Exercise 2
THE PLANT CELL
In beginning to examine in the internal structure of a cell requires a light microscope of sufficient quality to
magnify (i.e., increase the apparent size of the object) with adequate resolution (i.e., ability to visually separate two
objects that are close to each other), and contrast (i.e., proper lighting conditions so that objects can be seen more
clearly and not appear washed out against the brightness or hidden in shadows by the darkness). Proper use of the
microscope is necessary to make the most use out of this instrument so that you can maximize its capabilities and
usefulness. Your lab instructor will discuss the parts and proper usage of the light microscope before you
begin this laboratory exercise.
Compare what you see in lab with the figures given in the photographic
atlas (i.e., Figure 1.1).
I. PREPARING A WET MOUNT
A. ELODEA
Unless you are using a prepared slide, most of your microscopic work will require the use of a wet mount (a
slide which you prepare yourself by placing the object in a drop of water on a slide and adding a cover slip). Except for
very elementary observations, all objects viewed through the microscope with transmitted light must be mounted in
some medium. Obtain a clean, dry slide and cover slip from the supply table. With forceps, remove a young leaf from
the actively growing tip of Elodea. Place the leaf in a small drop of water on your slide and carefully add the cover
slip. You may find it best to hold the cover slip by the edges, touch it to the slide, and then move it toward the drop of
water at an angle of about 45 degrees. As the lower edge of the cover slip contacts the water, gently lower it to the
slide. If you have done this carefully, you should have the Elodea leaf mounted in the center of your slide in just
enough water to fill the space between the slide and cover slip and there should be no entrapped air bubbles. Your
instructor will demonstrate the proper procedure.
Study your wet mount under low power and then select an area near the tip about mid-way between the
margin and the mid-rib. Using higher magnification, carefully focusing, and changing diaphragm settings. What
structures do you see? In some cases you may see movement of organelles and cytoplasm within a living Elodea leaf
cell. This movement is known as cyclosis or cytoplasmic streaming.
Compare what you see in lab with the figure given in the
photographic atlas (i.e., Figure 1.5).
B. ONION EPIDERMAL CELLS
The outermost or surface layer of cells in plants is referred to as the epidermis. In most cases the epidermis is
composed of only a single layer of cells and can frequently be removed by the “peel technique”. Your instructor will
demonstrate this process. Following this procedure, prepare a wet mount of a small piece of epidermis from the inner
side of an onion bulb scale. Mount the tissue so that it is not folded and so that the surface of the epidermis is toward
the top in your slide. Examine your preparation under the microscope and notice the general shape, size, and
arrangement of the cells. Switch to high power and regulate the diaphragm setting so that you can see clearly the cell
wall and the living protoplast with composed of the cytoplasm and the nucleus ( plural, nuclei). If you look closely
you should also be able to see a nucleolus (plural, nucleoli) within the nucleus of some cells.
By careful focusing and light adjustment you may be able to discern the central vacuole containing the
so-called cell sap (water and dissolved substances).
Thus far you have been observing a living cell and have probably noted that some of its features are difficult
to observe In order to enhance details of the cell contest, place a drop of iodine stain at one edge of the cover slip. With
a small piece of paper towel gently touch the opposite edge of the cover slip in order to “drop” the iodine under the
cover slip. Now observe your preparation.
Compare what you see in lab with the figure given in the photographic
atlas (i.e., Figure 1.3). For a detailed list of the parts of a “typical plant
cell” check out Table 1.1 on page 2 in the photographic atlas.
II.
PERMANENT SLIDES
During this semester you will be examining a wide variety of plant materials with your microscope. Much of
this material will be in the form of permanent mounts, in which the tissue is cut into very thin slivers, stained to bring
out finer details or differentiate tissues, and embedded in a permanent mounting medium. In order to produce a
permanent slide of good quality a long tedious process must be followed that may extend over a period of several days
or even weeks. The main steps in this process are usually as follows:

A small piece of tissue such as a portion of a root, stem, or leaf is placed in a special solution to “kill” and
“fix” cellular detail as nearly as possible to the living state.

