三重 PCR 反应对于 16S rRNA 基因扩增子测序来说是不必要的。
Triplicate PCR reactions for 16S rRNA gene amplicon sequencing are unnecessary.
机构信息
Department of Pediatrics, University of California, San Diego, La Jolla, CA, USA.
Division of Bioscience, Argonne National Laboratory University of Chicago, Chicago, IL, USA.
出版信息
Biotechniques. 2019 Jul;67(1):29-32. doi: 10.2144/btn-2018-0192. Epub 2019 May 24.
Abstract
Conventional wisdom holds that PCR amplification for sequencing should employ pooled replicate reactions to reduce bias due to jackpot effects and chimera formation. However, modern amplicon data analysis employs methods that may be less sensitive to such artifacts. Here we directly compare results from single versus triplicate reactions for 16S amplicon sequencing and find no significant impact of adopting a less labor-intensive single-reaction protocol.
摘要
传统观点认为，为了减少 jackpot 效应和嵌合体形成引起的偏倚，测序的 PCR 扩增应采用合并的重复反应。然而，现代扩增子数据分析采用的方法可能对这些伪影不太敏感。在这里，我们直接比较了 16S 扩增子测序中单反应与三重复反应的结果，发现采用较少劳力的单反应方案没有显著影响。
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