Oxford Nanopore sequencing, hybrid error correction, and de novo assembly of a eukaryotic genome.
Oxford Nanopore sequencing, hybrid error correction, and de novo assembly of a eukaryotic genome.
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DOI：
10.1101/gr.191395.115
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发表时间：
2015-11
期刊：
影响因子：
7
通讯作者：
McCombie WR
中科院分区：
文献类型：
--
作者：
Goodwin S;Gurtowski J;Ethe-Sayers S;Deshpande P;Schatz MC;McCombie WR
监测DNA分子通过膜孔的进程已经被假定为几十年来用于DNA测序的方法。最近，一种基于纳米孔的测序仪器Oxford Nanopore MinION已经上市，我们使用它对酿酒酵母基因组进行测序。为了利用这些数据，我们开发了一种新的开源混合纠错算法Nanocorr，专门用于Oxford Nanopore读取，因为现有的软件包无法以如此高的错误率（错误率在0.5%和40%之间）组装长读取长度（5-50 kbp）。通过这种新方法，我们能够使用互补的MiSeq数据对纳米孔读数进行混合错误校正，并产生高度连续和准确的从头组装：重叠群N50长度比仅Illumina组装大十倍以上（678 kb对59.9 kbp），并且与参考相比具有>99.88%的一致性。此外，具有长纳米孔读段的组装呈现了基因组特征的更完整的表示，并且正确组装了基因盒、rRNA、转座因子和其他基因组特征，这些特征在仅Illumina组装中几乎完全不存在。
Monitoring the progress of DNA molecules through a membrane pore has been postulated as a method for sequencing DNA for several decades. Recently, a nanopore-based sequencing instrument, the Oxford Nanopore MinION, has become available, and we used this for sequencing the Saccharomyces cerevisiae genome. To make use of these data, we developed a novel open-source hybrid error correction algorithm Nanocorr specifically for Oxford Nanopore reads, because existing packages were incapable of assembling the long read lengths (5–50 kbp) at such high error rates (between ∼5% and 40% error). With this new method, we were able to perform a hybrid error correction of the nanopore reads using complementary MiSeq data and produce a de novo assembly that is highly contiguous and accurate: The contig N50 length is more than ten times greater than an Illumina-only assembly (678 kb versus 59.9 kbp) and has >99.88% consensus identity when compared to the reference. Furthermore, the assembly with the long nanopore reads presents a much more complete representation of the features of the genome and correctly assembles gene cassettes, rRNAs, transposable elements, and other genomic features that were almost entirely absent in the Illumina-only assembly.