Describe how to prepare squashes of cells from plant root tips
- Cut a thin slice of root tip (5mm from end) using scalpel and mount onto a slide
- Soak root tip in hydrochloric acid then rinse
- Stain for DNA eg. with toluidine blue
- Lower coverslip using a mounted needle at 45° without trapping air bubbles
- Squash by firmly pressing down on glass slip but do not push sideways
- Why are root tips used?
- Why is a stain used?
- Why squash / press down on cover slip?
- Why not push cover slip sideways?
- Why soak roots in acid?
- Where dividing cells are found / mitosis occurs
- • To distinguish chromosomes
• Chromosomes not visible without stain - • (Spreads out cells) to create a single layer of cells
• So light passes through to make chromosomes visible - Avoid rolling cells together / breaking chromosomes
- • Separate cells / cell walls
• To allow stain to diffuse into cells
• To allow cells to be more easily squashed
• To stop mitosis
Describe how to set-up and use an optical microscope
- Clip slide onto stage and turn on light
- Select lowest power objective lens (usually x 4)
- 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 - Adjust fine focusing dial to get clear image
- Swap to higher power objective lens, then refocus
What are the rules of scientific drawing?
✓ Look similar to specimen / image
✓ No sketching / shading- only clear, continuous lines
✓ Include a magnification scale (eg. x 400)
✓ Label with straight, uncrossed lines
Explain how the stages of mitosis can be identified
In interphase (not mitosis), chromosomes aren’t visible but nuclei are. In mitosis, chromosomes are visible.
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
What is a mitotic index?
● Proportion of cells undergoing mitosis (with visible chromosomes)
● Mitotic index = number of cells undergoing mitosis / total number of cells in sample
Explain how to determine a reliable MI from observed squashes
● 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
Suggest how to calculate the time cells are in a certain phase of mitosis
- Identify proportion of cells in named phase at any one time
● Number of cells in that phase / total number of cells observed - Multiply by length of cell cycle
Describe how to calculate dilutions
Use the formula: C1 x V1 = C2 x V2
• C1= concentration of stock solution
• V1 = volume of stock solution used to make new concentration
• C2 = concentration of solution you are making
• V2 = volume of new solution you are making
V2 = V1 + volume of distilled water to dilute with
Describe how you would use a 0.5 mol dm-3 solution of sucrose (stock solution) to produce 30 cm3 of a 0.15 mol dm-3 sucrose solution
- Volume of stock solution required, V1 = (C2/C1) x V2
(0.15 ÷ 0.5) x 30 = 9 cm3 - Volume of distilled water to top up with = V2 - V1
30 - 9 = 21 cm3 distilled water
Describe a method to produce of a calibration curve with which to identify
the water potential of plant tissue (eg. potato)
Collecting data:
1. Create a series of dilutions using a 1 mol dm-3 sucrose solution (0.0, 0.2, 0.4, 0.6, 0.8, 1.0 mol dm-3) (CONTROL volume of solution)
2. Use scalpel / cork borer to cut potato into identical cylinders (CONTROL size, shape and surface area of plant tissue ; source of plant tissue ie variety or age)
3. Blot dry with a paper towel (to remove excess water before weighing) and measure / record initial mass of each piece
4. Immerse one chip in each solution and leave for a set time (20-30 mins) in a water bath at 30°C (CONTROL length of time in solution ; temp ; regularly stir / shake to ensure all surfaces exposed)
5. Blot dry with a paper towel and measure / record final mass of each piece
Repeat (3 or more times) at each concentration
Processing data:
6. Calculate % change in mass = (final - initial mass)/ initial mass
7. Plot a graph with concentration on x axis and percentage change in mass on y axis (calibration curve)
● Must show positive and negative regions
8. Identify concentration where line of best fit intercepts x axis (0% change)
● Water potential of sucrose solution = water potential of potato cells
9. Use a table in a textbook to find the water potential of that solution
Why calculate % change in mass?
