areas.
Shift to HPO and study your specimen. Answer the questions on your worksheet.
Vacuoles also contain ergastic substances like crystals. Crystals may be composed of calcium
oxalate or calcium carbonate. These plant crystals come in various shapes. The commonly
known crystals are the prismatic (diamond shape), druse (star-like), rosette (star-like with
many points), raphide (needle-like), cystolith (like a cluster of grapes).
2. Make thin sections of the specimens listed on your worksheet and look for crystals. Draw the
crystals you see and identify the type present on your worksheet.
16
E.
Examining various types of cells
Did the cells you examined look the same? No, since these cells vary in size, shape, and function.
Their work or function determine how they look like; what cell parts are more developed than
others. There are three basic cell types found in plants. These are parenchyma, collenchyma, and
sclerenchyma.
1. Parenchyma cells. Most of the cells you examined above are parenchyma cells. These cells
have thin cell walls, and the protoplast is alive at maturity. They may have various functions
such as making food, storing food, or developing into other specialized function. Make thin
cross-sections of the specimens listed on the worksheet and observe under the microscope.
Look for cells with thin walls. Draw the specimens you see on your worksheet.
2. Collenchyma cells. Collenchyma cells are found in groups along the sides of the young stems
and in the stalk or midrib of leaves. The walls of these cells are unevenly thickened and the
thickening occur at the corners where the cells meet. The protoplast is alive at maturity just
like the parenchyma. The cells are also somewhat elongated. They have various functions
although most of them function for support and for strength of a young plant or support of a
leaf. Make thin cross-sections of the plants listed on the worksheet and observe under the
microscope. Look for cells in groups which have unevenly thickened walls. Draw the
specimens you see on your worksheet.
3. Sclerenchyma cells. Sclerenchyma cells have very thick walls and differ from the first two
types of cells because these cells die at maturity. The thick walls are impregnated with lignin.
The common sclerenchyma cells are the fibers and sclereids or stone cells. Cell walls of
sclerenchyma cells absorb stains so that they are often very visible in prepared slides. Since
the protoplast is dead, these cells look empty. These cells usually function for transport of
water and for support. Some cells also protect the more delicate parts of the plant. Draw the
specimens you see on your worksheet.
17
Name _______________________________________
Course/Year/Section ___________________________
Date Performed ___________________
Date Submitted ___________________
WORKSHEET NO. 3.
THE PLANT CELL
I.
The Parts of the Plant Cell
A. Non-green epidermal cell of Allium cepa (onion):
a. Do you see rectangular boxes adjacent to each other?
__________________________________________________________________________
b. Each “box” and its contents compose the cell/ What part of the cell forms the “box”?
__________________________________________________________________________
c. Can you see an inner “envelope’ surrounding the cell?
__________________________________________________________________________
Can you find any communication lines between the boxes? Why or why not?
__________________________________________________________________________
__________________________________________________________________________
__________________________________________________________________________
d. Do you think Robert Hooke was correct in giving the name “cell” (meaning “small rooms”)
to the specimen he saw in cork? Support your answer.
__________________________________________________________________________
__________________________________________________________________________
__________________________________________________________________________
e. What part of the cell becomes visible after the addition of the iodine stain?
__________________________________________________________________________
f. What is the function of this part of the cell?
__________________________________________________________________________
g. Look at the cell wall. What is responsible for the organized arrangement of cells?
__________________________________________________________________________
__________________________________________________________________________
__________________________________________________________________________
18
h. What is the cementing substance between cells called? What is the chemical composition of
this substance?
__________________________________________________________________________
__________________________________________________________________________
__________________________________________________________________________
i. Do you think Robert Browne was right when he gave the name “nucleus” to the “nut-like” part
he saw inside the cells? Support your answer.
__________________________________________________________________________
__________________________________________________________________________
__________________________________________________________________________
j. Do you see darker stained part/s inside the nucleus? What is/are this/these?
__________________________________________________________________________
__________________________________________________________________________
__________________________________________________________________________
B.
Green cell of Hydrilla verticillata
a. What are the green round bodies that you see?
________________________________________________________________________
b. What are the functions of the green bodies?
__________________________________________________________________________
c. About how many are there?
__________________________________________________________________________
d. Are those green bodies moving or stationary?
__________________________________________________________________________
e. If the green bodies are moving, what do you call this movement? What is the direction of this
movement?
__________________________________________________________________________
f. Is the nucleus visible among the green bodies? Why or why not?
__________________________________________________________________________
__________________________________________________________________________
C. Colored bodies in a cell
1.
Lycopersicon esculentum (Tomato) cells:
a. Do you see orange or red round or oblong bodies in cells?
________________________________________________________________________
b. What are they called?
________________________________________________________________________
c. How would you compare the arrangement of these cells to the cells of the onion skin?
________________________________________________________________________
________________________________________________________________________
d. Aside from tomato, what other plants contain these orange or red bodies?
________________________________________________________________________
________________________________________________________________________
e. What are the functions of these orange or red colored bodies?
________________________________________________________________________
________________________________________________________________________
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2.
Solanum tuberosum (Potato) scrapings:
a. What do you call the bodies stained by the iodine solution?
________________________________________________________________________
b. Are these bodies of the same sizes?
________________________________________________________________________
c. What are the functions of these bodies?
________________________________________________________________________
D. The Vacuole
a. What colored substance or pigment is found inside the vacuole?
