Abstract: The invention disclosed relates to a PCR primer pair for amplication of chaperonin-60 (cpn60) targets having high G+C content and to a PCR primer “cocktail” to improve the representation of diverse sequences in chaperonin-60 based PCR product libraries derived from complex templates. In previous cpn60-based and 16S rDNA-based studies of mammalian intestinal microbiota, it has been observed that some classes of organisms such as the Actinobacteria, which are known through culture-based studies to be present in large numbers in these environments, are underrepresented or even absent from PCR product libraries. Using library sequence data and reference cpn60 sequence data from cpnDB, the chaperonin sequence database, we designed a pair of PCR primers which can be used alone for higher G+C content targets and, when used in combination with a previously developed degenerate, universal cpn60 primer pair, improve the representation of complex templates with high G+C content.

Abstract: Methods of characterizing and identifying microorganisms within a community of microorganisms are described. The methods include amplifying variable regions of the cpn60 gene of samples taken from the microbial community. Primers designed from the analysis of phylogenetic comparisons of nucleotide sequences of the variable regions of the cpn60 gene are also disclosed.

Abstract: cpn60 nucleic acid-based methods for determining microbial profiles are provided, as are cpn60 primers and probes for use in methods of the invention, and kits containing such primers and probes.