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PRACTICAL SKILLS Flashcards

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1
Q

What 5 things can you are 5 variables you can change on rate of enzyme controlled reaction

A

enzyme conc
substrate conc
temperature
pH
inhibitors

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2
Q

What can you use as a control when testing pH with enzyme/

A

use water instead of trypsin solution (buffer + trypsin)
proves that temp or buffer itself is not causing the digestion of the substate (only the enzyme is)

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3
Q

What does it mean if no change has occurred beyond around 25mins/

A

the enzyme has denatured

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4
Q

What must you do before you find means between trials

A

eliminate the anomalies

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5
Q

what is important when you are plotting data on a table

A

all figures go to 3sf

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6
Q

how do you find rate of reaction

A

1 / time taken

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7
Q

what do you plot on y and x axis when finding effect of variable on rate in enzyme controlled reaction

A

mean reaction rate (yaxis)
variable e.g pH (x-axis)

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8
Q

When do you go dot to dot or find best line of fit line?

A

when not enough data points to predict a patter, go dot to dot

if you can easily predict the patter then draw line of best fit

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9
Q

You must always critique the method and not the experimenter, hence what are some method limitations when testing variable against rate on enzyme controlled reaction?

A
  1. end point is subjective, e.g the point at which the film turned clear is subjective leading to ‘inaccurate’ time taken measurements
  2. water bath isnt thermostatically controlled so temperature is deacuresing throughout duration of practical
  3. not enough intermediate variables being tested to accurately identify the optimum values
  4. range of variable is not wide enough hence unable to find the point at which the enzyme denatures
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9
Q

You must always critique the method and not the experimenter, hence what are some method limitations when testing variable against rate on enzyme controlled reaction?

A
  1. end point is subjective, e.g the point at which the film turned clear is subjective leading to ‘inaccurate’ time taken measurements
  2. water bath isnt thermostatically controlled so temperature is deacuresing throughout duration of practical
  3. not enough intermediate variables being tested to accurately identify the optimum values
  4. range of variable is not wide enough hence unable to find the point at which the enzyme denatures
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10
Q

Why is HCL, acetic orcein stain, mounted needle and root tip of plant used?

A

HCL - softens and loosens the root tissues so when it comes to squashing the root its easier

acetic orecien stain - stains chromosomes in the nucleus so they are visible under the microscope

mounted needle - lowers the cover slip at 45 degree angle to prevent air bubbles under the slide

root top- growing region where mitosis is occurring

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11
Q

describe process of producing mounted sample to see mitosis

A

heat 5mm root top with HCL in a 60 degree water bath for 10 mins

pour out acid and place root tip on watch glass, rinse with distilled water

place root tip on microscope slid and soak any excess water with filter paper

add 2 drops of acetic orcein stain to root tip

use mounted needle and lower coverslip and leave for 10 mins (provides time to stain chromosomes)

place filter paper on top of slide and gently press down (dont press to hard and break glass or slide glass to side)

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12
Q

why must you squash the root tip

A

speeds out the cells to get a single layer 1 cell thick so light can pass thru when viewing through microscope

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13
Q

what might happen if u slide the cover slip?

A

end up sliding a layer of cells ontop of another layer and the light won’t pass thru or damage the chromosomes so it ruins image

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14
Q

What are hazards, risk and precaution in mitotic index rp

A

scalpel , cut urself, cut ontop of white tile to prevent slipping and cut away from body

HCL, irritant, wear goggles and wash hands

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15
Q

how do you calculate mitotic index

A

number of cells in mitosis / total number of cells (x100)

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16
Q

why are

A
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17
Q

why is root tip stained

A

so chromosomes are visible

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18
Q

How can you ensure accuracy counting of cells to find mitotic index

A

examine for large number of fields of view then find average value

this ensures its more representative of entire root tip

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19
Q

Why is temp controlled when experimenting on osmosis

A

kinetic energy will affect the rate of osmosis so it must be kept the same for all experiments

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20
Q

Why must all test subjects of osmosis be the same length/have the same surface area

A

it affects rate of diffusion

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21
Q

why must you blot the test subject dry before weight

A

so that the excess water on the outside of the cells dont effect the initial mass

