General/Last year Flashcards
What is % (w/v)?
- g/100ml
What is M?
- mol/l
What is accuracy?
- whether all values are close to true value
What is consistency (precision)?
- whether all values are close to each other
Why would you dilate bacteria in saline rather than water?
- saline has same Ψ as bacterial cells
- pure water would cause water to enter cells and burst
What is the difference between viable cell count and total cell count?
- viable is actively growing cells
- total inc dead cells
What is a streak plate culture and what does it allow?
- cooled medium poured into petri dish and allowed to set in smooth even layer
- inoculum streaked onto surface of medium to prod well separated colonies after incubation
- allows cells to be spread more evenly , so individual colonies can be isolated
What is a spread plate culture?
- plate prepared like streak
- inoculum is drop of liquid bacterial culture
- transferred w/ pipette and spread using spreader dipped in alcohol and passed through bunsen flame (sterilising it)
What is a pour plate culture, and when is it a useful technique?
- inoculum added to melted and cooled agar medium
- gently mixed and poured into plates
- or inoculum placed into petri dish and liquid medium added
- to ensure even distribution, moved w/ rotating movement
- useful for isolation of bacterial cultures from mixed inoculum, or where heavy even growth needed
What is an overlay culture?
- similar to pour plates
- inoculum mixed w/ smaller amount of melted agar medium
- poured in thin layer to solidify on top of solid medium already in petri dish
What are stages of a gram stain?
- heat fixed smear
- stain w/ methyl violet
- iodine to trap stain in peptidoglycan layer
- ethanol wash
What do the results of a gram stain indicate about whether cells are gram +ve or -ve?
- +ve = purple
- -ve = pink/red
What is the difference between gram +ve and gram -ve cells?
- gram +ve have thick cell wall, so retains methyl violet
- gram -ve have thin cell wall
What does G banding do?
- stains gene-poor, heterochromatic regions
- leaving transcriptionally active regions unstained
- generates specific banding pattern for each chromosome
Would you expect standard curve for spectrophotometer to be linear and how long for?
- in theory always linear
- in practise it wouldn’t, as machine can’t accurately measure v small amount of light being transmitted
- compound becomes more conc, will reach point where no longer be soluble
What does absorbance equal?
- A = ECt
- E is extinction coefficient
- C is conc
- t is path length
How is pH calculated from [H+]?
- -log[H+]
What is the equation for Ka?
- Ka = [H+][A-] / [HA]
What is the Henderson-Hasselbalch equation?
- pH = pKa + log( [A-] / [HA] )
How is pKa calculated from [Ka]?
- -log[Ka]
What is the relationship between pH and pKa when [A-] = [HA]?
- pH = pKa
What are the limitations of Lineweaver-Burk plot?
- difficult to find x-intercept as never any minus values
- bunching of lower values relied on to draw line to bigger values
- larger values unreliable as at low substrate conc
What is an autoclave?
- strong heated container used to sterilise equipment by subjecting them to high pressure steam at 138kPa, heated to 121º for 20 mins
How can optical density be used to measure bacterial growth?
- cells in suspension scatter light
- only proportion of light passes through to be detected by photocell in spectrophotometer
- as cell density increases w/ growth, more light scattered and optical density increases
What features do plasmids used as vectors generally have?
- contain origin of replication functional in E. coli
- multiple restriction sites, which can be used to insert foreign DNA
- encode gene for resistance for antibiotic as means of selection
- relatively small and present at high copy no. per cell so can be purified in large amounts
What % agarose gels do labs typically use and what can this resolve?
- 1%
- can adequately resolve DNA fragments ~1-10kb
What factors can proteins be purified by?
- size
- shape
- charge
- solubility
- stability
- binding specificity
What is the formula for chi-squared?
Σ { (O-E)2 / E }
How is dilution factor calculated?
- total final vol / vol of original solution
- starting conc / final conc
How is total dilution factor of serial dilutions calculated?
- multiply individual dilution factors together
What are doubling dilutions?
- series of 2 fold dilutions
What are the prefixes for the different powers of 10 from 6 to -15?
- 6 = mega (M)
- 3 = kilo (k)
- 2 = hecto (h)
- 1 = Deca (da)
- -1 = deci (d)
- -2 = centi (c)
- -3 = milli (m)
- -6 = micro (μ)
- -9 = nano (n)
- -12 = pico (p)
- -15 = femto (f)
How can volumes in L be converted to volumes in m^3?
- 1L = 1dm^3
- 1ml = 1cm^3
- 1μl = 1mm^3
How do you calculate standard deviation?
- √ of Σ {(x – x)^2} / (n-1)
How is density calc?
- mass / vol
What is the density of water?
- 1g/mol
What is the mass of 1 mol C-12?
- 12g
What does Avogadro’s no. tell you?
- = no. units in a mole
- = atoms of 12C in 12g of 12C
What equation can be done if you know a vol and 2 concs (or a conc and 2 vols)?
- c1v1 = c2v2
How do you get ATP/EGTA/EDTA to dissolve in water?
- pH will drop to 3 –> stirring and heating will have no effect
- titrate w/ base to neutralise and dissolve acid, then correct to req pH after dissolved
When should serial dilutions be carried out, and why?
- when adding v small vols to v large vols
- as error would be too large and mixing not even
How do you calibrate a pH meter?
- remove cap and rinse in distilled water
- switch on (slide to right)
- calibrate in pH7 buffer and wait for it to settle
- take small screwdriver and adjust top left screw (ACW to decrease reading)
- calibrate in pH4 or 9.2 (whichever brackets pH aiming for)
- adjust right hand screw
- rinse w/ distilled water
How do you make a solution of 250ml w/ 2 solid components?
- weigh out and transfer to beakers
- add <250ml (as solids contribute to vol and need to add acid and alkali to get to desired pH) –> about 200ml
- put in magnetic stir bar
- make to desired pH (acid to lower) add dropwise w/ pasteur pipette, w/ stirrer still in
- rinse probe and replace cap
- make up to 250ml w/ distilled water, by pouring all back into cylinder
- mix in beaker
What is the difference between technical and biological repeats?
- technical repeats = repeated measurements of same sample –> look at measurement error w/ equipment etc.
- biological repeats = repeating whole experiment to get biologically distinct samples –> look at random biological variation
