Influence of commonly used primer systems on automated ribosomal intergenic spacer analysis of bacterial communities in environmental samples

PloS One
Witoon PurahongDirk Krüger

Abstract

Due to the high diversity of bacteria in many ecosystems, their slow generation times, specific but mostly unknown nutrient requirements and syntrophic interactions, isolation based approaches in microbial ecology mostly fail to describe microbial community structure. Thus, cultivation independent techniques, which rely on directly extracted nucleic acids from the environment, are a well-used alternative. For example, bacterial automated ribosomal intergenic spacer analysis (B-ARISA) is one of the widely used methods for fingerprinting bacterial communities after PCR-based amplification of selected regions of the operon coding for rRNA genes using community DNA. However, B-ARISA alone does not provide any taxonomic information and the results may be severely biased in relation to the primer set selection. Furthermore, amplified DNA stemming from mitochondrial or chloroplast templates might strongly bias the obtained fingerprints. In this study, we determined the applicability of three different B-ARISA primer sets to the study of bacterial communities. The results from in silico analysis harnessing publicly available sequence databases showed that all three primer sets tested are specific to bacteria but only two primers sets a...Continue Reading

References

Oct 7, 2004·Applied and Environmental Microbiology·Massimiliano CardinaleDaniele Daffonchio
Jul 20, 2010·BMC Genomics·Gentile Francesco FicetolaFrançois Pompanon
Sep 11, 2012·Environmental Microbiology·Aline FrossardMark O Gessner

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Citations

Jun 18, 2015·The Science of the Total Environment·Joana P FernandesAna P Mucha
Oct 24, 2018·Conservation Biology : the Journal of the Society for Conservation Biology·Witoon PurahongFrançois Buscot
Oct 28, 2017·Ecology and Evolution·Julia TiedeClaudio Gratton

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Methods Mentioned

BETA
Genome Sequencing
PCR
amplicon sequencing

Software Mentioned

ecoPCR
wgs
embl
FastM
ARISA
Genomatix

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