Abstract:
BACKGROUND:Massively parallel pyrosequencing systems have increased the efficiency of DNA sequencing, although the published per-base accuracy of a Roche GS20 is only 96%. In genome projects, highly redundant consensus assemblies can compensate for sequencing errors. In contrast, studies of microbial diversity that catalogue differences between PCR amplicons of ribosomal RNA genes (rDNA) or other conserved gene families cannot take advantage of consensus assemblies to detect and minimize incorrect base calls. RESULTS:We performed an empirical study of the per-base error rate for the Roche GS20 system using sequences of the V6 hypervariable region from cloned microbial ribosomal DNA (tag sequencing). We calculated a 99.5% accuracy rate in unassembled sequences, and identified several factors that can be used to remove a small percentage of low-quality reads, improving the accuracy to 99.75% or better. CONCLUSION:By using objective criteria to eliminate low quality data, the quality of individual GS20 sequence reads in molecular ecological applications can surpass the accuracy of traditional capillary methods.
journal_name
Genome Bioljournal_title
Genome biologyauthors
Huse SM,Huber JA,Morrison HG,Sogin ML,Welch DMdoi
10.1186/gb-2007-8-7-r143subject
Has Abstractpub_date
2007-01-01 00:00:00pages
R143issue
7eissn
1474-7596issn
1474-760Xpii
gb-2007-8-7-r143journal_volume
8pub_type
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