Long-read amplicon denoising
Long-read amplicon denoising
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DOI：
10.1093/nar/gkz657
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发表时间：
2019-10-10
影响因子：
14.9
通讯作者：
Murrell, Ben
中科院分区：
文献类型：
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作者：
Kumar, Venkatesh;Vollbrecht, Thomas;Murrell, Ben
长时间阅读的下一代扩增子测序显示了从复杂和多样化的种群中研究完整基因或基因组的前景。目前的长读测序技术具有挑战性的错误特征，阻碍了数据处理和纳入下游分析。在这里，我们考虑了如何在没有测序错误的情况下从PacBio读数中重建真实的序列变体及其相关频率的问题。这个问题被称为‘扩增子去噪’，已经被广泛地研究用于短读测序技术，但目前的解决方案并不总是成功地推广到具有高INDel错误率的长读。我们介绍了两种方法：一种几乎立即运行，对于中等长度的读取和高模板覆盖率非常准确；另一种较慢的方法在读取非常长或覆盖率较低时更健壮。在两个具有地面事实的模拟病毒社区数据集上，每个数据集都在不同的PacBio仪器上测序，以及在一些模拟数据集上，我们将我们的两种方法彼此比较，并与现有算法进行比较。我们在准确度上超过了所有测试的方法，即使对于速度较慢的方法，我们的运行时间也具有竞争性，成功地区分了只有一个核苷酸差异的模板。Julia实现的快速扩增去噪(FAD)和稳健扩增去噪(RAD)以及网络服务器接口都是免费提供的。
Long-read next-generation amplicon sequencing shows promise for studying complete genes or genomes from complex and diverse populations. Current long-read sequencing technologies have challenging error profiles, hindering data processing and incorporation into downstream analyses. Here we consider the problem of how to reconstruct, free of sequencing error, the true sequence variants and their associated frequencies from PacBio reads. Called 'amplicon denoising', this problem has been extensively studied for short-read sequencing technologies, but current solutions do not always successfully generalize to long reads with high indel error rates. We introduce two methods: one that runs nearly instantly and is very accurate for medium length reads and high template coverage, and another, slower method that is more robust when reads are very long or coverage is lower. On two Mock Virus Community datasets with ground truth, each sequenced on a different PacBio instrument, and on a number of simulated datasets, we compare our two approaches to each other and to existing algorithms. We outperform all tested methods in accuracy, with competitive run times even for our slower method, successfully discriminating templates that differ by a just single nucleotide. Julia implementations of Fast Amplicon Denoising (FAD) and Robust Amplicon Denoising (RAD), and a webserver interface, are freely available.