amplisas: a web server for multilocus genotyping using next-generation amplicon sequencing data
amplisas: a web server for multilocus genotyping using next-generation amplicon sequencing data
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DOI：
10.1111/1755-0998.12453
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发表时间：
2016-03-01
影响因子：
7.7
通讯作者：
Radwan, Jacek
中科院分区：
文献类型：
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作者：
Sebastian, Alvaro;Herdegen, Magdalena;Radwan, Jacek
下一代测序(NGS)技术作为扩增子测序(AS)的有力工具，正在给生物和医学领域带来革命性的变化。使用引物和条形码的组合，可以在一次实验中对数百个、甚至数千个个体的目标基因组区域进行深度覆盖。这对于基因家族的基因分型是非常有价值的，在这些基因家族中，位点特异的引物往往很难设计，例如主要组织相容性复合体(MHC)。然而，由于NGS技术固有的测序错误率很高，以及其他错误来源，如聚合酶扩增或嵌合体形成，AS的应用受到限制。纠正这些错误需要对NGS数据进行广泛的生物信息后处理。扩增序列分配(Amplisas)是一种工具，它以简单高效的方式执行AS结果分析，同时为高级用户提供定制选项。Amplisas被设计为包括(I)读解复用、(Ii)唯一序列聚类和(Iii)错误序列过滤的三步流水线。等位基因序列和频率以EXCEL电子表格格式检索，使它们易于解释。Amplisas的性能已经成功地与以前发表的利用各种NGS技术获得的MHC基因分型数据集进行了基准比较。
Next-generation sequencing (NGS) technologies are revolutionizing the fields of biology and medicine as powerful tools for amplicon sequencing (AS). Using combinations of primers and barcodes, it is possible to sequence targeted genomic regions with deep coverage for hundreds, even thousands, of individuals in a single experiment. This is extremely valuable for the genotyping of gene families in which locus-specific primers are often difficult to design, such as the major histocompatibility complex (MHC). The utility of AS is, however, limited by the high intrinsic sequencing error rates of NGS technologies and other sources of error such as polymerase amplification or chimera formation. Correcting these errors requires extensive bioinformatic post-processing of NGS data. Amplicon Sequence Assignment (amplisas) is a tool that performs analysis of AS results in a simple and efficient way, while offering customization options for advanced users. amplisas is designed as a three-step pipeline consisting of (i) read demultiplexing, (ii) unique sequence clustering and (iii) erroneous sequence filtering. Allele sequences and frequencies are retrieved in excel spreadsheet format, making them easy to interpret. amplisas performance has been successfully benchmarked against previously published genotyped MHC data sets obtained with various NGS technologies.