6.1 Microbial Techniques
- Created by: phoebeschofield02
- Created on: 29-12-18 14:44
Microbial Techniques
-important to take care when culturing microorganisms because:
- even if the microorganism you are culturing is harmless, a mutation may cause some to become pathogenic
- there is a risk of contaminating the culture with pathogenic microorganisms from the environment
- when a pure strain is grown, any entry of other microorganisms from the environment contaminates it
-all equipment must be sterile before the culture is started
-once the culture is grown, it must not leave the lab
-all cultures must be disposed of by sealing in plastic bags and then sterilising at 121 degrees C for 15 mins under high pressure before disposal
-microorganisms must be grown with nutrients, wither using nutrient broth (nutrients in liquid form) or nutrient agar (which is a jelly made from seaweed, sets at 50 degrees C and doesn't melt until 90 degrees C)
-selective medium= specific nutrients so only a certain type of microorganism can be grown in it
-selective medium is useful for identifying bacteria that have been genetically modified
-innoculation= getting microorganism onto the agar or broth is called innoculation and is done using an innoculation loop
-when using nutrient broth, it is stoppered with cotton wool to stop contamination with the air and regularly shaken to keep it aerated , allowing oxygen for the growing microorganism
-to isolate bacteria --> grow under anaerobic conditions, use selective medium or use an indicator medium that causes bacteria to change colour
Measuring the growth of bacterial cultures
-Cell Counts:
- counted using a haemocytometer which is a specialised thick microscope slide with rectangular chambers hat can hold 0.1mm3 liquid
- the sameple of nutrient broth is dilued by half with an equal volume of tryptan blue which stains dead cells blue
- each corner of the haemocytometer has a square divided into 16 smaller squares
- the no. of cells in each of the 4 sets of 16 squares is counted and a mean calculated
- the haemocytometer is calibrated so that the no. of bacterial cells in one 16 square is equal to no of cells x 10^4 per cm^3
- measurements are taken at regular intervals
-Optical Methods (turbidity)
- light is passed thorugh a sample
- the more light that gets through, the les turbid the solution therefore the less bacteria are growing in it
- the more turbid a solution, the less light that can get through, therefore there is more bacteria in it
- a calibration curve is produced using a control culture
- turbidity is measured and a cell count using a haemocytometer is doen to give a relationship between turbidity and cell growt
-Dilution Plating
- used to find the total viable cell count
- an orginal culture is diluted until a point is reached when the colonies can be counted
- if the no. of colonies is multiplied by the dilution factor then a total viable cell count can be found
- a mean can then be reached by repeating
-Area and mass of fungi
- when fungi are grown, the diameter of the mycelium can be measured
- this can be…
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