Core Practicals Flashcards
Investigate the effect of caffiene on the heart rate of Daphnia:
Why do we use daphnia?
Method?
Hypothesis?
Control variables?- solution, daphnia and environments
Why?
-daphnia have a translucent body (easy to observe heart rate)
- Daphnia have a simple nervous system (not complex enough to suffer physical and mental stress)
Method?
- dilute the caffiene solution with distilled water to produce several concentrations
- place cotton wool - to restrict the movement of the daphnia- on a cavity slide.
- allow the daphnia to acclimitise for a minute
- use a stopwatch and a clicker to measure the heart rate
- repeat the experiment and calculte a mean
- T test (when comparing caffiene with no caffiene) but Spearman’s rank when comparing a range of data.
Hypothesis?
Caffiene increases the heart rate of daphnia as it increases the release of stimulatory neurotransmitters
Control Variables?
- use distilled water as a control
- age, gender, species of daphnia
Investagate the vitamin C concentration of food and drink:
Other name for vitamin C:
Method:
Control variables:
Ascorbic acid (vitamin C) is a good reducing agent therefore it can be easily oxidised
Method:
1. Make up a series (5 known vitamin C concentrations and one control experiment with distilled water)
2. Use a measuring cylinder to measure out 1 cm3 of DCPIP solution in a test tube
3. Ad. one of the vitamin C solutions, drop by drop, to the DCPIP solution using a granulated pipette or burette
4. Shake the tube for a set period of time using a stop watch (control variable)
5. When the solution turns colourless record the volume of vitamin C solution added
6. Repeat steps and calculate an average
8. Results can be plotted showing the average volume of vitamin C needed to decolourise DCPIP against the concentration of Vitamin C- calibration curve (can be used to find the concentration of vitamin C in unknown samples
Control variables:
- concentration and volume of DCPIP
- volume of vitamin C
Investagate cell membrane structure including permeability changes:
- specifically investagating how temperature and pH affect the cell membrane.
Method: for temperature
1. Using a cork borer and a scalpel, cut five equally sized sections of the beetroot (equal surface areas and volumes)
2. Rinse the beetroot sections in distilled water and dap with paper towels
3. Add the beetroot pieces to five different test tubes, each containing the same volume of water (5cm3)
4. put each test tube in a water bath at 5 different temperatures for the same length of time (e.g. 10,20, 30 etc)
5.remove the beetroot pieces from the test tubes.
6. Use pipettes to transfer samples of the coloured liquid to colorimeter cuvettes
7. Use a colorimeter of the blue filter to measure how much light is absorbed as it passes through each of the five samples of the coloured liquid- a greater absorbance means that the solution is darker
-for pH, add different concentrations of acid/ alkali of different pHs
Investagating the rate of enzyme controlled reactions:
can be investagated by…
how to calculate initial rate of reaction:
Method:
- measuring the rate of the formation of a product
- measuring the rate of the disappearance of a substrate
- initial rate of reaction measured by drawing a tangeant on a product/ substrate over time graph when t=0 and calculate the gradient of the line
Method:
1. Ad. a set olume of hydrogen peroxide solution to a boiling tube
2. Add a set volume of buffer solution to the same boiling tube
3. Invert a full measuring cylinder into a trough of water
4. Place the end of a delivery tube into the open end of the measuring cylinder and attach the other end to a bung
5. Add a set volume of one concentration of catalase to the boiling tube and quickly place the bung into the boiling tube
6. Record the colime of oxygen collected in the measuring cylinder by the water displaced every ten sconds for 60 seconds
7. Repeat the experiment twice more and calculate the average volume of oxygen produced ay each ten second interval
8. Repeat the whole experiment for different concentrations of catalase
9. Plot the average volume of gas produced against time for each concentration
10. compare the initial rate of reaction of each of the concentrations.
as the volume of catalase increases, the volume of oxygen produced would increase. Oxygen woud plateau eventually due to substrate concentration decreasing. - can be done with a range of substrate concentration using amylase and iodine and starch to create a blue-black appearance as starch is produced.
