生物多样性 ›› 2019, Vol. 27 ›› Issue (5): 534-542.DOI: 10.17520/biods.2018201

• 综述 • 上一篇    下一篇

基于三代测序技术的微生物组学研究进展

许亚昆1,2,马越1,2,胡小茜1,王军1,*()   

  1. 1 中国科学院微生物研究所, 北京 100101
    2 中国科学院大学, 北京 100049
  • 收稿日期:2018-07-30 接受日期:2018-12-25 出版日期:2019-05-20 发布日期:2019-05-20
  • 通讯作者: 王军
  • 基金资助:
    科技部重点研发子课题(2018YFC2000504);国家自然科学基金(31771481)

Analysis of prospective microbiology research using third-generation sequencing technology

Xu Yakun1,2,Ma Yue1,2,Hu Xiaoxi1,Wang Jun1,*()   

  1. 1 Institute of Microbiology, Chinese Academy of Sciences, Beijing 100101
    2 University of Chinese Academy of Sciences, Beijing 100049
  • Received:2018-07-30 Accepted:2018-12-25 Online:2019-05-20 Published:2019-05-20
  • Contact: Wang Jun

摘要:

微生物在人类生活中无处不在, 过去人们对微生物的认识仅停留在单菌培养和定性研究上, 而测序技术的发展极大地促进了微生物组学的研究。越来越多的证据表明: 人体共生微生物、特别是肠道微生物与人类健康息息相关。 二代测序技术凭借其高通量、高准确率和低成本的特点, 成为微生物组学研究中的主流测序技术。但是随着研究的深入, 二代测序技术的短读长(< 450 bp)增加了后续数据分析和基因组拼接难度, 也限制了该技术在未来研究中的应用。在此背景下, 第三代测序技术应运而生。第三代测序技术又称单分子测序, 能够直接对单个DNA分子进行实时测序, 而不需要经过PCR扩增。第三代测序技术的平均读长在2-10 kb左右, 最高可以达到2.2 Mb, 实现了长序列的高通量测序。凭借其超长的测序读长、无GC偏好性等优势, 三代测序技术为微生物基因组全长测序, 组装完整可靠的基因组提供了新的方法。本文在描述三代测序的技术特点和原理的基础上, 重点介绍了三代测序技术在微生物16S/18S rRNA基因测序、单菌的基因组组装以及宏基因组中的研究应用和进展。

关键词: 微生物, 三代测序, 16S/18S rRNA, 宏基因组

Abstract:

Microbes are ubiquitous in human life. In years past, the study of microbes has only focused on single-bacteria cultures and qualitative analyses. The development of sequencing technology has greatly enhanced progress in microbiology research and more and more evidence shows that human symbiotic microbes, especially intestinal microbes, are closely related to human health. Second-generation sequencing technology is currently mainstream in microbiology research because of its high throughput, high accuracy and low cost. However, with the deepening complexity of research, the disadvantages of second-generation technology, i.e. short read length (< 450 bp), lead to subsequent challenges in data analysis and genome assembly, and limit the applicability to future research. In this context, the third-generation sequencing technology comes into being. The third-generation of sequencing (TGS) technology is also called single molecule sequencing. It directly carries out real-time sequencing of single DNA molecules without PCR amplifications. TGS technology significantly increases read length up to 2-10 kb or even 2.2 Mb. Because of its advantages of long read and no preference for GC, TGS provides a new method for full-length gene sequencing that facilitates the assembly of complete and reliable genome maps in microbes and that further reveals the diversity of microbial structures and functions. This review summarizes the technical characteristics and principles of TGS, and then mainly analyzes its applications and progress in 16S/18S rRNA gene sequencing, complete bacterial genome mapping and metagenomics research.

Key words: microbes, third-generation sequencing, 16S/18S rRNA, metagenomics