Sub culturing of aerobic bacteria, fungus and yeast << Back

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Introduction

In most of the labs, microbes are generally grown in solid agar media or liquid media. It is important to revive new cultures of microbes former to using them in the laboratory exercises. These growth of microbes are to be maintained for it’s longer use. This can be done through the subculturing the organisms.

Definition of subculturing

Aseptic technique of transfer of microorganisms from a used media to a freshly prepared medium. This freshly medium is incubated at organism growth temperature for it’s revival.

General procedure for subculturing

  1. On solid media: the transmission of bacteria or fungi from on agar slant or petri plate containing growth medium.
  2. From solid to liquid media: transmission of bacteria or fungi from on agar slant or petri plate containing liquid growth medium.
  3. From liquid to soild media: transmission of bacteria or fungi from on liquid growth medium on petri plate containing growth medium.
  4. From Liquid to liquid medium:  the transfer of bacteria or fungi from a liquid growth culture to other liquid growth culture.

 

General requirements for subculturing of microbes

Ø  Laminar air flow

Ø  Wire loops

Ø  Lighter/Match sticks

Ø  Petri plates

Ø  Test tubes

Ø  Growth media for microbes

Ø  Burners

Ø  Autoclave

Ø  70% ethanol

Ø  Incubator

Ø  Distilled water

Ø  Glass rod for spreading technique

Ø  Inoculation needle for stab culture technique


 

Method for subculturing the microbes

Generally the subculuring of bacteria is done by different methods depending type of organism i.e. either it is aerobic or anaerobic microbes.

Method for sub- culturing aerobic organism

  1. Spread plates
  2. Streak plate

Streak Plate method

It is fast and quick isolation method for microbe subculture. It is one of the important technique in which the loopful of microbial culture will be grown on nutrient media comprising agar in it. These will lead to the spreading of the whole viable cells and individual cells will be grown as lone colony/single cell on the media.

Procedure for the subculturing by streaking

1)      Prepare the fresh growth media for the microbes in case of mesophilic bacteria we can use the nutrient agar/broth, for common fungus and yeast potato dextrose agar can be used.

2)      Sterilise the media using autoclave (wet sterilisation technique)

3)      After sterilisation fill the test tubes/petri plates with the growth media for the microbes.

4)      Let the media to be cooled at room temperature.

5)      Red hot the wire loop for inoculation

6)      Take of loopful of culture and streak it on the media filled in petri plates or in test tubes.

7)      In case of test tubes for preparing slants, tilt the test tubes at 45 degrees for usage.

8)      Incubate the petri plates/slants at growth temperature of the microbes in incubator.

Different methods for streaking

Ø  Sector streaking

Ø  Four flame streaking

Spread plate technique

Principle of the spread plate technique is same that of the streak plate technique but in this techniques isolation of micorbes is not done directly by applying culture on to the growth media but instead it is first diluted externally either in distilled water or buffer and then culture is applied on growth medium for getting isolated colonies.