Lab Exercise 5 - Pure Culture Techniques

pure culture

a culture containing a single microbial species

very rare (or impossible) in environmental and patient samples

important in clinical and research settings

goal of pure culture techniques

to dilute bacterial samples in order to separate individual cells that can grow into isolated colonies when plated

3 different types of media

general purpose

selective

differential

When are general purpose media often used?

when trying to isolate microbes from a mixed culture

Are solid or liquid media preferred when trying to obtain pure cultures?

solid media are often preferred

agar

a solid medium often used when obtaining pure cultures

a solidifying agent derived from seaweed

cannot be metabolized by microbes

3 different methods of culture preparation

pour plate method

spread plate isolation

isolation streak plate

pour plate method

culture is diluted in molten agar, mixed well, and poured into an empty sterile Petri dish, allowed to solidify

solidification occurs at 40*C

colonies can be observed on top of, embedded in, or underneath the agar

can be used quantitatively or non-quantitatively depending on the dilution method used

spread plate isolation method

serial dilution is performed in tubes

a small sample from each dilution is plated, spread across an agar plate surface

when dilute enough, individual cells will replicate into colonies on the surface of the agar (only on the surface)

isolation streak plate method

sample is plated across agar surface in 3 zones (I, II, and III, following a template), then in a single zigzag across zone IIIB

a primary streak can yield a pure culture or mixed isolated cultures

a secondary streak from one of the isolated cultures can yield a pure culture

isolation should be achieved in zone IIIB