CP 3 - mitosis

Question 1

What is mitotic index?

Answer 1

The mitotic index
of a sample is the ratio of cells undergoing mitosis to the total number of cells in a
sample.

Question 2

What is special about cells in mertistem tissues?

Answer 2

Cells in the meristems are totipotent and retain the ability to differentiate .

Question 3

Method

Answer 3

1. Heat 1 mol dm -3 HCl at 55 °C in a water bath for 15 minutes.
2. Place the garlic clove in the bottle of HCl and leave it for 5 minutes.
3. Take the clove out and rinse the roots thoroughly in tap water
4. Fill a small bottle of vial with about 2mm depth of acetic orcein stain.
5. Place a pinprick hole in the bottle cap (can also use cling film with holes)
6. Cut a small sample of the root tip , 5-10mm in length, using a scalpel .
7. Transfer root tip to acetic orcein stain and heat in the water bath for 5 minutes.
8. Place tip on a microscope slide. Macerate with a needle to spread the cells out.
9. Lower the cover slip down carefully onto the slide. Make sure there are no air bubbles in the slide which may distort the image, and that the coverslip doesn’t slide sideways which could damage the chromosomes. Wrap in a paper towel and gently ‘squash’ the slide.
10. Remove paper towel
11. Place under a microscope and set the objective lens on the lowest
    magnification, then use the coarse adjustment knob to move the lens down to just above the slide.
12. Use the fine adjustment knob to carefully re-adjust the focus until the image is clear (can use a higher magnification if needed).
13. Count the cells in the various stages of mitosis and divide this by the total number of cells to calculate the percentage of the total cell cycle that stage represents.
14. To calculate mitotic index , cells undergoing mitosis must be counted (cells with chromosomes visible ), as well as the total number of cells .

Question 4

Why are the garlic close root tips put in HCl?

Answer 4

Increases the permeability of the cell, so allowing the stain to penetrate more easily
To soften the tissue/hydrolyse cellulose/loosen cells.

Question 5

Why is it a bad idea to put the acetic orcein stain in an open watch glass?

also why do we put holes in the lid?

Answer 5

it will often evaporate before the stain has penetrated the cells.

prevent it from popping off when heated

Question 6

Why do we squash the root?

Answer 6

It spreads the cells so there is just a single layer to be seen under a the microscope. This makes the individual cells easier to see.

Question 7

Safety

Answer 7

• eye protection
• take care with needle and scalpel
• keep glassware away from edges

Question 8

Why is the plant tissue sometimes put in some other kind of acid or alcohol or both mixture before being added to HCl?

Answer 8

kills the cells and stops mitosis

to ‘fix’ tissue

Question 9

Why do we remove the very tip of the root only?

Answer 9

Mature plant cells lose the ability to divide; the root tip is one region where mitosis occurs/is a meristem.

Question 10

Why is acetic orcein used?

Answer 10

Allows chromosomes to be seen; since (unstained) they have no colour.

Question 11

Why do we squash the cells?

Answer 11

Produce a thin layer; so individual cells can be seen.