The tissue is dehydrated by passing it through a progressively stronger series of alcohols.

Alcohol is then replace with a solvent, which in turn is replaced by melted paraffin (embedded).

After cooling, the paraffin (which now contains the tissue) is cut into thin sliver by a precision cutting device
called a microtome.

Sections are affixed to a slide and the paraffin is then removed with solvents.

Slides are passed through one or more stain series to enhance details and differentiate tissues within the
section.

Finally, after another dehydration sequence, a drop of special permanent mounting medium is place on the
specimen and cover slip added.
As you can see, a great deal of time, effort, and skill is required to produce a permanent slide. Therefore,
these slides are often quite expensive and should be handle with extreme care.
So far, you have gained experience in using the compound microscope and examined, among other things,
the living cells in a leaf of Elodea. The leaves of plants, as well as all other parts of plants, are composed of cells or
cell products. Although we often study the plant as an individual organism or as an intact unit, it is very important to
understand as much as possible about individual cells since they are the fundamental structural and functional
components of the plant as a whole. As you might suppose, plant cells vary widely in size, shape, origin chemical
composition, function, and in almost every other conceivable way. As a matter of fact, it is probably safe to say that no
two cells are exactly alike. During this exercise we will study some of this great variation and diversity among plant
cells.
III. OTHER PLANT CELL STRUCTURES
A. CHROMOPLASTS
Prepare a wet mount of a small amount of macerated red or yellow pepper fruit pulp and examine it under your
microscope. Locate an intact cell. How do these cells compare in size and shape to the onion epidermal cells?
Notice the large number of chromplasts located in the cell. In most preparations these plastids will be seen all over the
slide since the thin-walled cells are easily rupture thus releasing the cell contents. The primary pigments present in
chromplasts are fat-soluble carotenoids? The presence of chromoplasts in the cytoplasm and the pigments these
possess impart the color seen in many fruits and flowers.
B. LEUCOPLASTS
Another type of plastid found in many plant cells is the leucoplast (a food-storage plastid containing starch or
oil). Leucoplasts that contain starch are called amyloplasts. Those that contain oil are known as elaioplasts. Prepare
a wet mount of a small amount of tissue scraped from the cut surface of a potato. Examine your preparation under low
and then medium power. In this slide most of your potato cells will have ruptured and released the amyloplasts but
you may be able to locate a few intact cells. Now add a drop of iodine to the edge of the cover slip and draw it under
as you did earlier.
What color are the leucoplasts (amyloplasts with starch grains) now? _____________________
Compare what you see in lab with the figure given in the photographic
atlas (i.e., Figure 1.6).
Under high power focus on a single starch grain and manipulate the light and fine adjustment until you observe
maximum clarity and detail. Are concentric rings or layers evident in the starch grain? If so, these rings represent the
daily deposition of starch from one day to the next that were stored between extended periods of photosynthesis (day
time) and periods when food could not be manufactured (night time).
C. CRYSTALS
Prepare a hand cross section of the stem of Zebrina or Transcantia, or some available herbaceous plant. In making
hand sections it is very important that you use a sharp razor blade and cut the section as thin as possible. Your
instructor will demonstrate the technique of making hand sections but do not be discouraged if your first few attempts
are not successful. After you have cut what appears to be a good section, prepare a wet mount and examine under low
power. In your preparation you will see (in cross section) a great variety of cell types and tissues, which you will study
in more detail in a later exercise. For now, you are mainly concerned with locating crystals. Are you able to locate any
long needle-like crystals either individually or in clusters? The needle-like crystals are raphide crystals, while the
more faceted diamond-like ones are druse crystals. Most crystals are found in the cell sap and are formed from the
precipitation of calcium oxalate. Plants with large stores of calcium oxalate are poisonous to animals and humans.
D. PLASMODESMATA
