1. Mitotic Cell Division

2. Objectives
Learn preparing and staining procedure to identify the stages of mitosis in onion root tip.
To differentiate between the different stages of mitosis.
Calculate the mitotic index.

3. Types of Cell Division Mitosis Meiosis
Produces two new daughter cells with the same number and kind of chromosomes as the parent cell
Meiosis
Reduction division produces progeny cells with one-half the genetic content and number of chromosomes as parent cell
Produces gametes/spores

4. Mitosis Purpose: Karyokinesis Cytokinesis
Mitosis occurs in order for organisms to grow and develop.
In order to replenish dead or dying cells such as skin cells, and cells in the digestive tract.
Karyokinesis
process of nuclear division (division of genetic material)
Cytokinesis
Process of dividing cytoplasm/cell

5. The Cell Cycle
The life of a cell is divided into three stages known as the cell cycle:
1. Interphase:
cell carries out normal functions and prepares to divide
2. Mitosis:
nucleus divides splits into two
3. Cytokinesis:
cell and contents divide into two daughter cells.

6. Cell Cycle

7. G0

8. Interphase
This phase consist of the G1,S, and G2 phases of the cell cycle.
The chromatin is diffuse.
It may not look like much is going on here, but there is a lot of activity because the cell must prepare for Mitosis:
protein synthesis,
DNA synthesis,
replication of other cellular structures.

9. Mitosis There are 4 main phases:
Prophase,
Metaphase,
Anaphase,
Telophase.
Cytokinesis (division of the cytoplasm) follows and one cell becomes two.

10. Mitosis: Prophase Major processes during this phase:
Chromosomes condense and form visible bodies
Chromosomes become thicker, shorter, and easily visible when stained under the light microscope.
Two “sister chromatids” join near their middle at a structure called the centromere.
The nucleolus and the nuclear membrane disappear. The mitotic apparatus the spindle, begins to organize within the cell

11. Mitosis: Metaphase
Chromosomes become aligned at midpoint or equator between poles of the cell
are at their thickest and shortest structure.
They are easily identified as two longitudinally double sister chromatids.
Chromatids are connected (at their centromeres) to the spindle apparatus, which has formed between the two centrioles located at the poles of the cell. In many plants, the centrioles are absent. The spindle is still present, however, and the plant chromosomes are similarly attached to the spindle microtubular fibers.

12. Mitosis: Anaphase The centromere replicates and splits
The sister chromatids separate and are pulled to opposite sides of the cell

13. Mitosis: Telophase Chromosomes now uncoil
Nuclear envelope reappears and surrounds the chromosomes
Cytokinesis
The cytoplasm and all its contents are divided between the 2 daughter cells (cytoplasmic division)
membrane creates between the 2 new daughter cells
In plants, such as the onion root tip cells, this is seen as the formation of a cell plate

14. Stages of mitosis in onion root tip cells

15. Mitosis in Root Tip
In a growing plant root, the cells at the tip of the root are constantly dividing to allow the root to grow.
Because each cell divides independently of the others, a root tip contains cells at different stages of the cell cycle.
This makes a root tip an excellent tissue to study the stages of cell division

19. Experiment

20. Materials Slides & cover slips Microscope Fresh onion root tips
Fixative ( methanol-acetic acid 3:1 v/v)
Forceps
1 M HCl
Razor blade
Stain
Paper towel, or absorbent paper

21. Method Cut 2-3 mm of onion root
Use forceps to transfer an onion root tip into the cup of HCl.
Leave for 4 minutes
Transfer the root tip to the cup containing fixative and leave it for 4 minutes.
Then place the root tip on a slide.
Cover the root tip with a few drops of stain for 2 minutes
Cover the root tip with one to two drops of 45% acetic acid
Put a cover slip over the root, put a paper towel or other absorbent paper and with your thumb firmly press on the cover slip.

22. Observe your preparation under the low power (X10) of a microscope
Search the slide to find cells in various stages of cell division, once you have located cells in division, change to high power (X40) & try to observe several stages of division.
Record the number of cells in each stage. Count at least three full fields of view. You should have counted over 200 cells.
Record your data in the table
Calculate the percentage of cells in each phase and record in the table

23. Mitotic index
A measure for the proliferation status of a cell population.
It is defined as the ratio between the number of cells in mitosis and the total number of cells.

24. Animation

25. Interphase
Prophase
Metaphase
Anaphase
Telophase