Download Lab 2: The cell cycle and The Stages of Mitosis

BIOLOGY SEMESTER ONE
LAB 2
LAB 2: THE CELL CYCLE AND THE STAGES OF MITOSIS
Lab format: this lab is designed for the Remote Web-based Science Laboratory (RWSL)
Relationship to theory: In the textbook (Reece et al., 9th ed.), this lab is related to the following unit: 12.
The Cell Cycle
LEARNING OBJECTIVES
After completing this laboratory, you should be able to:
1. Describe the cell cycle.
2. Identify stages of mitosis from prepared slides.
3. By examining the mitotic region of an onion root tip (Allium cepa), calculate the percentage of
time a cell spends in the various stages of the cell cycle.
THE CELL CYCLE
Mitosis is the process of cellular division that is responsible for cell and tissue growth and development.
For this reason it occurs primarily in areas known as ‘zones of growth’. In plants this happens in the
cambium layers and one of these areas of cell division is located at the root tip.
The root tip of an onion is a location of growth and consists of a terminal quiescent zone, surrounded by
actively dividing cells and covered with a protective root cap. Behind the root cap, three regions are
apparent as the cells begin differentiating: the region of cell division at the tip of the root; behind this
lies the region of elongation; and behind this, the region of differentiation.
Within the region of cell division, you should be able to locate many stages of mitosis taking place.
Cells come into existence through the division of their parent cells and most of the cells divide in turn
producing daughter cells. Usually, this occurs during mitosis, when genetic material is duplicated, and
one copy is passed on to each daughter cell. Mitosis is generally followed by cytokinesis, or cytoplasmic
division, in which the rest of the cell divides in half forming two new cells. Sometimes, however,
cytokinesis does not occur and a cell with many nuclei is formed.
Mitosis is just one part of the cell cycle (see Figure 1)
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FIGURE 1: THE CELL CYCLE
The cell cycle illustrated in the figure above can be summarized as follows:
TABLE 1.1 – A HYPOTHETICAL CELL CYCLE
Phase
Percentage of Time at
Phase
Interphase
(G1, S-phase, G2)
87.5
Mitosis
Prophase
6.9
Metaphase
2.2
Anaphase
1.7
Telophase
1.7
Total
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ACTIVITY 1: OBSERVING THE STAGES OF MITOSIS
(5 marks per drawing- total 20 marks)
Equipment



Unlined paper
Pencil
Computer access (for RWSL microscope)
Procedure
1. Log into the RWSL Microscope. You will be examining a slide of the mitotic region of an onion
root tip (Allium cepa).
2. Make proper biological drawings of cells in each of the four stages of mitosis (see Appendix A).
Drawings should be large (one per page) on a blank (unlined) background. They must be
correctly titled and labelled and include drawing magnification. Please include an explanation
of how you calculated the magnification of your drawing on the back of the page. Figure 2
illustrates the kind of drawing you should submit.
3. Submit completed drawings to your instructor (i.e. by scanning or photographing, then emailing
the image you create; by creating the drawing on a tablet or touch screen; or by mail).
Discussion Questions
1. Why might it be useful to know how much time a cell spends in each phase of mitosis?
2. Can you think of any medical implications for this information?
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FIGURE 2: EXAMPLE DRAWING OF A STAGE OF MITOSIS IN AN ONION ROOT CELL
Onion Root Cell
Allium cepa (* see below)
Metaphase
X1100
*
As mentioned in Appendix A, underline genus and species names in your drawings.
ACTIVITY 2: CALCULATING THE PERCENT TIME SPENT IN EACH STAGE OF MITOSIS
At the time when a slide of an onion root tip was prepared, the cells in the region of cell division were
arrested at their current phase within the cell cycle. Some were fixed at the time of interphase and others
were fixed at some stage of mitosis. The duration of each stage in the cell cycle of the onion root tip can be
estimated by determining the proportion of cells arrested at each stage of mitosis and interphase.
Let’s assume that you examined a slide and determined the stage at which 100 cells were arrested at
time of fixation. Table 1.2 is a summary of your results.
It is known that onion root tip cells take about 16 hours to complete the cell cycle. By determining the
percentage of cells in each stage of mitosis and in interphase, you can calculate the amount of time
spent in each stage. For example, if ten cells out of 100 were found to be in prophase, the percentage of
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cells is 10/100 x 100 = 10%. This shows that any one of the hypothetical cells spends 10% of the time in
prophase, so they spend 0.10 x 16 hours or 1.6 hr (1 hr and 36 min) in that stage.
Procedure
Examine the slide provided of an onion root tip. Count and record the stages of the cycle of each of the
cells in your field of view, then calculate the percentage of cells in each stage, and enter the results in
Table 1 (there is no calculation if you have the patience to count exactly 100 cells; just enter the number
of cells). Keep in mind that you must count enough cells (at least 70 cells) to make a representative
sample. If you count too few, your data will likely NOT reflect the hypothetical estimates provided
earlier. If you are using the highest power ocular, you may have to record more than one field of view.
