Multiplexed highly-accurate DNA sequencing of closely-related HIV-1 variants using continuous long reads from single molecule, real-time sequencing.
Multiplexed highly-accurate DNA sequencing of closely-related HIV-1 variants using continuous long reads from single molecule, real-time sequencing.
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DOI：
10.1093/nar/gkv630
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发表时间：
2015-11-16
影响因子：
14.9
通讯作者：
Hunter E
中科院分区：
文献类型：
--
作者：
Dilernia DA;Chien JT;Monaco DC;Brown MP;Ende Z;Deymier MJ;Yue L;Paxinos EE;Allen S;Tirado-Ramos A;Hunter E
单分子实时（SMRT®）测序（Pacific Biosciences，门洛帕克，CA，USA）提供了目前可用的最长连续DNA测序读数。然而，原始读取数据中相对高的错误率需要新的分析方法来解卷积源自复杂样品的序列。在这里，我们提出了一个新的计算机算法的工作流程，能够重建病毒变异体基因组中存在的混合物与> QV 50的准确性。这种方法完全依赖于连续长读段（Continuous Long Reads，简写为STR），这是SMRT测序过程中生成的原始读段。我们成功地实现了这个工作流程，同时测序的混合物含有多达40个不同的>9 kb的HIV-1全基因组。这是使用单个SMRT Cell为每种混合物和桌面计算能力实现的。这种新颖的方法为解决目前缺乏解决方案的复杂测序任务提供了可能性。
Single Molecule, Real-Time (SMRT®) Sequencing (Pacific Biosciences, Menlo Park, CA, USA) provides the longest continuous DNA sequencing reads currently available. However, the relatively high error rate in the raw read data requires novel analysis methods to deconvolute sequences derived from complex samples. Here, we present a workflow of novel computer algorithms able to reconstruct viral variant genomes present in mixtures with an accuracy of >QV50. This approach relies exclusively on Continuous Long Reads (CLR), which are the raw reads generated during SMRT Sequencing. We successfully implement this workflow for simultaneous sequencing of mixtures containing up to forty different >9 kb HIV-1 full genomes. This was achieved using a single SMRT Cell for each mixture and desktop computing power. This novel approach opens the possibility of solving complex sequencing tasks that currently lack a solution.