required practicals

Question 1

outline required practical 2 (root tip squash) (3 points)

Answer 1

• preparation of stained squashes of cells from plant root tips
• set up & use of an optical microscope to identify the stages of mitosis in these stained squashes
• calculation of a mitotic index

Question 2

what equipment is needed for RP2 (root tip squashes)? (10 items)

Answer 2

• optical microscope
• microscope slides & cover slips
• water bath
• hydrochloric acid
• toludine blue stain
• distilled water
• scalpel
• forceps
• 100ml beaker
• garlic root tip

Question 3

outline the 10 step method of the root tip squash practical

Answer 3

• STEP 1 = heat 1 mol dm -3 HCl as 60˚C in a water bath
• STEP 2 = cut a small sample of the garlic root tip using a scalpel
• STEP 3 = transfer the root tip to the HCl & incubate for 5 minutes
• STEP 4 = remove the root tip from the HCL & wash the sample in cold distilled water. Remove the very tip using a scalpel
• STEP 5 = place the tip on a microscope slide & add a few drops of the toludine blue stain. This makes the chromosomes visible & will therefore show which cells are undergoing mitosis
• STEP 6 = lower the coverslip down carefully onto the slide. Make sure there are no air bubbles in the slide that may distort the image, & that the coverslip doesn’t slide sideways (which could damage the chromosomes)
• STEP 7 = place under a microscope & set the objective lens to the lowest magnification
• STEP 8 = use the coarse adjustment knob to move the lens down to just above the slide
• STEP 9 = use the fine adjustment knob to carefully re-adjust the focus until the image is clear. Use a higher magnification if needed
• STEP 10 = to calculate the mitotic index, the number of cells undergoing mitosis (cells with visible chromosomes) must be counted, as well as the total number of cells. Use the equation (number of cells with visible chromosomes/total number of cells) to calculate the mitotic index

Question 4

outline a risk assessment for the root tip squash practical

Answer 4

• hydrochloric acid may cause harm/irritation to eyes or cuts so wear eye protection, avoid contact with skin & tie up long hair. In an emergency, wash off skin immediately & flood eyes/cuts with cold water
• toludine blue stain may cause harm/irritation to eyes or cuts so wear eye protection & avoid contact with skin. In an emergency, wash of skin immediately & flood eyes/cuts with water
• scalpels way cause cuts so cut away from fingers, use forceps to hold sample whilst cutting & keep away from the edge of desks. In an emergency, elevate cuts, apply pressure & seek medical assistance
• broken glass may cause cuts so take care when handling slides/coverslips & keep glassware away from the edge of desks. In an emergency, elevate cuts, apply pressure, do not remove glass from wound & seek medical assistance

Question 5

outline required practical 4 (beetroot membranes)

Answer 5

• investigation into the effect of a named variable on the permeability of cell-surface membranes

Question 6

what equipment is needed for the beetroot cell membrane practical? (16 items)

Answer 6

• beetroot
• scalpel
• forceps
• cutting board
• ruler
• tongs
• distilled water
• boiling tubes
• boiling tube rack
• colorimeter
• cuvettes
• filter paper
• timer
• water bath
• thermometer
• ethanol

Question 7

describe an 8 step method for the beetroot membranes practical for both temperature & solvents (RP4)

Answer 7

• STEP 1 = cut beetroot into 6-10 identical cubes using a scalpel
• STEP 2 = wipe/rinse to clean off any pigment released as a result
• STEP 3 = (investigating temperature) - place each of the cubes of beetroot in an equal volume if distilled water (5-15ml)
• STEP 4 = (investigating temperature) - place each test tube in a water bath at a range on temperatures (30˚-80˚C)
• (in investigating solvents: create a dilution series of ethanol using distilled water. Ethanol concentrations should range from 0-100% ethanol)
• STEP 5 = leave the samples for 20 minutes so that the pigment leaks out of the beetroot
• STEP 6 = set the colorimeter to a blue filter & zero using a cuvette filled with distilled water
• STEP 7 = filter each sample into a cuvette using filter paper
• STEP 8 = measure the absorbance for each solution. A higher absorbance indicates a higher pigment concentration, & hence a more permeable membrane

Question 8

outline a risk assessment for the beetroot membranes practical

Answer 8

• scalpels may cause cuts so cut away from fingers, use forceps to hold sample while cutting & keep scalpels away from the edge of desks. In an emergency elevate cuts, apply pressure & seek medical assistance
• broken glass may cause cuts so take care when handling glassware & keep away from the edge of desks. In an emergency elevate cuts, apply pressure & seek medical assistance
• ethanol is an irritant & is flammable so wear eye protection & keep away from naked flames. In an emergency, wash eyes & skin with cold water
• hot liquids may cause scalding so handle with care, sue tongs to remove boiling tubes from water bath, wear eyes protection & keep away from the edge of desks. In an emergency run burn under cold water & seek medical assistance

Question 9

what should be on the axis of the graph for the beetroot membranes practical?

Answer 9

• plot a graph of absorbance against ethanol concentration/temperature

Question 10

what conclusions can be made about the affect of temperature on the permeability of beetroot membranes? (RP4)

Answer 10

• as the temperature increases, the permeability of the cell-surface membrane also increases.
• this is because the proteins in the membrane denature as the heat damages the bonds in their tertiary structure
• this creates gaps in the membrane so it is easier for molecules to pass through it
• at low temperatures phospholipids have little energy & are packed closely together to make the membrane rigid
• this causes a decrease in permeability & restricts molecules from crossing the membrane
• at very low temperatures ice crystals can form which can pierce the cell membrane & increase permeability

Question 11

what can we conclude about ethanol concentration on the permeability of beetroot membranes? (RP4)

Answer 11

• ethanol causes the cell-surface membrane to rupture, releasing the betalain pigment from the cell
• higher concentrations of ethanol will cause more disruption to the membrane & more gaps will form
• therefore as the concentration of ethanol increases, the permeability of the cell-surface membrane also increases