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22
Q

why do you measure the initial and final mass of test subject

A

to find change in mass to see if there was gain or loss due to osmosis

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23
Q

Why do you find percentage change in mass

A

fair comparisons as it takes into account that not all test subjects will have had the same initial mass

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24
Q

What are the axis of a calibration curve when testing osmosis

A

y-axis = percentage change in mass
x-axis = sucrose concentration

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25
Q

How do you find the water potential of test subject from the calibration curve

A

find the point at which the curve crosses the x-axis

use a conversion table to look up the conversion of mol dm=3 into water potential

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26
Q

how do you make a serial dilution

A
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27
Q

What should you undergo if asked to find either the qualitative or quantitative results on membrane permeability

A

qualitative = observations of colour

quantitative = use colorimeter and compare absorbance

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28
Q

How is a colour standard made when comparing the pigments release from eating cell membrane permeability?

A
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29
Q

What are the axis on the colour standard calibration curve?

A

y-axis = absorbance at 520nm (or any wavelength depending on the colour of the pigment)

x-axis + percentage extract (of pigment extract in solution)

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30
Q

when testing permeability of cell membranes, what must be controlled and why

A

temperature as it affects the permeability of cell membranes
should be around 30 degrees as anything above may begin to denature membrane proteins causing more pigment to be released

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31
Q

Why must a bung be placed in each test tube when testing cell permeability

A

to ensure the alcohol does not evaporate and change the concentration of the solutions that the “beetroot” is in

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32
Q

Why must 2 discs of beet root be put into each test tube?

A

so theres same surface area and amount so theres same potential maximum amount of vettelin that could move out

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33
Q

Why is beetroot blotted dry before placed in test tube

A

remove excess water and remove excess betalin that is caused by the cutting of the beetroot

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34
Q

Why should you shake the tubes with alcohol and beetroot every min

A

ensures it mixes fully with the alcohol and ensures the betalin can be released from the vacuole

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35
Q

How do we find the percentage extract of beetroot from our results when testing beetroot pigment with different concentrations of alcohol

A

make calibration curve of percentages conc of ethanol (x-axs) with absorbance (y-acxis)

extrapolate the chosen conc of alcohol back to the line and find the absornace

use this absorbance on the colour standard calibration curve and extrapolate back to find percentage extract

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36
Q

Why would these conditions cause betalin to be released : higher temp, alcohol, acid

A

high temp = membrane protein denature so more betalin released

alcohol = phospholipid soluble in alcohol so alcohol will start to dissolve the phospholipid bilayer causing damage and pigment to be released

acid = causes membrane proteins to denature and pigment can be released

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37
Q

Whats the use of dehydrogenase enzyme + where is it found

A

naturally occurs in chloroplasts and catalyses reactions involved in NADP accepting electrons (from photoionisation of chlorophyll and photolysis)

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38
Q

What is DCPIP

A

redox indicator that turns from blue to colourless when it gets reduced/gains electrons

(picks up electrons from LDR instead of NADPH)

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39
Q

Why is ammonium hydroxide used in finding effect of a named factor on the rate of dehydrogenase activity in extracts of chloroplasts.

A

its an alkaline which can denature the dehydrogenase enzyme

it can also accept electrons instead of DCPIP and NADP

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40
Q

what is the hypothesis on effect of a ammonium hydroxide on the rate of dehydrogenase activity in extracts of chloroplasts.

A

rate of reaction decrease with addition of ammonium hydroxide

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41
Q

explain the 5 different test tubes you have when testing the effect of a ammonium hydroxide on the rate of dehydrogenase activity in extracts of chloroplasts.

A
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42
Q

How do you form the chloroplast suspension and explain

A

homogenise/blend spinach leaves and isolation medium which is ice cold with salts to make it isotonic

filter through muslin cloth

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43
Q

What do you do with ur 5 test tubes to test effect of a named factor on the rate of dehydrogenase activity in extracts of chloroplasts.

A

place at a set distance from light (as its the light dependant reactant)

wait for the colours of test tubes to turn the same colour as the testable with only with chloroplast solution (proved that DCPIP has decolourised)

time how long that took

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44
Q

limitations in effect of testing ammonium hydroxide on the rate of dehydrogenase activity in extracts of chloroplasts.