● Enables comparison / shows proportional change
● As plant tissue samples had different initial masses
Why blot dry before weighing?
● Solution on surface will add to mass (only want to measure water taken up or lost)
● Amount of solution on cube varies (so ensure same amount of solution on outside)
Explain the changes in plant tissue mass when placed in different concentrations of solute
Increase in mass
● Water moved into cells by osmosis
● As water potential of solution higher than inside cells
Decrease in mass
● Water moved out of cells by osmosis
● As water potential of solution lower than inside cells
No change
● No net gain/loss of water by osmosis
● As water potential of solution = water potential of cells
Describe a method to investigate the effect of a named variable (eg.
temperature) on the permeability of cell-surface membranes
- Cut equal sized / identical cubes of plant tissue (eg. beetroot) of same age / type using a scalpel
- Rinse to remove pigment released during cutting or blot on paper towel
- Add same number of cubes to 5 different test tubes containing same volume of water (eg. 5cm3)
- Place each test tube in a water bath at a different temperature (eg. 10, 20, 30, 40, 50 degrees)
- Leave for same amount of time (eg. 20 mins)
- Remove beetroot and measure intensity of colour of surrounding solution:
● Semi-quantitatively
- Use a known conc. of extract & distilled water to prepare a dilution series (colour standards)
- Compare results with colour standards to estimate conc.
● Quantitatively
- Measure absorbance (of light) of known concentrations using a colorimeter
- Draw a calibration curve → plot a graph of absorbance (y) against conc. of extract (x) and draw a line / curve of best fit
- Absorbance value for sample read off calibration curve to find associated extract conc
What are the issues with comparing to a colour standard?
● Matching to colour standards is subjective
● Colour obtained may not match any of colour standards
Why wash the beetroot before placing it in water?
● Wash off any pigment on surface
● To show that release is only due to [named variable]
Why regularly shake each test tube containing cubes of plant tissue?
● To ensure all surfaces of cubes remain in contact with liquid
● To maintain a concentration gradient for diffusion
Why control the volume of water?
● Too much water would dilute the pigment so solution will appear lighter / more light passes through in colorimeter than expected
● So results are comparable
How could you ensure beetroot cylinders were kept at the same temperature throughout the experiment?
● Take readings in intervals throughout experiment of temperature in tube using a digital thermometer /temperature sensor
● Use corrective measure if temperature has fluctuated
What does a high absorbance suggest about the cell-membrane?
● More permeable / damaged
● As more pigment leaks out making surrounding solution more concentrated (darker)
Explain how temperature affects permeability of cell-surface membranes
● As temperature increases, permeability increases
- Phospholipids gain kinetic energy and fluidity increases
- Transport proteins denature at high temperatures as H bonds break, changing tertiary structure
● At very low temperatures, permeability increases
- Ice crystals can form which pierce the cell membrane and increase permeability
Explain how pH affects permeability of cell-surface membranes
● High or low pH increases permeability
- Transport proteins denature as H / ionic bonds break, changing tertiary structure
Explain how lipid-soluble solvents eg. alcohol affect permeability of cell-surface membranes
● As concentration increases, permeability increases
● Ethanol (a lipid-soluble solvent) may dissolve phospholipid bilayer (gaps form)
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3.1 Biological molecules90
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3.2 Cells84
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3.3 Organisms exchange substances with their environment71
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3.4 Genetic information, variation and relationships between organisms64
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3.5 Energy transfers in and between organisms (A-level only)53
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3.6 Organisms respond to changes in their internal and external environments (A-level only)85
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3.7 Genetics, populations, evolution and ecosystems (A-level only)71
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3.8 The control of gene expression (A-level only)82
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3.1 RP7
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3.2 RP24
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3.3 RP3
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3.4 RP11
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3.5 RP28
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3.6 RP (incomplete)8
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3.7 RP8
