The cell cycle in plant cells
The onion root tip is one of the most widely used materials for the study of the cell cycle because it is readily available, preparation of the dividing cells is easy, and the chromosomes are large and few in number - hence, easier to study than the cells of many other organisms. Since root tips are regions of active cell division, chances are good that in a specimen of such tissues, one can find every stage of mitosis. You should recognize that the tissue has been particularly selected for this property; most tissues have few mitotic figures, and one needs to search extensively to find the number of stages that are easy to find in a single root tip.
It is a temptation to think of the four stages of mitosis as discrete phases with no overlap; however, mitosis is a continuous process in which one phase fades into the next. You may find cells which are intermediate between two stages rather than clearly showing all of the characteristics of a single stage.
Obtain a slide of onion root tips (Allium), and note a series of dark streaks. Each streak is a longitudinal section through an onion root tip (Perry & Morton, Figure 10a). Observe the longitudinal section part of the model of the onion root tip. At the pointed end of the root (tip), there is a root cap of loosely organized cells, which protect the delicate growth region of the root. Inside of the root cap, you will find a clear, organized set of rapidly dividing, cubic cells called the root meristem. These are the cells you are concentrating upon in this exercise. Note that as you move away from the root cap, the cells become more elongated. The majority of cell division takes place in the meristem; further elongation of the root occurs as the cells further back continue growth and elongation, but no further cell division. Finally, the central portion of the root is differentiated into the vascular tissue that carries water and minerals from the root into the remainder of the plant.
Place your slide on the stage of your microscope and locate one of the sections under low power. Because the section is very thin, not all will be equally good for study. After this preliminary screening, with the meristem in the center of your field, change to high power to observe mitosis. Keep in mind the sequence in which the stages occur (as described below), but do not try to find them in sequence. Because cells remain in interphase and prophase longer than the other stages, chances are that most of the cells you observe will be in interphase, many will be in prophase, and only a few will be in metaphase, anaphase and telophase. Examine as many regions of meristem as necessary to see all the stages. Diagram cells that you see in each stage; use the figures referred to in your "Atlas" to assist you in recognizing them.
Interphase: Although these cells are resting with respect to nuclear division, they are actively undergoing respiration and even synthesis of DNA, RNA and protein, in preparation for mitosis. The nucleus is obvious, but no chromosomes are clearly visible in these cells. (Perry & Morton, Figure 10b)
Prophase: During
prophase, the DNA, which was originally in long, thin strands, becomes
condensed as a result of coiling and supercoiling. The nuclear membrane begins to
break down, and the chromosomes are distributed throughout the
nucleoplasm. During prophase
in the onion root tip, the chromosomes often appear as a coiled mass
(Perry & Morton, Figure 10c/d). Even at this early stage, the DNA of
each chromosome has doubled, though this is difficult to see on a
slide. Under very high
magnifications, it is possible to see that each chromosome is composed of
two separate strands, the sister chromatids. The two sister chromatids are
identical in structure, chemistry, and the genetic information they carry,
because one was replicated (copied) from the original DNA of the other
during the last S
phase.
The sister chromatids are joined together at a
region of attachment called the centromere. Within this region, each chromatid
contains a disc-shaped kinetochore. Microtubules (called kinetochore
microtubules) insert into the kinetochores and run from them outward to
the two poles of the cell.
Other polar microtubules become organized into the spindle
fibers.
Metaphase:
During early metaphase,
some of the polar microtubules break down and new attachments are made
between the kinetochore microtubules and tubules from the opposite
pole. This results in what
appear to be rather aimless chromosome movements, aptly described as
"dancing chromosomes."
As metaphase
progresses, a random breaking and reattachment of kinetochore microtubules
to the polar microtubules of the same or opposite poles occurs until
(randomly) the kinetochore of one sister chromatid is attached to
microtubules from one pole and the kinetochore of the other sister
chromatid is attached to tubules from the opposite pole. Then the polar microtubules pull
in such a way that the kinetochores become positioned in a region halfway
between the poles (Perry & Morton, Figure 11a). This region, which occupies a
plane near the center of the cell (and at right angles to the long axis of
the spindle fibers) is called the metaphase plate (or equatorial
plate.)
The cell is considered to have reached metaphase when the kinetochores of all chromosomes have arrived at this metaphase plate region. At this time, the centromeres divide in preparation for separation of the sister chromatids during the following stage.
Anaphase: The sister chromatids that make up each chromosome are separated from each other and are pulled by the microtubules to opposite poles of the cell. As the centromeres are pulled apart, the arms of these daughter chromatids (as they are now called) are passively dragged along. Thus, anaphase in onion cells can be recognized by the two groups of V-shaped chromosomes on opposite sides of the cell. The sharp, pointed end of the V is oriented toward the pole of the spindle (Perry & Morton, Figure 11b).
Reduce the light by adjusting the diaphragm of the microscope, and try to locate any spindle fibers near the center of the cell. They appear as very fine lines between the two groups of chromosomes. Anaphase ends when the newly separated chromosomes arrive at the opposite poles of the cells.
Telophase: Karyokinesis is completed during telophase, and reorganization of the contents of the two daughter cells (cytokinesis) begins (Perry & Morton, Figure 11c). It is often difficult to distinguish late anaphase from early telophase in the cells of plants. During telophase, however, a cell plate, the first indication that cytokinesis is beginning, starts to form as a fine line across the center of the cell. When complete, the cell plate divides the original cell into two daughter cells. As telophase progresses, the nuclei begin to reorganize: the chromosomes uncoil and become longer and thinner, the nuclear membrane reforms, and the nucleoli reappear.
Mitosis ends with the assembly of two interphase nuclei, each with one complete set of single-stranded chromosomes (Perry & Morton, Figure 11d). The daughter cells (Perry & Morton, Figure 11e) resulting from mitotic division have the same number and kinds of chromosomes (and therefore the same genetic makeup) as the original cell.