Can composition and structural features of oligonucleotides contribute to their wide-scale applicability as random PCR primers in mapping bacterial genome diversity?

Abstract

Among current genotypic methodologies, random amplification of polymorphic DNA (RAPD or AP-PCR) represents a widely employed assay for the evaluation of bacterial genomic diversity. A common bottleneck of this technique, however, is represented by the screening of useful informative primers to discriminate among isolates of a particular bacterial species. In an attempt to simplify this process, we evaluated here the utility of degenerate oligonucleotides to act as informative AP-PCR primers. For this purpose, a number of features (G+C contents, degeneracy rate, modifications at the 5' end) of related degenerate primers was tested for their effects in the generation of informative arrays from a set of bacterial genomes. Our results indicate that a combination of a wide base composition and a common palindromic structure at the 5' end of the sequences that compose the degenerate primers tested here beneficially resulted for the generation of informative arrays aimed to evaluate the bacterial genome heterogeneity.