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比较评估一种新型基于磁珠的粪便样本 DNA 提取方法,用于下游的新一代 16S rRNA 基因测序。

Comparative evaluation of a new magnetic bead-based DNA extraction method from fecal samples for downstream next-generation 16S rRNA gene sequencing.

机构信息

Department of Psychiatry and Behavioral Sciences, Duke University Medical Center, Durham, North Carolina, United States of America.

Mouse Behavioral and Neuroendocrine Analysis Core Facility, Duke University Medical Center, Durham, North Carolina, United States of America.

出版信息

PLoS One. 2018 Aug 23;13(8):e0202858. doi: 10.1371/journal.pone.0202858. eCollection 2018.

DOI:10.1371/journal.pone.0202858
PMID:30138447
原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC6107275/
Abstract

We are colonized by a vast population of genetically diverse microbes, the majority of which are unculturable bacteria that reside within the gastrointestinal tract. As affordable, advanced next-generation sequencing technologies become more widely available, important discoveries about the composition and function of these microbes become increasingly possible. In addition to rapid advancement in sequencing technologies, automated systems have been developed for nucleic acid extraction; however, these methods have yet to be widely used for the isolation of bacterial DNA from fecal samples. Here, we adapted Promega's Maxwell® RSC PureFood GMO and Authentication kit for use with fecal samples and compared it to the commonly used Qiagen QIAamp® PowerFecal® kit. Results showed that the two approaches yielded similar measures of DNA purity and successful next-generation sequencing amplification and produced comparable composition of microbial communities. However, DNA extraction with the Maxwell® RSC kit produced higher concentrations with a lower fecal sample input weight and took a fraction of the time compared to the QIAamp® PowerFecal® protocol. The results of this study demonstrate that the Promega Maxwell® RSC system can be used for medium-throughput DNA extraction in a time-efficient manner without compromising the quality of the downstream sequencing.

摘要

我们被大量遗传上多样化的微生物所定植,其中大多数是无法培养的细菌,它们存在于胃肠道内。随着价格合理、先进的下一代测序技术越来越普及,对这些微生物的组成和功能的重要发现变得越来越有可能。除了测序技术的快速进步外,还开发了用于核酸提取的自动化系统;然而,这些方法尚未广泛用于从粪便样本中分离细菌 DNA。在这里,我们将 Promega 的 Maxwell® RSC PureFood GMO 和 Authentication 试剂盒进行了改编,使其可用于粪便样本,并将其与常用的 Qiagen QIAamp® PowerFecal®试剂盒进行了比较。结果表明,两种方法在 DNA 纯度、下一代测序扩增的成功以及微生物群落的组成方面都取得了相似的效果。然而,与 QIAamp® PowerFecal®方案相比,Maxwell® RSC 试剂盒提取的 DNA 浓度更高,所需粪便样本输入量更少,且所需时间也更少。这项研究的结果表明,Promega 的 Maxwell® RSC 系统可用于高效、高通量的 DNA 提取,而不会影响下游测序的质量。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/ae55/6107275/8e0354acbb56/pone.0202858.g007.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/ae55/6107275/fca17e9c2a72/pone.0202858.g001.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/ae55/6107275/8c27e4543d5e/pone.0202858.g002.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/ae55/6107275/1957cef42775/pone.0202858.g003.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/ae55/6107275/71215e6d192a/pone.0202858.g004.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/ae55/6107275/3b393c0b119c/pone.0202858.g005.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/ae55/6107275/7a9b2ed384e0/pone.0202858.g006.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/ae55/6107275/8e0354acbb56/pone.0202858.g007.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/ae55/6107275/fca17e9c2a72/pone.0202858.g001.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/ae55/6107275/8c27e4543d5e/pone.0202858.g002.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/ae55/6107275/1957cef42775/pone.0202858.g003.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/ae55/6107275/71215e6d192a/pone.0202858.g004.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/ae55/6107275/3b393c0b119c/pone.0202858.g005.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/ae55/6107275/7a9b2ed384e0/pone.0202858.g006.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/ae55/6107275/8e0354acbb56/pone.0202858.g007.jpg

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