As known, plasmid is an extra chromosomal DNA, which is capable to replicate independently within bacterial cell.
Plasmid may contain extra genes those code of some special type of proteins, utilized by bacteria. E.g. plasmid may contain code for antibiotic resistant genes.
Because of its capability to replicate independently it is also called as autonomously replicating extrachromosomal molecule of DNA.
In contrast, microbial isolates that are phenotypically distinct have different plasmid fingerprints. Plasmid fingerprint of many E. coli, Salmonella, Campylobacter, and Pseudomonas strains and species has demonstrated that this method often is more accurate than other phenotyping methods such as biotyping, antibiotic resistance patterns, phage typing and serotyping.
The technique of plasmid fingerprinting involves five steps:
- The bacterial strains are grown in broth or on agar plates.
- The cells are harvested and lyses with a detergent.
- The plasmid DNA is separated from the chromosomal DNA and then cut with specific restriction endonucleases.
- The plasmid DNA is applied to agarose gels and electrophoretically separated.
- the gel is stained with ethicist bromide, which binds to DNA, causing it to fluorescent under UV light. The plasmid DNA bands are then located.
Because the migration rate of plasmid DNA in agarose is inversely proportional to the molecular weight, plasmids of a different size appear as distinct bands in the stained gel.
The molecular weight of each plasmid species can then be determined from a plot of the distance that each species has migrated versus the log of the molecular weights of plasmid markers of known size that have electrophoresed simultaneously in the same gel.