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Updated: Jan 17, 2026

Microbiota Analysis Using Two-step PCR and Next-generation 16S rRNA Gene Sequencing

Microbiota Analysis Using Two-step PCR and Next-generation 16S rRNA Gene Sequencing

Published on: October 15, 2019

人类DNA劫持微生物群调查:与结肠相关的16s rRNA amplicon测序中的原因和后果

Leandro Di Gloria¹, Lorenzo Casbarra¹, Marta Bastiani¹

¹Department of Experimental and Clinical Biomedical Sciences, University of Florence, Florence, Italy.

Gut microbiome (Cambridge, England)

|September 25, 2025

概括

在16S测序中的宿主DNA污染可能导致微生物群分析不准确. 使用改性核酸的新策略可以在不改变标准协议的情况下减轻这些错误.

科学领域:

微生物学微生物学
基因组学就是基因组学.
生物信息学是一种生物信息学.

背景情况:

在低于最佳条件下,聚合酶链反应 (PCR) 的效率下降,导致错误的原始和错误的序列.
宿主非目标序列经常在人类肠道样本的16S安普利康测序中产生,特别是在V3-V4原始剂中.
这种污染引入了数据解释错误和废物测序深度.

研究的目的:

分析人类结肠中的V3-V4安普利康序列,以分析微生物群.
为了识别和阐明由宿主非目标序列引入的偏差.
在16S测序中提出对宿主DNA污染的缓解策略.

主要方法:

分析了1300多个公开可用的人类结肠V3-V4安普利康序列.
结肠微生物群组成的分析.
识别特定的人类染色体起源的目标外序列.

主要成果:

未被解决的宿主DNA污染导致错误的细菌识别,并掩盖了真正的微生物群差异.
来自5号,11号和17号染色体的人类序列被确定为非目标序列的主要来源.
提出了一种使用C3间隔器修改核酸准非目标序列的策略.

结论:

在使用V3-V4原料的人类肠道微生物群研究中,宿主DNA污染是一个重要的问题.
拟议的C3间隔器修改核酸策略提供了一个有前途的上游解决方案.
这种方法可以减轻宿主DNA的污染,而不会改变现有的V3-V4原始协议.

关键词:

主机的主机主机的主机主机人类人类人类人类人类人类人类低生物质的生物质.我们的微生物群.错误的初始化问题远离目标 - 远离目标测序的测序是指测序的测序.

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