A 16S rRNA gene sequencing and analysis protocol for the Illumina MiniSeq platform.
A 16S rRNA gene sequencing and analysis protocol for the Illumina MiniSeq platform.
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DOI：
10.1002/mbo3.611
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发表时间：
2018-12
期刊：
影响因子：
3.4
通讯作者：
Orsi WD
中科院分区：
文献类型：
--
作者：
Pichler M;Coskun ÖK;Ortega-Arbulú AS;Conci N;Wörheide G;Vargas S;Orsi WD
Illumina平台上的16S rRNA基因高通量测序通常用于评估环境样品中的微生物多样性。MiniSeq是Illumina最新的台式测序仪，相对于更大的Illumina测序平台（例如，MiSeq）。在这里，我们使用了一种改进的自定义引物测序方法来测试MiniSeq的保真度，以高通量测序环境样品中复杂群落的16S rRNA基因的V4高变区。为此，我们设计了额外的测序引物，使得能够在MiniSeq上应用双索引条形码方法。一个模拟社区测序的环境样品一起在四个不同的测序运行作为质量控制基准。我们能够在所有测序运行中重新捕获模拟社区的现实丰富性，并确定环境样本中α和β多样性的有意义差异。此外，稀疏分析表明，许多环境样品的多样性接近饱和。这些结果表明，与MiSeq相比，MiniSeq可以产生类似数量的高质量V4读段，但对于任何有兴趣进行高通量16S rRNA基因测序的实验室来说，MiniSeq都是一种具有成本效益的选择。
High‐throughput sequencing of the 16S rRNA gene on the Illumina platform is commonly used to assess microbial diversity in environmental samples. The MiniSeq, Illumina's latest benchtop sequencer, enables more cost‐efficient DNA sequencing relative to larger Illumina sequencing platforms (e.g., MiSeq). Here we used a modified custom primer sequencing approach to test the fidelity of the MiniSeq for high‐throughput sequencing of the V4 hypervariable region of 16S rRNA genes from complex communities in environmental samples. To this end, we designed additional sequencing primers that enabled application of a dual‐index barcoding method on the MiniSeq. A mock community was sequenced alongside the environmental samples in four different sequencing runs as a quality control benchmark. We were able to recapture a realistic richness of the mock community in all sequencing runs, and identify meaningful differences in alpha and beta diversity in the environmental samples. Furthermore, rarefaction analysis indicated diversity in many environmental samples was close to saturation. These results show that the MiniSeq can produce similar quantities of high‐quality V4 reads compared to the MiSeq, yet is a cost‐effective option for any laboratory interested in performing high‐throughput 16S rRNA gene sequencing.
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影响因子：
2.6
作者：
Flynn, Jullien M.;Brown, Emily A.;Chain, Frederic J. J.;MacIsaac, Hugh J.;Cristescu, Melania E.
通讯作者：
Cristescu, Melania E.
DOI：
10.1038/ismej.2012.8
发表时间：
2012-08
期刊：
The ISME journal
影响因子：
--
作者：
通讯作者：
--
影响因子：
1.4
作者：
Apprill, Amy;McNally, Sean;Weber, Laura
通讯作者：
Weber, Laura
影响因子：
4.4
作者：
Bartram, Andrea K.;Lynch, Michael D. J.;Neufeld, Josh D.
通讯作者：
Neufeld, Josh D.
影响因子：
4.6
作者：
Orsi, William D.;Coolen, Marco J. L.;Giosan, Liviu
通讯作者：
Giosan, Liviu