investagating and observing the stages of mitosis:
-growth in plants occurs in the meristems
-root tip meristem can be used to study mitosis
method:
1. Garlic or onion root tips are most commonly used (the bulbs are encouraged to grow roots but suspending them in water for over a week or two)
2. Prepare a boiling tube of 1M hydrochloric acid and place in a water bath at 60 degrees celcius for ten minutes
3. Remove the tips of the roots (about 1cm) and place in warmed HCl for 5 minute (this softens the middle lamella in the roots)
4. Rinse the tips in cold water using a piupette and blot with a dry paper towel
5. Cut approximetely 2mm off the tip and place onto a microscope slide
6. Add a drop os a suitable stain (toluidine blue)
7. Macerate the root tip using a mounted needle (thinner so easier to see on the microscope)
8. View the slide under the microscope
9. Cells undergoing mitosis can be photographed and redrawn, including labels of the different stages of mitosis.
Identify tissue types within stems:
Method:
1. Cut a very thin cross- section of the stem using a scalpel
2. carefully transfer each section into a dish containing a suitable stain and leave for 1 minutes (e.g. toluidine blue)
3. Rinse off each section in water and mount onto a microscope slide, before adding a cover slip (take care to lower the cover slip slowly over the sample from one side to the other to avoid trapping air bubbles, which can be mistaken for plant structures)
4. View under a microscope and adjust the focus to form a clear image
5. Make a labelled drawing of the positions of the xylem vessels, phloem sieve tubes and sclerenchyma fibres (no arrows for labels)
Investagate plant mineral deficiencies:
Method:
-use 5 different concentrations of one nutrient (nitrogen ion, magnesium ion or calcium broth) or compare between the three and used a distilled water control experiment.
1. Fill the test tube with a set volume (e.g. 10cm3) of each of the nutrient solutions
2. Cover the top of each test tube with foil and create a small hole in the foil
3. For each test tube, take the plantlet and push the roots of the plantlet through the small hole in the foil, enduring that the roots are submerged in the nutrient solutions.
4. Place the test tubes in the same location and ensure that the light intensity and temperature are relatively constant (at least the same between all the plantlets) - ensure that the abiotic factors are kept constant for all the plants
5. After a set period of time (e.g. a week) observe the plantlets and take the qualitative and quantative measurement of their growth (e.g. measure the length of the stem or the number of leaves present for quantative and thew colour of the plant for qualitative)
Determine the tensile strength of plant fibres:
method:
1. Attach the fibre to a clamp stand
2. Attach a weight on the other end of the plant fibre
3. Carefully continue to add one weight at a time until the fibre breaks
4. Record the mass at which the fibre breaks
5. This is the tensile strength
6. To increase tje accuracy of your results, this process should be repeated with more samples of the same plant fibre- calculate a mean
- ensure that all the fibres are of the same length, from the same plant and that all other variables are kept constant
Investagate the antimicrobial properties of plants
Method:
Aseptic techniques:
-work near a lit bunsen burner, clean all surfaces and equipment with disinfectant, flame the nec of the bottle and the inoculating loop, close windows and doors to limit air current.
1. Prior to the practical, bacteria are grown in a nutrient broth before inoculation and incubation on an agar plate (at 25 degrees C- to prevent harmful bacteria thriving at body temp)
2. make sure the bacteria are evenly spread out using a sterile plastic spreader- open the lip of agar as little as possible
3. soak the plant fibre (garlic or mint) in methylated spirit
4. Equal sized discs cut from sterile absorbant paper should be dipped in the plant extract using sterile forceps
5. Leave the discs in the extract for the same amount of time- the discs that is the control will be dipped in only methylated spirit
6. Spread the discs out evenly on the agar plate, before taping the lid on (not all the way round- no oxygen can enter, promoting the growth of anarobic bacteria)
7. Incubate at 25 degrees C for 24 hours
- the larger the zone of inhibition, the greater the antimicrobial properties of the plant
- repeate experiment and calculate a mean
Carry out a study on the ecology of a habitat: using a transect
method:
1. Choose a site where there is an obvious grdient in a abiotic variable and place the transect down. Select a species that chznges in abundance along the gradient.
2. Place a quadrat at each of the marks on the transectm placing the bottom left corner on the mark every time
3. Record the percentage cover for the chosen species. This can be done by recording how many of the quadrat’s 100 squares contain the species ( a square should only be counted if half or more is covered)
4. At each corrdinate, a measure of the independent variable should be taken. For example, investagating light intensity, a photometer can be used to take. areading for the light intensity at each coordinate.
- plot a graph of percentage cover against the chosen independent variable
- carry out the statistical test- spearmans rank, T-test, Chi squared
Investagate photosynthesis using isolated chloroplasts:
Method:
1. Remove stalks from leaf samples. Cut into small sections. Grind samples using a pestle and mortar and place into a chilled isolated solution
2. Place several layers of muslin cloth into a funnel and wet with isolation medium to filter the sample into a beaker
3. Suspend the beaker in an ice water bath to keep sample chilled
4. Transfer to centrifuge tubes and centrifuge at high speed for ten minutes- this will seperate chloroplasts into a pellet
5. Remove supernatant and add pellet tp fresh isolation medium
6. Store isolation solution on ice
7. Set the colorimeter to the red filter. Zereo using cuvette containing chloroplast extract and distilled water
8. Place test tube in a rack 30 cm from light source and add DCPIP. Immediately take a sample and add to a cuvette
9. Measure the absorbance of the sample using the colorimeter
10. Take a sample and measure its absorbance every 2 minutes for 10 minutes.
11. Repeat for different distances from lamp up tp 100 cm. This will vary the light intensity
- plot a graph of absorbance against time for each distance from the light
Investagate the effect of temperature on an enzyme catalysed reaction:
Method:
1. prepare 5 water baths and moniter their temperatures using thermometres.
