Mitotic Cell Division lab
BIOL 150 Lab– Mitotic Cell Division
MITOTIC CELL DIVISION
______________________________________________________________________________LEARNING OBJECTIVES
Upon completion of this exercise the student should be able to:
1. Describe the events that characterize the 4 stages of the cell cycle.
2. Describe the behavior of the chromosomes during the 4 phases of mitosis.
3. Recognize the various phases of mitosis in representative plant and animal cells.
4. Recognize metaphase chromosome arrangements from both lateral and polar views.
5. Distinguish between the mechanisms of cytokinesis of plant and animal cells.
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IMPORTANT TERMS
cell cycle – the cycle of events from the formation of a cell to its next cell division
interphase – phase between mitotic divisions consisting of G1, S, and G2
mitosis – division of the nuclear contents; consists of prophase, metaphase, anaphase, and telophase
cytokinesis – division of the cytoplasmic contents
centrioles – small bodies in animal cells that organize the spindle
spindle fibers – network of microtubules that move the chromosomes during mitosis
metaphase plate (or equatorial plane) – center of the cell where chromosomes align during metaphase
sister chromatids – duplicated chromosomes that remain attached at the centromere until anaphase
kinetochore – proteins at the centromere that allow attachment of chromosomes to the spindle
cell plate – new cell wall that separates daughter cells during cytokinesis of plant cells
cleavage furrow – invagination produced by a belt of microfilaments during cytokinesis of animal cells
THE CELL CYCLE
The first three stages of the cell cycle are characterized by specific biochemical activities.
· The newly formed cell actively produces RNA and protein as it grows. This stage is called G1.
· The cell then stops producing RNA and switches to DNA duplication during the second stage, called S.
· Once the DNA has been replicated, the cell enters another round of RNA and protein synthesis and minor growth. This stage is called G2.
During G1, S, and G2 stages, the chromosomal material is too diffused to be resolved with the light microscope; thus, chromosomes cannot be seen as specific entities at this time.
The cells are stained with acidophilic dyes, that color the nuclei. This colored chromosomal material is called chromatin.
In some cells, one or more nucleoli may be visible within the nucleus. These are areas that are rich in a specific type of RNA called rRNA, and are susceptible to the stain.
When cells are in G1, S, and G2, they are said to be in interphase—which literally means between mitotic divisions. Interphase accounts for 90% of the cell cycle.
Finally, during the M stage, the cell will divide through the process of mitosis and cytokinesis. At the end of the M stage, two new daughter cells have formed
I. The Phases of Mitosis
Although the process of mitosis is a continuous, uninterrupted sequence of activities, it has arbitrarily been divided into, traditionally, four phases for convenience. These phases are prophase (early and late), metaphase, anaphase, and telophase . Late prophase is sometimes elevated to a phase of its own, called prometaphase .
A. Prophase
When mitosis begins, several changes occur in the cell. The chromosomal material (called chromatin) in the nucleus starts to condense, and eventually reaches thickness that makes them visible under the light microscope.
At the same time, the nuclear membrane is disassembled and the nucleoli begin to disperse. In animal cells, a pair of small bodies—the centrioles (also called centrosomes)—begin to migrate toward opposite poles of the cell. There are no centrioles in plant cells. The centrioles seem to be involved in the organization of the spindle fibers. The spindle fibers are a network of microtubules that will later assist in moving the chromosomes from the center of the cell to each of the poles.
Prophase is typically the longest of the phases of mitosis, and it is also the most variable in terms of the appearance of the chromosomes.
As prophase proceeds, the chromosomes continue to get shorter and thicker. During late prophase we can first see that the chromosomes have been duplicated. It is important to note that the chromosomes are duplicated during the S stage of the cell cycle (a part of interphase), but it is not until late prophase that this duplicate nature becomes clearly visible under the light microscope. Late prophase is sometimes elevated to a phase of its own, called prometaphase.
The duplicated chromosomes are separated from one another, except at the centromere. When the duplicate chromosomes are in this attached condition, as they have been since they were replicated, each duplicate is called a sister chromatid. The kinetochore is a protein on the centromere that allows for attachment of a spindle fiber from each pole.
B. Metaphase
By the end of prophase the chromosomes have become maximally short and thick and spindle fibers have attached to the kinetochore proteins.
Some of the spindle fibers still run from one pole to the other (non kinetochore fibers), while others will extend from one pole to a kinetochore.
During metaphase, the chromosomes that are attached to the spindle fibers eventually begin to move toward the equator of the cell, midway between the poles.
At the end of metaphase, each centromere, holding two identical sister chromatids, is attached to two spindle fibers, one from each pole and they are lined up along the metaphase plate.
C. Anaphase
Anaphase begins with the separation of sister chromatids of each duplicated chromosome as the spindle fibers retract.
Each chromatid is moved by motor molecules that ratchet along the spindle fibers toward the polar ends. ATP energy is required for this cellular movement. As the sister chromatids move apart, enzymes degrade the tubulin protein that comprise the microtubules of the spindle fibers. This degradation occurs medial to the kinetochores, not at the polar ends as was once thought.
The original chromosome and its exact duplicate are pulled to opposite poles of the cell. This happens for each different chromosome that the cell possesses; thus, each pole will eventually obtain exactly the same set of chromosomes that were in the original nucleus.
