required practicals

Question 1

RP 2 - prep of stained squashes of cells from plant root tips

Answer 1

1) add 10ml of 5mol^dm-3 hydrochloric acid to a beaker placed on a bench mat
2) cut small sample of root tip using scalpel
3) transfer root tip immediately to hydrochloric acid in beaker and leave for 15 minutes
4) set up microscope whilst waiting
5) remove root tip and rinse in distilled water in watch glass
6) cut off root tip (1mm) and place on microscope slide
7) place few drops of toluidine blue stain and macerate with mounted needle to separate cells
8) add cover slip and spread root tip using gentle pressure and blot simultaneously using filter paper between finger and slide
9) place on microscope stage
10) set objective lens to lowest magnification and use coarse adjustment knob to readjust focus until image clear, using higher magnification if necessary
11) calculate mitotic index - no.of cells in mitosis/ total number of cells

Question 2

what is purpose of HCl - RP 2

Answer 2

-destroy substances uniting cells to break into tissue and individual cells to halt process of mitosis

Question 3

how what u use an eyepiece graticule to determine the mean diameter of an organelle

Answer 3

-place stage micrometer on stage
-look through eyepiece graticule and align with stage micrometer to calibrate
-calculate how many known values on stage micrometer equal one eyepiece graticule unit
-measure each organelle using eyepiece graticule
-take number of measurements to calculate mean

Question 4

why is the lowest mag needed - RP 2

Answer 4

-larger area of slide can be viewed
-specfic areas can be located easier
-focusing is easier

Question 5

where does mitosis occur in the root - RP 2

Answer 5

meristem

Question 6

how is the mitotic index ensured to be reliable

Answer 6

-select multiple fields of view/many cells to ensure representative sample
-pick fields of view at random (e.g. using grid and random number generator)
-repeat count to ensure figures correct
-count only whole cells
-calculate mean of different mitotic indexes

Question 7

RP 3

Answer 7

1) make a serial dilution of sucrose solutions using distilled water - at ​0.0, 0.2, 0.4, 0.6, 0.8 and 1.0 mol sucrose
2) measure 5cm^3 of each dilution into separate test tubes
3) use cork borer to cut out six potato chips and cut down sections into identically sized chips using the scalpel
4) dry each chip using a paper towel to remove excess water
5) weigh each before start of experiment and record
6) place a potato chip in each test tube and leave for 20 minutes
7) remove each potato chip and blot dry using paper towel before weighing each again
8) calculate percentage change in mass for each potato chip
9) plot graph of percentage change in mass (y-axis) against concentration of sucrose solution (x-axis)
10) x axis intercept is equal to the concentration of the potato
11) use this to find data for the water potential of the potato - compare against another source/ graph

Question 8

what are the variables in RP 3

Answer 8

independent - conc of sucrose
dependent - percentage change in mass
control- vol of sucrose soltuion, size of potato chips

Question 9

RP4 - permeability of membranes - often used with beetroot or carrots or smth else

Answer 9

1) cut beetroot into slice of uniform thickness using scalpel
2) cut slice into 10 identical discs using cork borers
3) rinse discs with distilled water to remove any excess pigment
4) create a dilution series of ethanol ranging from 0 - 100% using distilled water
5) place two discs in each test tube
6) leave samples for 5 minutes
7) set colorimeter to a blue filter and zero using a cuvette with distilled water
8) filter each sample into a cuvette using filter paper
9) measure absorbance for each solution
10) higher absorbance = higher pigment concentration = more permeable membrane
11) plot graph of absorbance (y-axis) against ethanol concentration/ temperature (x-axis)

Question 10

variables and controls (group) for RP4

Answer 10

control group - 2 beetroot discs in distilled water to show any pigment released from the action of the alcohol
independent - alcohol conc or temp
dependent - absorbance
control - size of beetroot discs, vol of solution, time in beetroot solution

Question 11

notes for RP 4

Answer 11

-beetroot contains purple pigment betalain, which is contained within vacuole
-can indicate permeability as betalain leaks out of cell membrane when membrane permeability increased
-ethanol is non-polar so acts as solvent for phospholipids, dissolving them

Question 12

in RP 4 why would using a calorimeter improve the results

Answer 12

-quantative method
-colour change is subjective
-standardises the method

Question 13

why wash beetroot discs at start - RP 4

Answer 13

-wash off any betalain pigment
-show that any pigment released is from the effect of alcohol conc/temp