Calculate the hours and minutes spent in each stage, assuming the entire cell cycle takes 16 hours (0.1 hr = 6
min).
TABLE 1.2 DATA ON STAGES IN PLANT CELLS
Cell Cycle Stage
Number of Cells
% of Total Cells
in the Stage
in the Stage
Hours and
Minutes
in Stage
Interphase
Prophase
Metaphase
Anaphase
Telophase
Total
100
16:00
(Calculations 5 marks)
Discussion Questions
1. How do your data compare to that of the hypothetical times given in Table 1.1? If it is not
exactly the same, discuss some of the reasons this might be. (3 marks)
2. Why is the tip of the onion root chosen for this activity? (1mark)
3. How do the sizes of cells in cytokinesis compare with those in prophase? Why? (2 marks)
4. Compare and contrast mitosis and meiosis. Please include a description about where the
process takes place and the outcome of each type of division. You may use drawings to illustrate
and add to your discussion. (10 marks)
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Bonus:
5. Do the processes of mitosis and meiosis differ between plant and animal cells? Explain.
6. Arrange the following types of cells in order of increasing rates of mitosis: skin, nerve, muscle,
bone. Explain your reasoning.
7. How would you predict the cell cycle of cancerous cells to compare with the cycle in healthy
cells?
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APPENDIX A: USE OF THE COMPOUND MICROSCOPE
BACKGROUND
The microscope is the instrument used in biology to extend the sense of vision to encompass very small
objects. Microscopes were developed during the 17th century and their use revolutionized the way in
which the world was viewed. Biologists discovered that a single drop of water, transparent to the eye,
could be packed with tiny living organisms. In another major discovery, scientists observed that the
tissues of all living things possess a regular, microscopically visible substructure (the units of which came
to be called cells). Biologists today still use fairly unsophisticated microscopes to make observations
about living things. Advances in more sophisticated technology, including scanning and transmission
electron microscopes, have vastly improved our understanding of cell structure and function.
GENERAL CARE OF MICROSCOPES
The compound microscope is a sensitive instrument. It is easily damaged and expensive to repair or
replace. Please handle it with care!
1.
To lift or carry a microscope, always grasp the arm of the microscope with one hand while
supporting the base of the scope with the other hand.
2.
Always set a microscope down on a bench or in its cabinet gently and avoid knocking the
eyepieces or other parts against hard surfaces. Rough handling can damage the alignment of
the lenses.
3.
Always keep the microscope right side up. Many fittings are loosely held in place on the
instrument and could fall off and be damaged if the microscope were turned on its side or
upside down.
4.
Keep the microscope clean. If you spill water or any other fluid on the stage, wipe it up
immediately to avoid corrosion of the stage.
5.
Use lens paper only to clean lenses. Optical glass is very soft and can be scratched by other
types of paper. First, gently blow off loose dust, then gently wipe the lens with clean lens
paper. Avoid polishing the lens vigorously.
6.
Always look from the side when you rotate the objectives, to make sure that the lens does
not hit the slide. A collision could damage the lens, the slide, or both.
7.
Always look from the side when you raise the stage, to avoid raising it too high. If the slide
hits the lens, both could be damaged.
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PARTS OF THE COMPOUND MICROSCOPE
As you read the following description, locate the indicated structures on your microscope.
Microscopes magnify objects through a system of lenses. The compound microscope uses two lenses, so
that double magnification takes place:
i.
The ocular lens is located in the eyepiece. Our microscopes have two eyepieces; that is,
they are binocular. The ocular lens usually provides ten-fold magnification. We call this
"ten-power magnification"; it is usually expressed in written form as 10X.
ii.
The second series of lenses is composed of the objective lenses, mounted on the revolving
nosepiece. Our microscopes have four objectives: a 4X, a 10X, a 40X, and a 100X. Note that
the higher the magnification, the longer the lens. These lenses may also be referred to as
the "scanning" or low-power (4X), medium-power (10X), high-power (40X), and oilimmersion lenses.
The total magnification with which you view an object is the product of the magnification provided by
the ocular and objective lenses. In other words, if you view a cell using a 10X ocular and a 4X objective,
your total magnification is 10  4 = 40X.
The slide to be viewed is placed on the mechanical stage, with the coverslip uppermost. The slide is
placed so that the object to be viewed is directly over the round opening in the stage. Metal arms are
used to hold the slide in place. The slide is moved by using the adjustment knobs that are located at the
side of the stage.
Our microscopes have a built-in light source located in the base of the scope. The light is turned on and
off using the switch on the side of the base; the intensity of the light is controlled using the dial located
next to the on-off switch. The condenser is a system of lenses that concentrates and focuses the light on
the specimen. It is located immediately below the stage and is adjusted by a small knob under the stage
at one side. Below the condenser is the iris diaphragm; it opens and closes to adjust the amount of light
reaching the slide.
