肠道菌群核酸提取自动化流程的优化

doi:10.13523/j.cb.20171102

中国生物工程杂志

2017, Vol. 37

Issue (11): 6-11 DOI: 10.13523/j.cb.20171102

计算生物学与生物信息学专栏

肠道菌群核酸提取自动化流程的优化

李金蔓^1,2, 裴广倩², 范航², 黄勇², 童贻刚^1,2

1. 安徽医科大学合肥 230032;
2. 中国人民解放军军事科学院微生物与流行病研究所北京 100071

Optimization of Automatic Flow Chart for Nucleic Acid Extraction from Intestinal Microflora

LI Jin-man^1,2, PEI Guang-qian², FAN Hang², HUANG Yong², TONG Yi-gang^1,2

1. Anhui Medical University, Hefei 230032, China;
2. Institute of Microbiology and Epidemiology, the Academy of Military Sciences PLA China, Beijing 100071, China

全文: PDF(1408 KB) HTML

摘要： 目的：优化肠道菌群核酸提取自动化流程。方法：收集粪便样本，分别采用手工方法、核酸提取工作站提取核酸，然后利用液体工作站制备PCR反应液进行PCR扩增，最后采用文库工作站建库测序。结果：400μl样品用QIAamp^® Fast DNA Stool Mini Kit试剂盒裂解液裂解后，用MagMAX^TM Express 96机器提取核酸的方法与用QIAamp^® Fast DNA Stool Mini Kit试剂盒手工提取核酸的方法相比，测序得到的序列数基本一致，分析结果也没有明显区别；而单独采用MagMAX^TMViral RNA Isolation Kit试剂盒提取核酸由于样品投入体积受限（50μl）、核酸浓度低、测序得到的序列数太少，不能满足后续的分析要求。结论：通过结合使用两种不同的试剂盒，可以实现核酸提取、PCR反应液配制及文库制备的全流程自动化操作，从而大大提高工作效率和结果的稳定性。

关键词： 自动化; 肠道菌群; 鸟枪法测序

Abstract: Objective:Optimize the automatic flow of nucleic acid extraction from intestinal flora. Methods:Fecal samples were collected, and nucleic acids were extracted by manual method and nucleic acid extraction workstation. Then PCR reaction liquid was prepared by liquid workstation, and PCR was amplified and sequenced by library workstation. Results:400μl samples using QIAamp^® Fast DNA Stool Mini Kit, compared with nucleic acid extraction method with QIAamp^® Fast DNA Stool Mini Kit manual with MagMAX^TM Express 96 machine nucleic acid extraction method, sequence number sequencing analysis results obtained are basically the same, there were also no significant differences; the single use of MagMAX^TMViral RNA Isolation Kit to extract nucleic acid sequencing results due to sample input limited volume (50μl), low concentration of nucleic acid and littlesequence number, can't meet the requirements of the follow-up analysis.Conclusion:By using two different kits, the whole process automation of nucleic acid extraction, PCR reaction preparation and library preparation can be realized, thus greatly improving the efficiency and stability of the results.

Key words: Intestinal flora Shotgun sequencing Automation

收稿日期: 2017-09-20 出版日期: 2017-11-15

ZTFLH:

Q-3

基金资助: 病原微生物生物安全国家重点实验室课题资助项目（SKLPBS1518）

通讯作者: 童贻刚 E-mail: tong.yigang@gmail.com

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引用本文:

李金蔓, 裴广倩, 范航, 黄勇, 童贻刚. 肠道菌群核酸提取自动化流程的优化[J]. 中国生物工程杂志, 2017, 37(11): 6-11.

LI Jin-man, PEI Guang-qian, FAN Hang, HUANG Yong, TONG Yi-gang. Optimization of Automatic Flow Chart for Nucleic Acid Extraction from Intestinal Microflora. China Biotechnology, 2017, 37(11): 6-11.

链接本文:

https://manu60.magtech.com.cn/biotech/CN/10.13523/j.cb.20171102 或 https://manu60.magtech.com.cn/biotech/CN/Y2017/V37/I11/6

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