RP2

Question 1

Describe how to prepare squashes of cells from plant root tips

Answer 1

1. Cut a thin slice of root tip (5mm from end) using scalpel and mount onto a slide
2. Soak root tip in hydrochloric acid then rinse
3. Stain for DNA eg. with toluidine blue
4. Lower coverslip using a mounted needle at 45
   o without trapping air bubbles
5. Squash by firmly pressing down on glass slip but do not push sideways

Question 2

Why are root tips used?

Answer 2

● Where dividing cells are found / mitosis occurs

Question 3

Why is a stain used?

Answer 3

● To distinguish chromosomes
● Chromosomes not visible without stain

Question 4

Why squash / press down on cover
slip?

Answer 4

● (Spreads out cells) to create a single layer of cells
● So light passes through to make chromosomes visible

Question 5

Why not push cover slip sideways?

Answer 5

● Avoid rolling cells together / breaking chromosomes

Question 6

Why soak roots in acid?

Answer 6

● Separate cells / cell walls
● To allow stain to diffuse into cells
● To allow cells to be more easily squashed
● To stop mitosis

Question 7

Describe how to set-up and use an optical microscope

Answer 7

1. Clip slide onto stage and turn on light
2. Select lowest power objective lens (usually x 4)
3. a. Use coarse focusing dial to move stage close to lens
   b. Turn coarse focusing dial to move stage away from
   lens until image comes into focus
4. Adjust fine focusing dial to get clear image
   5 Swap to higher power objective lens, then refocus

Question 8

What are the rules of scientific drawing?

Answer 8

✓ Look similar to specimen / image
✓ No sketching / shading - only clear, continuous lines
✓ Include a magnification scale (eg. x 400)
✓ Label with straight, uncrossed lines

Question 9

Explain how the stages of mitosis can be identified

Answer 9

Prophase
● Chromosomes visible / distinct → because condensing
● But randomly arranged → because no spindle activity / not attached
to spindle fibre

Metaphase
● Chromosomes lined up on equator → because attaching to spindle

Anaphase
● Chromatids (in two groups) at poles of spindle
● Chromatids V shaped → because being pulled apart at their
centromeres by spindle fibres

Telophase
● Chromosomes in two sets, one at each pole

Question 10

What is a mitotic index?

Answer 10

● Proportion of cells undergoing mitosis (with visible chromosomes)
● Mitotic index = number of cells undergoing mitosis / total number of cells in sample

Question 11

Explain how to determine a reliable MI from observed squashes

Answer 11

● Count cells in mitosis in field of view
● Count only whole cells / only cells on top and right edges → standardise counting
● Divide this by total number of cells in field of view
● Repeat with many / at least 5 fields of view selected randomly → representative sample
● Calculate a reliable mean

Question 12

Suggest how to calculate the time cells are in a certain phase of mitosis

Answer 12

1. Identify proportion of cells in named phase at any one time
   ● Number of cells in that phase / total number of cells observed
2. Multiply by length of cell cycle