Question 14

why would the results be different if used cooked beetroot

Answer 14

-heating would damage the plasma membranes
-denatures the protein and disprupts phospholipid bilayer
-lots of pigment released during cooking

Question 15

why do u shake the tt every min - RP 4

Answer 15

-increase contact of all surfaces with the alcohol
-prevent discs sticking tg
-maintain diffusion gradient for pigment

Question 16

RP 5 - heart

Answer 16

Method:
1. Wear protective equipment (goggles and plastic gloves) throughout the whole practical.
2. Disinfect the workspace before proceeding with the dissection.
3. Observe the heat and try to identify the aorta and the pulmonary artery and locate the valves.
4. Then run water into these arteries and see how the valves work. To release the water, squeeze the heart gently over the sink.
5. To dissect the heart, make a cut along the side of the heart to open the left and right atria and ventricles. Then look for the atrioventricular valves between the atria and the ventricles.
6. Identify the chambers and out pins or labels in them once you have identified them and take a photo to record this. Then you can compare the thickness of the muscle tissue between the chambers

Question 17

rules for drawing a labelled diagram

Answer 17

• Use a sharp HB pencil
• Make sure drawing is large enough for detail and labels (at least half a page advisable)
• No shading
• Use single and continuous lines (no sketching)
• Label lines should be drawn with a ruler
• Do not cross label lines
• Lines should not have any arrows
• Label all the structures
• Do not use colour
• Include a magnification/scale

Question 18

RP 6 - aseptic techniques

Answer 18

1) use sterile pipette/wire hoop to transfer bacteria from broth to agar plate
2) spread bacteria evenly over plate using plastic spreader
3) use sterile forceps to place multi-disc antibiotic ring on plate by holding its centre
4) tape lid on (do not tape around entire lid), invert and incubate at 25°C for 48 hours
5) sterilise equipment and disinfect surfaces
6) measure diameter of inhibition zone for each antibiotic ensuring lid of agar plate not removed
7) calculate area of inhibition zone A = πd / 4
8) plot bar chart of area of inhibition zone (y-axis) against antibiotic (x-axis)

Question 19

controls (group) and variables for RP 6

Answer 19

control variable - add water to bacteria on agar plate to show that the bacteria do not die regardless of antibiotic

indpendent - antiobiotic
dependent - area of inhibitor zone
control -time disk in antibiotic conc

Question 20

aseptic techniques - RP 6

Answer 20

1) use sterile pipette/wire hoop to transfer bacteria from broth to agar plate
2) spread bacteria evenly over plate using plastic spreader
3) use sterile forceps to place multi-disc antibiotic ring on plate by holding its centre
4) tape lid on (do not tape around entire lid), invert and incubate at 25°C for 48 hours
5) sterilise equipment and disinfect surfaces
6) measure diameter of inhibition zone for each antibiotic ensuring lid of agar plate not removed
7) calculate area of inhibition zone A = πd / 4
8) plot bar chart of area of inhibition zone (y-axis) against antibiotic (x-axis)

Question 21

what factors affect the rate of bacterial growth

Answer 21

-increased conc of glucose - increase resp
-increased conc of oxygen so increase resp
-increased temp so increased emzyme activity
-increased conc of phosphate so increased ATP/DNA/RNA
-increased conc of nucleotides so increased DNA synthesis

Question 22

why is it important to have sterile techniques - RP 6

Answer 22

-prevent release of bacteria into the air
-prevent contamination of appartus with bacteria

Question 23

why do u not tape the entire petri dish - RP 6

Answer 23

-prevents oxygen entering
-promotes growth of more harmful anaerobic bacteria

Question 24

why are there clear zones around some of the antibiotics - RP 6

Answer 24

-antibiotic has diffused into agar
-killed bacteria

Question 25

why might an effective antibiotic produce a small clear zone

Answer 25

-antibiotic may diffuse slowly

Question 26

conclusion - RP 6

Answer 26

-If there is a larger inhibition zone around the antibiotic, it has killed more bacteria. Therefore, the larger the inhibition zone, the better the antibiotic works.
-Some antibiotics will have no/very little inhibition zone. This shows that the bacteria are resistant to this antibiotic and are not killed by it.