Carefully observe the demonstration microscope showing plasmodesmata (plasmodesma, singular) in the endosperm
of the persimmon, Diospyros virginianta. What are plasmodesmata and what is their function?
Compare what you see in lab with the figure given in the photographic
atlas (i.e., Figure 1.22).
IV. MITOSIS AND CYTOKINESIS
A. OVERVIEW OF THE CELL CYCLE
Now that you have examined the plant cell and some of its organelles you are ready to study one of the more
important processes which plant cells undergo, that of cell division. Growth in higher plants generally involves an
increase in number of cells resulting from the formation of new (daughter) cells by the division of parent cells. Such
division usually takes place only in special meristematic tissues (tissue in which cells retain their capacity for repeated
division). Meristematic tissues is found in the root and stem tips, which allows them to increase in length, and in the
vascular cambium and cork cambium which allows roots and stems to increase in diameter during secondary growth.
The type of cell division we are considering here is generally referred to as mitosis, although in the strict sense mitosis
refers to division of the nucleus (karyokinesis) while cytokinesis refers to division of the cytoplasm. However, as you
will see, in most plants the process of cell division is a continuous and dynamic process in which nuclear division and
cytoplasmic division blend together smoothly and seem o occur as part of the same overall process.
After cell division, some of the daughter cells mature and as a result of differentiation develop into special
tissues and organs in the growing plant. Other daughter cells remain as meristematic cells and after a period of time
these in turn undergo cell division.
Cytokinesis results in an approximately equal division of the cytoplasm and its components. Nuclear
division, on the other hand, is an extremely precise process in which each daughter cell receives the same genetic
information as that contained in the parent cell.
For sake of convenience, mitosis has been divided into distinct stages, although the actual process is a
continuous even in which on stage merges smoothly into the next.
STAGES OF MITOSIS
i.
INTERPHASE. Strictly speaking, interphase is not a part of mitosis, but events that occur during
interphase make the process of mitosis possible. The nucleus, as well as the entire cell, is extremely active
during the period prior to active division. Many materials such as proteins, carbohydrates, and lipids are
synthesized or accumulated during this period. Perhaps the most important synthesis that occurs is that of
DNA (deoxyribonucleic acid). The DNA with the nucleus must be precisely replicated at this time so that an
ext replication of genetic material may be transmitted to each daughter nucleus. The nuclear cont
(chromatin) when viewed with a light microscope during interphase appears as a dispersed network of faintly
staining material. In Allium, one or more nucleoli may be observed within the nucleus during interphase.
The nucleus is relatively large in those cells, which are in interphase immediately prior to division. The
nucleus at this stage is sharply defined, indicating that the nuclear envelope is intact. Also, the tin cell wall
and the presence of small vacuoles can be seen.
ii.
PROPHASE. The initiation of prophase is indicated by the appearance of long, thin, thread-like
structures (chromosomes) in the nucleus. A gradual shortening and thickening of these
chromosomes occurs as the condensation of chromatin continues and by late prophase they appear
as rod or bar-shaped bodies. Each species has a characteristic number of chromosomes in its cells.
In onion, the characteristic diploid (2n) number is 16, while lily has 24 and humans have 46. In
slides prepared by section, the chromosomes in many cells will be cut away or distorted and because
of this it is usually difficult to determine the number in a given cell.
At the end of prophase, the nuclear envelope and nucleoli begin to disappear, a spindle begins to
form, and chromosomes begin to move toward a median position in the cell. Some authors prefer to
call this prometaphase while others use the term early metaphase for this stage.
iii.
METAPHASE. At metaphase, the chromosomes become arrange so that their centromeres are in a
line parallel to the equatorial plane in a median position in the spindle. In order for normal division
to occur the centromere of each chromosome must be associated with the fibers (microtubules) of
the spindle. These chromosomal fibers along with eh continuous fibers make up the spindle
