Background: Different high-throughput nucleic acid sequencing platforms are currently available but a trade-off currently exists between the cost and number of reads that can be generated versus the read length that can be achieved. Methodology/Principal Findings: We describe an experimental and computational pipeline yielding millions of reads that can exceed 200 bp with quality scores approaching that of traditional Sanger sequencing. The method combines an automatable gel-less library construction step with paired-end sequencing on a short-read instrument. With appropriately sized library inserts, mate-pair sequences can overlap, and we describe the SHERA software package that joins them to form a longer composite read. Conclusions/Significance: This strategy is broadly applicable to sequencing applications that benefit from low-cost highthroughput sequencing, but require longer read lengths. We demonstrate that our approach enables metagenomic analyses using the Illumina Genome Analyzer, with low error rates, and at a fraction of the cost of pyrosequencing. © 2010 Rodrigue et al.
CITATION STYLE
Rodrigue, S., Materna, A. C., Timberlake, S. C., Blackburn, M. C., Malmstrom, R. R., Alm, E. J., & Chisholm, S. W. (2010). Unlocking short read sequencing for metagenomics. PLoS ONE, 5(7). https://doi.org/10.1371/journal.pone.0011840
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