A

end point is subjective

unequal distribution of light

foil isnt blocking out light fro all directions

45
Q

limitations in effect of testing ammonium hydroxide on the rate of dehydrogenase activity in extracts of chloroplasts.

A

end point is subjective

unequal distribution of light

foil isnt blocking out light fro all directions

46
Q

How does a respirometer work and what is it used for

A

used to estimate rate of respiration

works due to volume and pressure changes caused by gas ABSORPTION

47
Q

What are the 2 different test tubes in a repirometer and why are they sealed with bungs and what are they connected by

A

control tube and experimental tube

bung make test tubes air tight as we are looking at changes in volume and pressure so have to ensure no gases enter or leave

thin capillary tube known as manometer contains a red liquid and is connected to the 2 tubes

48
Q

Why is there a scale/rule behind the manometer?

A

so you can measure how far the liquid moves over a period of time

49
Q

Why is there a string on the control tube

A

if you wish to do repeats, you can reset your equipment so you can move the res liquid back to the starting point

50
Q

Why is there a string on the control tube

A

if you wish to do repeats, you can reset your equipment so you can move the res liquid back to the starting point

51
Q

What is in the experimental tube?

A

respiring organism e.g maggots separated from the soda lime (which absorbs carbon dioxide) by a metal mesh

52
Q

What is in the control tube?

A

glass beads and soda line

53
Q

Why is there a metal mesh separating the respiring organism from soda lime

A

just incase the soda lime irritates the organism

54
Q

Why is soda lime used

A

to absorb carbon dioxide and ensure any changes in volume and pressure are purely due to oxygen being absorbed

55
Q

Why are gas beads used

A

same mass of glass beads used as respiring organism as they won’t be retiring and can act as a control

56
Q

explain how the red liquid moves

A

in the experimental tube, the respiring organism is using up oxygen and the co2 it produces is absorbed by the soda lime so the overall pressure in the experimental tube decreases and volume of gas in experimental tube decreases

no respiration is occurring in control tube volume of gas and pressure in control tub is the same BUT it has a higher pressure compared to experimental tube

so force exerted on liquid in manometer and pushes it towards the experimental tube

57
Q

How do you find rate of respiration from respirometer

A

rate = volume / ( time x mass)

units= cm3 min-1 g-1

volume of manometer (cross section are of manometer x change in length of red liquid)

mass of respiring organisms

58
Q

What is the use of dehydrogenase enzyme in yeast

A

catalyses reactions involving removing H from coenzymes and carbon compounds
(involved in all stages of aerobic respiration when NADH is made but also is oxidative phosphorylation where NADH is re-oxidised to NAD.

59
Q

What is TTC

A

redox indicator which is colourless when oxidised and red when reduced

it can pick up a hydrogen removed by dehydrogenase in respiration

60
Q

What happens as you increase them temp to the yeast and TTC mixture

A

itll turn red faster indicating a faster rate of reaction as the temp increases

61
Q

describe the process of varying temp on the enzyme reaction of sugar and yeast with TTC and determining rate of reaction

A

place TTC and yeast sugar mixture in separate test tubes and place in thermostatic water bath, leave for 5 mins to equilibrate the temps of both solutions

when both solutions are same temp, combine the solutions and start a timer, wait for the solution to turn a certain standard colour of pink (not red because the yeast mixture is beige).

ensure to mix whilst waiting as the yeast can fall to the bottom of the tube

when the colour of solution has been reached, record the time taken

repeat experiment at varying temperatures

plot a table of the temps, time taken to turn pink in seconds, rate of reaction

plot a spearman’s rank graph with data

62
Q

What are the limitations of the TTC yeast experiment

A

end-point is subjective (different eyes have different views of pink end point in mind)

difficulties in seeing colour change in water bath

63
Q

Why did the TTC turn red

A

dehydrogenase enzyme which is naturally occurring in yeast removed H from NADH and TTC picked up this H.

TTC got reduced and formed red precipitate

64
Q

At what stages of respiration will dehydrogenase be removing hydrogen

A

NADH is made in glycolysis, links and Krebs as it removed H from carbon compounds.

iT ALSO REMOVED h FROM REDUCED COENZYMES IN OXIDATIVE PHOSPHORYLATION

65
Q

How do you make a control chamber when testing kinesis and taxis of organisms

A

leave all areas of the chamber uncovered so it can be assumed that there will be an even spilt of invertabrae across the chamber

66
Q

What are some risk assessments when using invertebrates to test kinesis and taxis

A

may have pathogens on them which cause infection so use equipment to transfer them instead of ur hands and wash hands after practical

67
Q

How can you test whether there was a significant difference in invertaebrea kinesis and taxis?