mechanism. In three-dimension, the spindle would be similar to two cons placed base to base. In
onion it is relatively easy to see the doubled nature of chromosomes at metaphase. Each
chromosome consists of two identical units (chromatids), which are tightly appressed and lie
parallel to each other.
iv.
ANAPHASE. Anaphase begins when the centromere of each chromosome divides allowing the two
chromatids to move toward opposite poles of the spindle. In onion 16 chromatids (now called
daughter chromosomes) move toward one pole while 16 others move toward the opposite pole.
Anaphase ends when the daughter chromosomes reach the poles of the spindle.
v.
TELOPHASE. Telophase begins with the daughter chromosomes aggregating or grouping at the
poles and becoming less distinct. In terms of appearance, the events of telophase are similar to those
of prophase in reverse. Chromosomes become diffuse, a nuclear membrane begins to form, nucleoli
reappear, and the spindle begins to disintegrate. As the daughter nuclei are formed, they assume an
interphase appearance.
vi.
In most plant cells the process of cytokinesis begins as the daughter nuclei are reforming. A cell
plate appears within the spindle along the equatorial plane of the cell and extends laterally until it
reaches the surrounding cell walls, thereby separating the cytoplasm and forming tow daughter
cells. As the cell plate forms, new cell wall material is deposited on each side to complete the newly
formed cells.
B. MITOSIS IN THE ONION ROOT TIP
Obtain a prepared slide of a longitudinal section of onion (Allium) root tip and examine it under low power of
your microscope. You will study root anatomy and function in detail at a later time. For now, a simple orientation to
the toot tip will suffice. The root cap forms a layer of loosely packed, thin-walled cells covering the tip of the root.
The region of cell division (i.e., meristematic region) is located just back of the root cap and beyond this lies
the region of elongation. Cells farthest from the root cap are in the region of maturation may be in various stages of
differentiation. Notice that some of the epidermal cells found on the outer surface of the root in this region of the root
have hair-like extensions (root hairs).
It should be obvious to you by now that these regions of the root are not sharply defined but rather there is a
gradation of one into the other. Now return to the meristematic region and switch to higher power.
Compare what you see in lab with the figures given in the photographic
atlas (i.e., Figure 1.27, 1.28, 1.29). Refer to these figures to help you
locate and study cells, which are in various stages of mitosis.
After you have located a certain stage and studied it carefully under high power, notify your instructor so that
she can verify your identification. Be prepared to give reasons for you answer or otherwise justify your identification.
Place the major characteristics for each of the following stages in the space provided.
Interphase
Prophase
Metaphase
Anaphase
Telophase
Daughter Cells
C. THE DURATION OF MITOTIC STAGES
You have probably suspected by now that the various stages of mitosis are not equal in duration. You should
also realize that the total process of mitosis might proceed at a slower or faster rate depending on the species, the
tissue, and the time of day, temperature, and a number of other factors. Now that you are familiar with appearance of
differentiation mitotic stages, carry out the following calculation, which should give a rough approximation of the
relative length of time required for completion of each stage. For this activity, examine the lowest part of the
meristematic region of your onion root tip slide. Proceeding upward, determine the stage of all mitotic cells until you
have recorded a total of 100 cells. Ignore cells that have nuclei that are in interphase. Record your results in the table
below. Remember, not overlook any cell in any stage of division (including early prophase and late telophase). After
you have completed your calculations, you may wish to compare your results with those of your neighbor.
NOTE: In the event that your slide does not contain a total of 100 mitotic cells, you may finish your count on another
onion root tip slide.
Number of Cells
% Of Total
Prophase
Metaphase
Anaphase
Telophase
Totals
Which stages of mitosis are most prevalent? If you think of the root tip mounted on the prepared slide as a
snap shot of the cellular events that were occurring in this region of the plant when the specimen was collected and
preserved, what do these data suggest as to the length of time spent by cells as they go through these stages of mitosis?