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Exploring Microbial Diversity and Taxonomy Using SSU rRNA Hypervariable Tag Sequencing
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@MISC{Huse_exploringmicrobial,
author = {Susan M. Huse and Les Dethlefsen and Julie A. Huber and David Mark Welch and David A. Relman and Mitchell L},
title = {Exploring Microbial Diversity and Taxonomy Using SSU rRNA Hypervariable Tag Sequencing},
year = {}
}
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Abstract
Massively parallel pyrosequencing of hypervariable regions from small subunit ribosomal RNA (SSU rRNA) genes can sample a microbial community two or three orders of magnitude more deeply per dollar and per hour than capillary sequencing of full-length SSU rRNA. As with full-length rRNA surveys, each sequence read is a tag surrogate for a single microbe. However, rather than assigning taxonomy by creating gene trees de novo that include all experimental sequences and certain reference taxa, we compare the hypervariable region tags to an extensive database of rRNA sequences and assign taxonomy based on the best match in a Global Alignment for Sequence Taxonomy (GAST) process. The resulting taxonomic census provides information on both composition and diversity of the microbial community. To determine the effectiveness of using only hypervariable region tags for assessing microbial community membership, we compared the taxonomy assigned to the V3 and V6 hypervariable regions with the taxonomy assigned to full-length SSU rRNA sequences isolated from both the human gut and a deep-sea hydrothermal vent. The hypervariable region tags and full-length rRNA sequences provided equivalent taxonomy and measures of relative abundance of microbial communities, even for tags up to 15% divergent from their nearest reference match. The greater sampling depth per dollar afforded by massively parallel pyrosequencing reveals many more members of the ‘‘rare biosphere’ ’ than does capillary sequencing of the full-length gene. In addition, tag sequencing eliminates cloning bias and the sequences are short enough to be completely sequenced
Keyphrases
hypervariable region tag microbial community microbial diversity capillary sequencing full-length gene hypervariable region relative abundance full-length ssu rrna rrna sequence deep-sea hydrothermal vent sequence taxonomy global alignment gene tree tag surrogate parallel pyrosequencing extensive database full-length rrna survey taxonomic census small subunit ribosomal rna ssu rrna reference match v6 hypervariable region certain reference taxon sequence read full-length rrna sequence microbial community membership full-length ssu rrna sequence human gut equivalent taxonomy single microbe experimental sequence
