Journal
PLOS ONE
Volume 5, Issue 7, Pages -Publisher
PUBLIC LIBRARY SCIENCE
DOI: 10.1371/journal.pone.0011840
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Funding
- Gordon and Betty Moore Foundation Marine Microbiology Initiative
- Center for Microbial Oceanography
- Department of Energy Genome-to-Life (DOE-GTL)
- Natural Science and Engineering Research Council of Canada (NSERC)
- Fonds Quebecois de Recherche sur la Nature et Technologies (FQRNT)
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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 high-throughput 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.
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