A

use chi squared statistic

if theres less than 5% probability that theres difference in distribution due to chance, it can be concluded that environmental variables cause a signig=ficant difference in distribution

68
Q

How can you test whether there was a significant difference in invertaebrea kinesis and taxis?

A

use chi squared statistic

if theres less than 5% probability that theres difference in distribution due to chance, it can be concluded that environmental variables cause a signig=ficant difference in distribution

69
Q

After invertabrae has moved along maze/chamber you need to wipe it down with cotton, why?

A

the invertabrae can leave a trail of chemicals where they move which can influence the choices other invertabrae make

70
Q

what is the purpose of the control maze

A

to show any differences in movement within maze are due to independent variables

71
Q

why is it important to repeat with at least 10 invertebrates

A

so that a mean and statistic can be calculated to see if the difference in turning direction is due to chance or significance

72
Q

How would you make the following conditions in a chamber: dry, humid, dark

A

dry - use silica beads (absorbs moisture)
humid - dampened filter paper
dark - cover with black paper and sellotape it

73
Q

Why is a nylon fabric placed between lid and base

A

easier for organisms to crawl over sheet rather than over the sections of the chamber

ensures they dont touch the silica gel beads

74
Q

Why is a nylon fabric placed between lid and base

A

easier for organisms to crawl over sheet rather than over the sections of the chamber

ensures they dont touch the silica gel beads

75
Q

How do you insert the bugs into the chamber

A

through a hole in the middle of the chamber lid so they all start in the middle

76
Q

why should you wait 5 mins after adding dampened filter paper to chamber

A

provides time for the water from the filter paper to evaporate to create a humid environment and reach the nylon mesh

77
Q

whats a more accurate way to count the invertabrea after removing the lids

A

remove lid and take instant photo and count from the picture

this ensures that you are able to count them before they move along to a different chamber once lid is removed

78
Q

why do invertebrate move to dark and humid chambers

A

humidity is advantageous to prevent desiccation (drying out0

darkness advantageous as its harder for predators to locate and may prevent desiccation

79
Q

What is the serial dilution equation

A

C1 V1 = C2 V2

c1= starting conc
v1 = volume you are trying to transfer
c2 = concentration your trying o make
v2 = final volume

79
Q

What is the serial dilution equation

A

C1 V1 = C2 V2

c1= starting conc
v1 = volume you are trying to transfer
c2 = concentration your trying o make
v2 = final volume

80
Q

How do you test a urine same for its glucose concentration?

A

make a series dilution of glucose concentration

add Benedictus reagent

place it in a boiling water bath for 5 mins and colour should change

use a colorimeter to measure the absorbance of each glucose concentration

plot a calibration curve of glucose concentration (x-axis) against absorbance (y-axis)

add Benedictus eacgent to urine samples and boil them in water bath

place the solution into colorimeter and find the rabsorbance

compare the absorbance of urine samples to calibration curve (extrapolate the absorbance back to the line and find the glucose conc in the urine)

81
Q

Why is sampling used and how do you ensure its accurate

A

to accurately represent population

  • random to eliminate bias
  • line transects to examine a change over distance

-large number of samples 30+

82
Q

How do you know to use a capture release method or quadrate

A

if population is mobile then use capture method

if population is slow moving or non-mobile, use quadrate

83
Q

what are the 2 method for using quadrate depending on if theres uniform or non uniform distribution

A

uniform = random sampling

non-uniform = line transect

84
Q

how do you do random sampling

A

place 2 tape measures act right angle to form gridded area

use random number generator to make 2 coordinates

place quadrate and collect data

repeat at least 30 times and find mead

85
Q

what is belt and interrupted belt transect

A

belt = quadrate placed at every position alone tape measure

interrupted = quadrate placed at uniform intervals along tape measure

86
Q

when would you choose interrupted belt transect over normal belt?

A

if there isnt much variation at every position alone the tape measurement

or to make things faster/more efficient

87
Q

when would you choose interrupted belt transect over normal belt?

