Abstract
The recent release of twenty-two new genome sequences has dramatically increased the data available for mammalian comparative genomics, but twenty of these new sequences are currently limited to ∼2× coverage. Here we examine the extent of sequencing error in these 2× assemblies, and its potential impact in downstream analyses. By comparing 2× assemblies with high-quality sequences from the ENCODE regions, we estimate the rate of sequencing error to be 1–4 errors per kilobase. While this error rate is fairly modest, sequencing error can still have surprising effects. For example, an apparent lineage-specific insertion in a coding region is more likely to reflect sequencing error than a true biological event, and the length distribution of coding indels is strongly distorted by error. We find that most errors are contributed by a small fraction of bases with low quality scores, in particular, by the ends of reads in regions of single-read coverage in the assembly. We explore several approaches for automatic sequencing error mitigation (SEM), making use of the localized nature of sequencing error, the fact that it is well predicted by quality scores, and information about errors that comes from comparisons across species. Our automatic methods for error mitigation cannot replace the need for additional sequencing, but they do allow substantial fractions of errors to be masked or eliminated at the cost of modest amounts of over-correction, and they can reduce the impact of error in downstream phylogenomic analyses. Our error-mitigated alignments are available for download.
Highlights
The field of comparative mammalian genomics has been given an enormous boost by the recent release of genome assemblies for 22 previously unsequenced species of eutherian mammals (26 Mammals Consortium, in prep.)
For each of the fifteen species of interest, we aligned the entire 26 assembly against the ENCODE sequences for that species, applied a series of filters to ensure that corresponding regions were aligned with high confidence
Half or more of the bases in the available ENCODE sequences were in alignment with the 26 assemblies, indicating good coverage in the 26 assemblies and a reasonably sensitive alignment procedure, despite the use of conservative filters
Summary
The field of comparative mammalian genomics has been given an enormous boost by the recent release of genome assemblies for 22 previously unsequenced species of eutherian (placental) mammals (26 Mammals Consortium, in prep.) These new assemblies increase the number of sequenced eutherian species by nearly fourfold, and provide an opportunity for many new functional and evolutionary insights in mammalian genomics. Twenty of these twenty-two genome sequences are currently available only as low- coverage (,26) assemblies (Table 1), produced using traditional, capillary sequencing methods. Low-coverage assemblies necessarily have elevated levels of sequencing error—that is, miscalled bases and erroneous insertions and deletions, which might otherwise be corrected through redundant sequencing of the same genomic region This issue of sequencing error in 26genomes is our focus in this article. Several potential limitations of low-coverage assemblies were examined by Margulies et al [2], but their study focused on assembly and alignment error, and its influence on the detection power for conserved elements in mammalian genomes
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