CHARACTERIZATION OF UNIVERSAL SMALL-SUBUNIT RIBOSOMAL-RNA HYBRIDIZATION PROBES FOR QUANTITATIVE MOLECULAR MICROBIAL ECOLOGY STUDIES

Universal oligonucleotide hybridization probes targeting the small-sub unit rRNA are commonly used to quantify total microbial representation in environmental samples. Universal probes also serve to normalize re sults obtained with probes targeting specific phylogenetic groups of m icroorganisms, In this study, six universal probes were evaluated for stability of probe-target duplexes by using rRNA from nine organisms r epresenting the three domains of Bacteria, Archaea, and Eucarya. Domai n-specific variations in dissociation temperatures were observed for a ll probes. This could lead to a significant bias when these probes are used to quantify microbial populations in environmental samples. We s uggest lowering the posthybridization wash stringency for two of the u niversal probes (S--Univ-1390-a-A-18 and S-*-Univ-1392-a-A-15) examin ed. These two probes were evaluated with traditional and modified hybr idization conditions to characterize defined mixtures of rRNAs extract ed from pure cultures and rRNA samples obtained from anaerobic digeste r samples. Probe S--Univ-1390-a-A-18 provided excellent estimations o f domain-level community composition of these samples and is recommend ed for future use in microbial ecology studies.