________________________________________________________________________
b. In what solvent is this pigment dissolved?
________________________________________________________________________
c. What is the membrane of this structure called?
________________________________________________________________________
d. What is the function of a vacuole?
________________________________________________________________________
Type of Crystals in various specimens
Specimen
1. X-section of
Begonia stalk
Drawing of Crystal
2. X-section of
Rhoeo discolor
leaf
3. X-section of
Portulaca
oleracea
4. X-section of
Sanseviera
zeylanica leaf
5. X-section of
Ficus sp. leaf
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Type of Crystal
Questions:
1.
How are the crystals formed?
__________________________________________________________________________________
__________________________________________________________________________________
__________________________________________________________________________________
2.
What is the chemical composition of crystals?
__________________________________________________________________________________
__________________________________________________________________________________
__________________________________________________________________________________
3.
What is the importance of crystal formation in plant cells?
__________________________________________________________________________________
__________________________________________________________________________________
__________________________________________________________________________________
E. Fundamental Cell Types
1. Parenchyma Cells
Specimens
Scrapings of Musa sp. (banana) fruit
Drawing of Parenchyma cells
X-section of Dieffenbachia sp. leaf midrib
X-section of Commelina benghalensis stem
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2. Collenchyma cells
Specimens
1. X-section of Coleus blumei young stem
Drawing of Collenchyma cells
2. X-section of Ipomea aquatica leaf stalk
3. X-section of Ixora sp. leaf midrib
3. Sclerenchyma cells
Specimens
Drawing of Sclerenchyma cells
Mongo seed coat
Luffa fibers
Wood fragments (macerated)
Coconut husks
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Questions:
1. What organelles did you see in the specimens?
____________________________________________________________________________
2. Differentiate a cell wall from a plasma membrane structurally and functionally.
____________________________________________________________________________
____________________________________________________________________________
____________________________________________________________________________
3. In a living cell, could the nucleus occur in the vacuole? Why or why not?
____________________________________________________________________________
____________________________________________________________________________
4. How do stains facilitate the study of cells?
____________________________________________________________________________
____________________________________________________________________________
5. What is the advantage of using fresh mounts in the examination of plant cells?
____________________________________________________________________________
____________________________________________________________________________
____________________________________________________________________________
23
Exercise 4
MITOSIS
I.
Introduction
Cell division is an important component of the growth of plants. It is a way of increasing the number
of cells in the plant body. In plants, this division of cells occurs in special tissues called meristems.
This cell division results in daughter cells that are exactly like the parent cells. There are two phases
of this division: mitosis (or nuclear division) followed by cytokinesis (or cytoplasmic division).
Mitosis also enables the plant to replace dead cells or to heal wounds.
II. Objectives
At the end of this exercise, you will be able to:
1. Identify the prophase, metaphase, anaphase and telophase, and distinguish the mitotic stages from
interphase.
2. Determine the duration of the different mitotic stages in the onion root tip.
3. Determine the mitotic index of the onion root tip.
III. Materials
Equipment: Microscope
Plant Specimen: Prepared slide of the Allium cepa (onion) root tip
IV. Procedure
A. Identification of the Stages of Mitosis
Examine a prepared slide of the Allium cepa (onion) root tip under the LPO. Locate the root cap,
the tip most portion of the root apical meristem. Focus on the area just behind the root cap. Most
of the cells in this part of the root tip are almost square in shape and many cells are dividing. Shift
to HPO. Select cells that show the features listed on the worksheet and indicate interphase as well
as different stages of mitosis. Answer the questions on your worksheet.
B.
Determination of Duration of Mitotic Stages
Center the most mitotically active region of the onion root tip in your field of view under high
power objective. Once centered, secure your slide and do not move the slide.
1.
Estimate the total number of cells in the field of view. Count all the cells in one segment of
the field, for example, one quadrant, then multiply this count by the proportion of the total
field not counted. For example, if you counted the cells in ¼ of the field, the total field would
contain approximately 4 times this number of cells. How many cells are there in your field
of view? Show your calculations on the worksheet.
2.
Count the number of cells in the recognizable stages of division and add these tallies to the
appropriate column on the worksheet. Estimate the number of interphase cells by
subtracting the number of cells in all other stages from the total number of cells in the field.
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The duration of the cell cycle in a typical onion root tip is about 16 hours. We can estimate
the duration of time spent by an individual cell in each stage of mitosis by the proportion of
cells in that stage at the instant the tissue was fixed. Use the following formula to calculate
the duration of each stage, in minutes.
Duration of stage = number of cells at stage/total number of cells x 16 hrs./cycle x 60
minutes/hour.
Calculate the duration of each stages (min./cell cycle) and enter this into the table on your
worksheet. Get also the class data.
Published values for prophase, metaphase, anaphase, and telophase respectively are 7.1, 6.5,
2.4 and 3.8 minutes.
Answer the questions on your worksheet.
C.
Determination of the Mitotic Index
1.
Examine under HPO a prepared slide of the longitudinal sections of root tips of onion (Allium
cepa). Locate the different stages of mitosis in cells near the region of formation of the root
tip and try to recognize the different stages of mitosis.
2.
Determine the proportion of the cells in the different stages of mitosis (mitotic index)
observed in 1 microscopic field at this region and fill in the table in the worksheet. Answer
the questions on your worksheet.
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