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An improved dual-indexing approach for multiplexed 16S rRNA gene sequencing on the Illumina MiSeq platform

机译：一种改进的双索引方法，用于在Illumina MiSeq平台上进行多重16S rRNA基因测序

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Background To take advantage of affordable high-throughput next-generation sequencing technologies to characterize microbial community composition often requires the development of improved methods to overcome technical limitations inherent to the sequencing platforms. Sequencing low sequence diversity libraries such as 16S rRNA amplicons has been problematic on the Illumina MiSeq platform and often generates sequences of suboptimal quality. Results Here we present an improved dual-indexing amplification and sequencing approach to assess the composition of microbial communities from clinical samples using the V3-V4 region of the 16S rRNA gene on the Illumina MiSeq platform. We introduced a 0 to 7 bp “heterogeneity spacer” to the index sequence that allows an equal proportion of samples to be sequenced out of phase. Conclusions Our approach yields high quality sequence data from 16S rRNA gene amplicons using both 250 bp and 300 bp paired-end MiSeq protocols and provides a flexible and cost-effective sequencing option.
机译：背景技术为了利用负担得起的高通量下一代测序技术来表征微生物群落组成，通常需要开发改进的方法来克服测序平台固有的技术局限性。在Illumina MiSeq平台上对低序列多样性文库（例如16S rRNA扩增子）进行测序一直存在问题，并且经常产生次优质量的序列。结果在这里，我们提出一种改进的双索引扩增和测序方法，以使用Illumina MiSeq平台上16S rRNA基因的V3-V4区评估临床样品中的微生物群落组成。我们在索引序列中引入了0到7 bp的“异质间隔子”，以允许等比例的样本异相测序。结论我们的方法使用250 bp和300 bp配对末端MiSeq协议从16S rRNA基因扩增子中获得高质量的序列数据，并提供了灵活而经济高效的测序方法。

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