A

if there isnt much variation at every position alone the tape measurement

or to make things faster/more efficient

88
Q

how do you line transects

A
  1. place tape measure at right angle TO SHORE LINE
  2. PLACE QUADRATE EVERY 5M OR POSITION
  3. collect data
  4. REPEAT at RIGHT ANGLES to shoreline about 30 times
89
Q

what are the different ways to collect data in a quadrate and how

A
  1. local frequency
    - find the percentage of squares in the quadrate occupied by the species
  2. Density
    - number of species in given area
    ( area of quadrate is 0.25m2 so
    (area of entire field/0.25m2 ) x number of species in quadrate = total number of species in field
  3. Percentage cover
    - proportion of ground occupied by spe died (how many squares do all the species in the quadrate take up)
90
Q

pros and cons of local frequency

A

quick way to sample large area
useful if too hard to identify individual organism

poor accuracy as doesnt consider overlapping plants or its size

91
Q

pros and cons of density

A

more accurate if species is easily distinguishable and not too many to count
can be used to estimate species richness

more time consuming

92
Q

pros and cons of percentage cover

A

faster than density
useful if too hard to identify individual organism or if too many to count

subjective so limits accuracy
doesnt consider overlapping or size of plants

93
Q

pros and cons of percentage cover

A

faster than density
useful if too hard to identify individual organism or if too many to count

subjective so limits accuracy
doesnt consider overlapping or size of plants

94
Q

Describe the process of mark-release-recapture

A
  1. initial sample of population captured
  2. marked and released back into wild. capture number is recorded
  3. after period of time to allow animals to randomly disperse throughout habitat
  4. second sample is captured
  5. total number captured in 2nd sample and unumber recaptured with marking is recorded
  6. size of population is estimated on principle that proportion marked in 2nd sample = proportion of marked individuals in population as a whole
95
Q

ow to increase reliability of mark-release-recapture

A

repeat several times
the more repeated the more reliable

96
Q

whats the equation for mark-release-recapture

A

estimated total population = ( no organisms initially caught x no organisms in 2nd sample) / no of marked organisms recaptured

97
Q

what are some considerations about marking animals

A

non toxic
must increase chance of predetaion
musnt reduce chances of reproduction

98
Q

what assumptions are made with mark-release-recapture

A

population size is constant (no birth, no death, no migration )

animals always redistribute evenly (can huddle unreality)

98
Q

what assumptions are made with mark-release-recapture

A

population size is constant (no birth, no death, no migration )

animals always redistribute evenly (can huddle unreality)

99
Q

What does a potometer measure

A

rate of transpiration

100
Q

why must plant be cut underwater for potometer

A

xylem has negative pressure so tis constantly pulling water up from lower down

so if cut in air, it will pull air into xylem (breaks column of water and transpiration dont work efficiently)

so you need to cut in water to maintain the water column in order for transpiration

101
Q

why is entire potometer assembled under water

A

to remove all air bubbles

102
Q

why is position where plant is attached to potometer smothered in petroleum jelly and have rubber seal

A

make equipment air tight

prevent any water leaking out

ensure water can only leave by evaporation out of stomata on plant

103
Q

how is air bubble introduced into the capillary section of potomenter

A

life the potometer out of the beaker of water for 5-20 seconds and replace

104
Q

how is air bubble introduced into the capillary section of potomenter

A

life the potometer out of the beaker of water for 5-20 seconds and replace

105
Q

how do you find rate of transpiration from potometer

A

wait for introduced air bubble to reach zero on capillary tube scale

have ur variable on

start the timer and measure the time taken for the air bubble to travel a distance

find the volume of water that the air bubble travels ( cross sectional area of capillary tube x distance travelled by air bubble) / time taken for air bubble to move

106
Q

how do you remove the air bubble to reset th epotometer

A

open the tap to the reservoir and flush the air bubble out

107
Q

what variable would be controlled between 2 different plant species testing for rate fo transpiration (not including light, humidity etc)

A

surface area of leaves / number and size of leaves

108
Q

what variable would be controlled between 2 different plant species testing for rate fo transpiration (not including light, humidity etc)

A

surface area of leaves / number and size of leaves

Biology (35 decks)

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