At the side of the arm is an adjustment knob for bringing an object on a slide into focus. The large knob
is the coarse adjustment. It permits large upward and downward movement of the stage to bring the
specimen into the range where it is subject to fine focusing. The smaller knob is the fine adjustment. It
permits small upward and downward movement of the stage for final focusing of the image. The
innermost narrow ring is a tension adjustment knob, which must be slackened off before using the
coarse adjustment. It is tightened when using fine focus to prevent “drift”.
USING THE MICROSCOPE
Getting Set Up
1.
Remove the dust cover.
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2.
Plug in the microscope and switch on the light source,adjusting it to about 1/3 maximum
brightness.
3.
Make sure the iris diaphragm is open and raise the condenser until its lens is slightly below
the level of the stage.
4.
Check the lenses for cleanliness. Clean them if necessary.(Use only lens paper.)
5.
Click the low-power (4X) objective into place. (It should be there already if the microscope
was put away properly.)
6.
Lower the stage and place a prepared slide on it, with the coverslip uppermost.
7.
While looking from the side, use the coarse adjustment knob to raise the stage to its highest
position. (Slacken the tension ring first.)
8.
Look into the microscope. Slowly turn the coarse adjustment knob counterclockwise to
lower the stage until the object on the slide comes into view. Then use the fine adjustment
to bring the object into sharper focus.
9.
Whenever you use any binocular microscope for the first time,you should adjust it for your
eyes. This allows both eyes to relax and yet each receive a focused image. Therefore it
reduces eyestrain. Adjust the ocular lenses to suit your own eyes as follows:
(a) With one hand on each side of the microscope, grasp the plate that holds the
eyepieces. Look through the eyepieces and adjust the distance between the ocular
lenses to agree with the distance between your eyes, so that the separate fields
viewed simultaneously by your two eyes merge into one field.
(b) Note the reading on the scale between the oculars. Set the ring on the right eyepiece
to the same reading as noted on the interocular scale.
(c) Close your left eye and use the fine adjustment knob to focus the microscope for your
right eye.
(d) Close your right eye and use the ring on the left eyepiece to bring the specimen into
sharp focus for the left eye.
(e) View with both eyes. The microscope can now be focused for both eyes by using the
normal coarse and fine adjustment knobs.
10.
Adjust the focus of the condenser by moving it all the way up and then lowering it slightly.
11.
Adjust the amount of light passing through the condenser by closing the iris diaphragm all
the way and then opening it as much as necessary while viewing the slide through the
microscope. Use only as much light as is necessary to show maximum detail. If you use too
much light you will miss fine details in the specimen. The iris diaphragm should be adjusted
for every slide and every magnification. You can also vary the intensity of the light by using
the dial beside the on-off switch.
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12.
After you have focused your specimen using low power and made all the necessary
adjustments, you may increase the magnification. Do not use the oil-immersion (100X) lens
unless instructed to do so.
13.
Look from the side and rotate the medium-power (10X) objective into place.
14.
Look through the microscope and adjust the focus. You should only need to use the fine
adjustment knob. Adjust the light.
15.
Look from the side and rotate the high-power (40X) objective into place. Once again, adjust
the focus and adjust the light. You must only use the fine focus adjustment when viewing a
slide under high power. Why? Gently tighten the tension ring to prevent drift. If you lose the
focus, go back to low power to refocus, then return to high power.
16.
After you have finished viewing the slide, lower the stage completely and rotate the 4X
objective into place. Then remove the slide from the stage. Be careful not to hit the slide
against the objective lens. Return the slide to the correct box.
Viewing a Series of Slides
Once the microscope is set up, you can view a series of slides with only a few adjustments.
1.
Always lower the stage and rotate the 4X objective into place before removing one slide and
placing another on the stage.
2.
Focus first with low power, then move to higher magnification. Remember to always look
from the side when you rotate the objectives into place.
3.
Once you have focused with low power, you should only need to use the fine adjustment
knob at higher magnification. Remember, with high power, you must only use fine focus.
4.
Remember to adjust the iris diaphragm and light intensity when you change slides and when
you change magnification.
Putting the Microscope Away
When you finish with the microscope, be sure to do the following:
1.
Rotate the 4X objective into place.
2.
Lower the stage completely.
3.
Remove the slide from the stage.
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4.
Clean all lenses, using lens paper only.
5.
Unplug the microscope and coil the cord neatly at the base of the scope, securing it
with a rubber band if necessary.
6.
Replace the plastic microscope cover.
7.
Return the microscope to a secure cabinet.
NANSLO Biology Core Units and Laboratory Experiments
by the North American Network of Science Labs Online,
a collaboration between WICHE, CCCS, and BCcampus
is licensed under a Creative Commons Attribution 3.0 Unported License;
based on a work at rwsl.nic.bc.ca.
Funded by a grant from EDUCAUSE through the Next Generation Learning Challenges.
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