Introductory Microbiology

Replica plating technique

• This is also a selection type of mutant identification procedure which will be useful to identify auxotroph mutants. Auxotrophic mutant is a metabolic defective mutant. The mutant which cannot synthesize a particular nutrient, required the nutrient through externally is referred as auxotrophic mutant. The strains which can synthesize the nutrients by themselves are referred as prototrophs. The auxotrophic mutants are very common for amino acids. Normally the wild strains can able to synthesize all the essential amino acids by themselves.
• By mutation if a strain lost its ability to synthesize particular amino acids (Ex. leucine), the mutant is referred as leu- auxotroph mutant. This mutant can grow only the medium supplemented with leucine,. Whereas the prototroth can grow in the medium without leucine. This type of auxotrophic mutant can be selected by replica plating technique.
• The procedure of replica plating technique is as follows; the master plate (mixture of prototroph and auxotroph) will be duplicated to medium with and without leucine by means of rubber stamp like device (called as replica plate). A clone which grows in medium with leucine and which cannot show growth on medium without leucine is an auxotroph.

Replica plating technique for isolation of leucine auxotrophs (leu-); CM, complete medium;, ML, medium devoid of leucine.