Unit 3 - Practical Flashcards
Practical Examination
What must you ensure when marking using the capture-mark-recapture technique?
Don’t use a toxic substance as you mustn’t cause them harm, only mark on underside to ensure you don’t make them vulnerable to predators. Mark must be permanent to endure for rest of investigation. Collect a large sample.
What is the equation for population estimation? What is it called?
(s1 x s2) / r s1 = total num captured first time s2 = total num captured second time r = total num recaptured Lincoln Index/Peterson Estimate
What is the method for the capture recapture technique?
Capture num of organisms. Record total caught + mark each. Release + allow to reintegrate for long enough, depending on typical movements. Recapture some. Record total recaptured + total of those marked. Use equation to estimate.
What assumptions are made when using capture mark recapture technique?
No significant gains/losses through immigration/emigration or births/deaths (closed conditions), trapping doesn’t affect animal so it becomes more wary and less likely to be trapped, marked animals have remixed.
What is a haemocytometer?
Instrument for counting cell nums (density). (Yeast/blood cells) Modified microscope slide with grid of squares slightly lower giving depth of 0.1mm so liquid has known volume.
What are the volumes of these squares:
Type A, B + C.
A = 0.1mm3 B = 0.004mm3 C = 0.00025mm3
How do you determine the number to multiply average num of cells by to get the number/mm3?
1/volume
How do you count the cells in a haemocytometer?
First, choose which size square to count from. Then, count using the North West Rule (only count touching cells if touching the top or left side of each square)
How do you set up a haemocytometer?
Clean haemocytometer + cover slip with lens tissue + ethanol. Mix sample well. Use pipette (sample at same depth in solution) to add few drops onto grooves of haemocytometer. Cover slip on. Place on stage + focus. Don’t get solution on top of cover slip or haemocytometer grooves. Ensure to replicate for reliability. Hard to tell between dead + alive cells so not good for decline + stationary phase.
What should you do if there are too many cells to count, even in a type C square?
Serial dilution. 1 part cell suspension to 9 parts isotonic buffer + multiply raw count by dilution factor (10)
What should you do if sampling from natural environment (phytoplankton from lake)?
Retrieve samples from same depth at same time of day.
How do you do the haemocytometer + yeast practical?
Use 3 flasks of diff sizes (150,250,500), pour 50ml apple juice into each + 1 drop yeast suspension. Cover each with muslin (keep out dust + allow air), leave flasks in warm place (25-30c, 3-4days). Set up haemocytometer + estimate population.
What is aseptic technique used for?
To prevent contamination when working with microorganisms, including preventing contamination of microbe culture + individuals working with microbes.
What can be used to culture microorganisms?
Solid agar or liquid broth. Care needed when transferring microorganisms to Petri dishes/other containers for investigation.
What is the procedure for transferring microorganisms?
Metal inoculating/disposable plastic loops can be used. When transferring from culture to petri, lid should be held in same hand as culture bottle + not allowed to touch bench with other hand holding inoc loo. After opening culture bottle, neck should be passed through Bunsen flame to sterilise lid region. Repeat before replacing lid. Lid of petri only raised enough to add microorganisms to agar.
What must you do with a metal inoculating loop?
Flame it in the hottest part of a Bunsen flame until it becomes red hot to sterilise it. Must then allow it to air cool as it would kill any microorganisms it came into contact with. After transfer, must resterilise. Don’t create microorganism-rich aerosol when doing this for safety reasons.
What must you do with disposable plastic loops?
Discard into a solution of disinfectant after use.
What are the 2 methods for transferring cultures to petri dishes?
Inoculating loop or spread plate method.
What is the spread plate method?
Used when cells in suspension used in inoculation. L shaped spreader used to spread inoculating bacteria over agar surface (plastic alt to glass which require repeated sterilisation)
What must you do after transferring culture to petri dish?
Label petri dish on outside of base. Incubate upside down in incubator at appropriate temp for it.
What are 2 different types of antibiotic applications?
Antibiotic discs or e-strips (more accurate)
How do you complete an experiment using antibiotic discs?
Place antibiotic disc on agar in petri dish. Inoculate agar with bacteria.
What are e-strips?
Prepared strips which contain concentration gradient of antibiotic running length of strip. More sophisticated way of measuring effect of diff concs of antibiotic on bacteria. Contain markings which show antibiotic concs at certain points on strip so can work out min antibiotic conc to inhibit bacterial growth.
How do you complete an experiment using e-strips?
Place e-strip on agar of petri inoculated with bacteria.
