Root Tip Squash

Question 1

What is step 1 of the root squash tip practical?

Answer 1

Heat 1 mol dm HCI at 60°C in a water bath

Question 2

What is step 2 of the root squash tip practical?

Answer 2

Cut a small sample of the root tip using a scalpel

Question 2

What is step 3 of the root squash tip practical?

Answer 2

Transfer root tip to HCI and incubate for 5 minutes

Question 3

What is step 4 of the root squash tip practical?

Answer 3

Remove from HCl and wash sample in cold distilled water and remove the very tip using a scalpel

Question 4

What is step 5 of the root squash tip practical?

Answer 4

Place tip on a microscope slide and add a few drops of stain (e.g. toluidine blue O). This makes the chromosomes visible and will therefore show which cells are undergoing mitosis

Question 5

What is step 6 of the root squash tip practical?

Answer 5

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

Question 6

What is step 7 of the root squash tip practical?

Answer 6

Place under a microscope and set the objective lens on the lowest magnification

Question 7

What is step 8 of the root squash tip practical?

Answer 7

Use the coarse adjustment knob to move the lens down to just above the slide

Question 8

What is step 9 of the root squash tip practical?

Answer 8

Use the fine adjustment knob to carefully re-adjust the focus until the image is clear (you can use a higher magnification if needed)

Question 9

What is step 10 of the root squash tip practical?

Answer 9

To calculate mitotic index, cells undergoing mitosis must be counted (cells with chromosomes visible), as well as the total number of cells

Question 10

What part of the root tip does mitosis occur in?

Answer 10

In plants, cell division by mitosis is restricted to specific tissues, called meristems, that are found at the growing root and shoot tips
The cells in the meristems are undifferentiated
Stem cells in plant meristems divide to produce cells that increase the height of the plant, length of the roots and girth of the stem

Question 11

Why is toluene blue stain used?

Answer 11

It binds to chromatin DNA, staining it deep blue and making the chromosomes visible.
Staining the chromosomes to be clearly visible.

Question 12

Why are the root tips suspended in water?

Answer 12

The root tips are suspended in a drop of distilled/pure water on the slide, the water can help to elongate the cells as it is absorbed by osmosis causing the cells to swell. This can separate the chromosomes and make them easier to see.

Question 13

Why is the acid used?

Answer 13

It helps to break down the cell walls and make them more permeable to allow the stain to penetrate the cell and reach the chromosomes.

Question 14

Why does the coverslip need to be pressed down?

Answer 14

The coverslip on the prepared slide is pressed gently to squash the cells and to produce a layer that is one cell thick this ensures light can travel through the slide.

Question 15

Why do you have to press the coverslip down firmly and not slide it?

Answer 15

Pressing down firmly and not sliding the coverslip prevents one layer of cells from rolling on top of another obscuring the view of the chromosomes. It also prevents chromosomes from breaking or being damaged. The root tip squash is performed gently so that the cells are not ruptured and to prevent the microscope slide and coverslip from cracking

Question 16

What will the results of the practical vary on?

Answer 16

The root tips used, different results may be seen if different types of plant were used and depending on the age of the root tip or time of day the roots were cut.
It could also depend on the conditions the plant was grown in (e.g temperature and water or nutrient availability)

Question 17

How could you make the practical more reliable?

Answer 17

In order for results to be reliable and representative of the plant at the time a large number of fields of view should be observed and an average taken.
Repeat counting the cells in different areas of the cell to get a mean to get a more accurate result

Question 18

What factors can affect the mitotic index?

Answer 18

Nutrients available
Amount of water
Type of plant root tip
Age of root
Temperature

Question 19

How do you calculate the mitotic index?

Answer 19

Number of cells in the field of view undergoing cell division -divided by- total number of cells in the field of view -x100

Question 20

Why is hydrochloric acid a hazard?

Answer 20

There is a risk of causing harm/irritation to eyes or in cuts

Question 21

What are the safety precautions for using hydrochloric acid?

Answer 21

Wear eye protection; avoid contact with skin, tie up long hair

Question 22

Why is toluidine blue O stain a hazard?

Answer 22

There is a risk of causing harm/irritation to eyes or in cuts

Question 23

What are the safety precautions for using toluidine blue O stain?

Answer 23

Wear eye protection; avoid contact with skin

Question 24

Why is using a scalpel a hazard?

Answer 24

There is a risk of there being cuts from sharp object

Question 25

What are the safety precautions for using a scalpel?

Answer 25

Cut away from fingers; use forceps to hold sample whilst cutting, keep away from the edge of the desk

Question 26

Why is possible broken glass a hazard?

Answer 26

There is a risk of there being cuts from sharp object

Question 27

What are the safety precautions for having broken glass?

Answer 27

Take care when handling slides and coverslips; keep glassware away from edge of desk

Question 28

What is the equipment list for the root squash tip practical?

Answer 28

* Optical microscope * Microscope slides and cover slips * Water bath * Hydrochloric acid * Toluidine blue O stain * Distilled water * Scalpel * Forceps * 100 ml beaker * Root tip