中国媒介生物学及控制杂志 ›› 2020, Vol. 31 ›› Issue (6): 735-737.DOI: 10.11853/j.issn.1003.8280.2020.06.024

• 调查研究 • 上一篇    下一篇

高通量半自动化细菌全基因组序列测定体系的构建

张景山1, 龙政2, 王紫鉴2, 肖文静1, 王群1, 赵宏群1   

  1. 1 中国疾病预防控制中心传染病预防控制所生物安全实验室, 北京 102206;
    2 河北北方学院, 河北 张家口 075000
  • 收稿日期:2020-06-11 出版日期:2020-12-20 发布日期:2020-12-20
  • 通讯作者: 赵宏群,Email:zhaohongqun@icdc.cn
  • 作者简介:张景山,男,主管技师,主要从事疾病控制工作,Email:zhangjingshan@icdc.cn

Establishment of a high-throughput semi-automatic bacterial whole-genome sequencing system

ZHANG Jing-shan1, LONG Zheng2, WANG Zi-jian2, XIAO Wen-jing1, WANG Qun1, ZHAO Hong-qun1   

  1. 1 National Institute for Communicable Disease Control and Prevention, Chinese Center for Disease Control and Prevention, Beijing 102206, China;
    2 Hebei North University
  • Received:2020-06-11 Online:2020-12-20 Published:2020-12-20

摘要: 目的 建立高通量、半自动化细菌全基因组序列测定体系,满足基于基因组序列的细菌性病原体监测的需求。方法 使用自动化液体处理器、可丢弃性机械手枪头、96孔板型核酸提取试剂盒、自动测序文库构建系统以及测序仪,实现细菌基因组序列测定的半自动化。结果 机械手枪头与加样孔底的距离、液体混合速度和每次混匀液体的体积比3个因素与样品之间的交叉污染密切相关,机械手枪头与加样孔底的距离为2 mm、液体混合速度为全速的15%、每次混匀液体的体积比为50%时污染率为0。与手工方法相比,高通量半自动化细菌全基因组序列测定体系的工作效率提高了4倍。结论 建立的高通量、半自动化细菌基因组序列测定体系,能够满足基于基因组序列的细菌性病原体监测的需求。

关键词: 自动化液体处理, 交叉污染, 全基因组序列测定, 病原体网络化监测

Abstract: Objective To establish a high-throughput semi-automatic bacterial whole-genome sequencing system to meet the needs of bacterial pathogen surveillance based on the genome sequence. Methods An automated liquid handler, disposable mechanical pipette tips, a 96-well nucleic acid extraction kit, an automated sequencing library construction system, and a sequencer were used to realize the semi-automation of bacterial genome sequencing. Results The distance between the mechanical pipette tip and the bottom of the sample well, the mixing speed of liquid, and the volume ratio of the mixed liquid to the sample each time were closely related to the cross-contamination of the samples. When the distance between the mechanical pipette tip and the bottom of the sample well was 2 mm, the mixing speed of liquid was 15% of the full speed, and the volume ratio of the mixed liquid to the sample was 50% each time, the contamination rate was 0. Compared with the manual procedure, this high-throughput semi-automatic bacterial whole-genome sequencing system improved the work efficiency by 4-fold. Conclusion A high-throughput semi-automatic bacterial whole-genome sequencing system is established, which can meet the needs of bacterial pathogen surveillance based on the genome sequence.

Key words: Automated liquid handling, Cross-contamination, Whole-genome sequencing, Pathogen surveillance network

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