The molecular techniques have been widely used for typing and assessing the genetic diversity of microorganisms. The techniques based on polymerase chain reaction (PCR) fingerprinting have been employed to examine genotypic diversity of rhizobial populations and to discriminate among rhizobial strains. These techniques include random amplified polymorphic DNA (RAPD), two-primers RAPD (TP-RAPD), repetitive sequence based PCR (rep-PCR) and amplified fragment length polymorphism (AFLP). The 3 main techniques of rep-PCR are enterobacterial repetitive intergenic consensus (ERIC)-PCR, repetitive extragenic palindromic (REP)-PCR and BOX-PCR. While RAPD uses a single primer to amplify the segments of DNA randomly throughout the genome, rep-PCR uses pairs of primers (for ERIC- and REP-PCR) or a single primer (for BOX-PCR) to amplify the intervals between conserved repeated sequences present in genome. In AFLP, total genomic DNA is digested and then ligated to oligonucleotide adapters. A pair of primer is used to amplify the product from restriction. RAPD, rep-PCR and AFLP are suitable for distinguishing strains at species or below levels but they are less valuable for taxonomic purpose. TP-RAPD has been developed for taxonomy purpose as the patterns of strains in the same species have been found to be identical. The TP-RAPD patterns supported the proposal of novel species of rhizobia.
Keywords: Amplified fragment length polymorphism (AFLP), BOX-polymerase chain reaction (BOX-PCR), Enterobacterial repetitive intergenic consensuspolymerase chain reaction (ERIC-PCR), Genotypic diversity, Polymerase chain reaction (PCR) fingerprinting, Random amplified polymorphic DNA (RAPD), Repetitive extragenic palindromic-polymerase chain reaction (REP-PCR), Repetitive sequence based polymerase chain reaction (rep-PCR), Rhizobia, Twoprimers random amplified polymorphic DNA (TP-RAPD).