Improving oligonucleotide fingerprinting of rRNA genes by implementation of polony microarray technology

Improvements to oligonucleotide fingerprinting of rRNA genes (OFRG) were obtained by implementing polony microarray technology. OFRG is an array-based method for analyzing microbial community composition. Polonies are discrete clusters of DNA, produced by solid-phase PCR in hydrogels, and derived from individual, spatially isolated DNA molecules. The advantages of a polony-based OFRG method include higher throughput and reductions in the PCR-induced errors and compositional skew inherent in all other PCR-based community composition methods, including high‐throughput sequencing of rRNA genes. Given the similarities between polony microarrays and certain aspects of sequencing methods such as the Illumina platform, we suggest that if concepts presented in this study were implemented in high‐throughput sequencing protocols, a reduction of PCR-induced errors and compositional skew may be realized.

► Polony OFRG distinguished 16S bacterial rRNA genes at or near the species level. ► Weighted UniFrac on polony OFRG data differentiates whole microbial communities. ► PCR artifacts are reduced via spatial isolation of genomic DNA prior to amplification.