2. use a pipette to at 2cm cubed of trypsin solution to test 5 tubes and place each one in a water bath.
3. then use the pipette to add 2 cm cubed of ilk to 5 test tubes and again place one in each water bath.
4. start the stopwatch and time for 5 minutes, allowing the contents of each tets tube to reach the temperature of the water bath.
5. pipette 2cm 3 of trypsin and 2cm3 of distilled water into a cuvette and take a colorimetry reading to set the reference absorbance of the colorimeter to zero.
6. now remove the milk test tube and the trypsin tets tube from the 20 degrees C water bath and add the contents of each to a single cuvette, shaking it and then immedietely placing it in the colorimeter.
7. taka an initial reading and then record the absorbance reading at 15 second intervals for 5 minutes or until there is a slight change in absorbance between readings.
8. repeat steps 6-7 fror different temperatures, being sure to rinse the cuvettes between each temperature and setting the absorbance to zero with the water and trypsin cuvette each time.
Investagate the effect of temperature on the development of organisms (to investagate climate change):
Method:
1. Place 2g of sea salt into a beaker containing 100 cm of dechlorinated water. Stir with stirring rod until the salt completely dissolves
2. Put some eggs (brine shrimp) onto a sheet of paper
3. Wet a piece of graph paper in salt water. Place it face down onto the sheet of paper so it picks up some eggs
4. Observe the graph paper under a microscope and count 40 eggs
5. Remove the rest of the eggs/ the paper so that there are only 40 eggs there
6. Place the graph paper upside down into the beaker and leave for 3 minutes until the eggs have detached in the water
7. Incubate the beaker at a set temperature (between 5 degrees and 35 degrees C- this mimics conditions in the wild) for 24 hours
8. Remove the beaker from the incubator and shine a bright light on the beaker. Any hatched larvae will swim towards the light and then can be removed with a pipette.
10. Return the beaker to the incubator and repeatedly remove and count hatched larvae
11. Repeat at a range of temperatures
- plot a graph of temperature against hatched larvae
- as the temperature increases, the number of shrimp hatched increases, up to an optimum of 25 degrees C.
- handle brine shirmp ethically- use salt water and release back into salt water environment after the experiment
Use gel electrophoresis to seperate gene fragments:
Method:
-DNA fragments are cut with endonuclease enzymes
1. Preparation of the gel: the concentration of agrose gel affects the resolution; lower concentrations are used for larger DNA fragments
2. Sample preparation: DNA samples are mixed with a loading buffer that contains dye for visibility and glycerol to increase density (buffer is able to conduct electricity)
3. Loading the samples: the prepared mixture is pepetted into the wells of the gel
4. Running electrophoresis: when an electric current is applies, DNA fragments start migrating through the gel matrix- smaller fragments more faster and therefore further into the gel. DNA is slightly negatively charged therefore, it moves towards the anode.
- resulting bands can be visualised using gene probes (with fluorescnet tags) and UV light
Investagate the effect of different antibiotics on bacteria:
Method:
1. Disinfect the worktop to prevent contamination when preparing the bacteria culture
2. Mark on the agar plate 6 letters (A-F) to show where to place the antibiotic discs
3. Use a bunsen burner to flame the bottle os the neck of the bacterial flask causing the air to rise and carry away undesires airborne microorganisms and then use a sterile pipette to transfer 2cm3 of the liquid broth on the agar plate
4. Use a sterile plastic spreader to evenly distribute the bacteria across the agar jelly.
5. Take the first filter paper disc and use the forceps to completely submerge it in the fist antibiotic for twn seconds. Slightly lift the lid of the agar plate at an angle allowing you to quickly transfer the disc to the agar jelly.
6. Repeat the steps for 4 other antibiotics as well as a controlled disc soaked in distilled water, sterilising the forcepts in between each antibiotic by holding them in the bunsen burner flame.
7. Tape the lid- not all the way to allow oxygen to enter (so anaerobic bacteria dont grow). Incubate the dish at 25 degrees C for 48 hours
8. Disninfect the work surfaces again and wash your hands
9. After the 48 hours have passed, use a ruler to measure the diameeter of the inhibition zone around each disc.
-the greater the inhibition zone, the greater the effects of the antibiotic