D. Telophase
Once the sister chromatids (now called daughter chromosomes) have reached their respective pole, the last phase of mitosis, telophase, begins. The events of telophase are essentially the reverse of those of prophase. The chromosomes begin to distend (uncoil), the spindle fibers begin to break apart, and a nuclear membrane and nucleoli are reconstituted at each pole. At this point, the division of the nucleus has been completed. There are now two nuclei, each having identical sets of chromosomes.
Watch the following video and refer to the diagrams below for a summary of the events that occur during each phase. https://www.youtube.com/watch?v=1cVZBV9tD-A Bozeman video
For a detailed description of chromosome behavior during mitosis (and meiosis) watch the following video from Montgomery College Rockville Professor Kate Monzo:
II. Cytokinesis
Cytokinesis, division of the cytoplasm, is usually well underway by late telophase and happens somewhat differently in plant and animal cells. In animal cells, cytokinesis involves the formation of a cleavage furrow, which pinches the cell in two. In plant cells, vesicles derived from the Golgi apparatus produce a cell plate at the middle of the cell, separating the new daughter cells. In animal cells, a cleavage furrow formed by a belt of microfilaments, divides the new cells.
Plant Cell Cytokinesis Animal Cell Cytokinesis
III. Observations of Cell Division
A. Plant Cell Division
In order to study mitosis, it is necessary to obtain tissues that contain cells that are actively dividing. One such tissue is the root tip, and you will examine the onion root tip in lab. The unspecialized, dividing cells are not undergoing mitosis in synchrony (i.e., they are not all at the same stage of mitosis at the same time).
The prepared slides contain longitudinal sections of the root tip, which have been chemically fixed and stained for nucleic acid. Since cells in this tissue typically divide in one plane (up-and-down relative to the long axis of the root), most of the cells will be seen from the side relative to the poles. This viewpoint is called a lateral view. If cells were cut along the equator, you would be viewing them from one of the poles (called a polar view). (How would the root tip have to be cut in order to provide polar views of the cells?) This staining procedure has “frozen” each cell at whatever point it had reached at the time of preparation. Each cell can be considered to be a single frame of a motion picture. By properly placing the frames in sequence, you can visualize the entire process of mitosis as a continuous event.
PROCEDURE: The image below shows cells in the growth zone of an onion root tip (Allium cepa) under 40 X and 100X total magnification.
40 X 100 X 400 X
Most of the cells you see are not in the M stage of the cell cycle. That is, most cells are in interphase. However, a sufficient number will be in undergoing various phases of mitosis.
Here is a virtual simulation which lets you zoom in and look at the cells in detail.
http://medsci.indiana.edu/a215/virtualscope/virtual/html5_root_tip_40x.html
See how many phases of mitosis you can identify in the simulation, by looking at the pictures of individual phases below.
Slides of specific stages of plant cell mitosis
Interphase Early prophase Late prophase (prometaphase)
Metaphase Early Anaphase Late Anaphase
Telophase and Cytokinesis Late Telophase and Cytokinesis (you can see (observe the formation of a cell plate) the formation of two new cells)
Here is a picture of the onion root tip with cells in various stages of cell division. Can you identify the phases?
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https://www.youtube.com/watch?v=0UPNcmZQ-KQ mitosis
https://www.youtube.com/watch?v=L61Gp_d7evo Mitosis real time
Plant Cell Cycle: Drawings of Onion Root Tip Cells In the spaces provided to the right and on the following page, draw a cell that is not dividing (in “interphase"), as well as cells in early and late prophase, metaphase, anaphase, and telophase. Label all indicated structures in your drawings. If the cell you draw in telophase is not undergoing cytokinesis, make a separate drawing of a cell in cytokinesis.
B. Animal Cell Division
Mitosis in animal cells is most readily seen in early embryonic stages. The blastula stage of an embryo is more or less a spherical ball of cells, and cell division occurs in every imaginable plane of the embryo. A section through this tissue will cut cells in cross, longitudinal and tangential sections; thus, careful attention to detail will be necessary to correctly interpret what you see. Prophase is not as distinctive in the whitefish embryo as it is in the onion root.
PROCEDURE: Below is an image of cells from a whitefish blastula observed at 40 X, 100 X and 400X total magnification.
40 X 100 X
The whitefish blastula is a ball of cells and each cell will be in a different phase of mitosis as seen under high power below. Most of the cells you see are not in the M stage of the cell cycle. That is, most cells are in interphase. However, a sufficient number will be in undergoing various phases of mitosis.
400 X
Slides of specific stages of animal cell mitosis as seen under 1000 X total magnification
Interphase Prophase Metaphase
Anaphase Early Telophase Telophase and Cytokinesis
Here is a picture of cells from a whitefish blastula undergoing various stages of cell division and is observed at 400X total magnification.
Remember, a lateral view refers to looking at a cell from the side, eye level to the metaphase plate. A polar view refers to looking at a cell from the top or bottom, through the poles.
Animal Cell Cycle: Drawings of whitefish blastula cells
In the spaces provided, draw a cell that is not dividing (in “interphase”), as well as cells in prophase; a lateral view of metaphase; a polar view of metaphase; anaphase; and telophase. Label all indicated structures that you include in your drawings. If the cell you draw in telophase is not undergoing cytokinesis, make a separate drawing of a cell in cytokinesis.
A review video covering Mitosis Lab (Exercise 8 in the Lab Manual) by Montgomery College Rockville Professor Kate Monzo can be accessed at the following link:
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