生物学杂志

生物学杂志 ›› 2022, Vol. 39 ›› Issue (3): 107-.doi: 10.3969/j.issn. 2095-1736.2022.03.107

• 技术方法 • 上一篇    下一篇

利用连接酶检测反应检测碎片化新冠病毒S基因D614G突变

  

  1. 1. 江苏海洋大学 江苏省海洋药物活性分子筛选重点实验室， 连云港 222005；
    2. 江苏省海洋资源开发研究院（连云港）， 连云港 222005
  • 出版日期:2022-06-18 发布日期:2022-06-17
  • 通讯作者: 罗志丹，博士，讲师，研究方向为分子生物学，E-mail: lzd_111@163.com
  • 作者简介:张新亚，硕士研究生，研究方向为分子诊断学，E-mail: Zhang_xinya6@163.com
  • 基金资助:
    国家自然科学基金项目（31501763）

Detection of D614 mutation in fragmented S gene of SARS-CoV-2 by ligation detection reaction

  1. 1.Jiangsu Key Laboratory of Marine Pharmaceutical Compound Screening, Jiangsu Ocean University,
    Lianyungang 222005, China; 2. Jiangsu Institute of Marine Resource Development (Lianyungang),
    Lianyungang 222005, China
  • Online:2022-06-18 Published:2022-06-17

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摘要： 新型冠状病毒 (SARS-CoV-2) S基因D614突变导致病毒感染能力显著增强，也是德尔塔变异毒株的主要来源。如何在病毒核酸检测时对此类高危SNP进行快速分型是研究者非常关注的技术问题。但新冠病毒属于单链RNA病毒，基因组易降解，现有的点突变鉴定技术并不适用高度降解的碎片化病毒核酸。研究利用连接酶检测反应 (LDR) 技术结合荧光定量PCR，建立一种检测碎片化新冠病毒S基因D614G的方法。结果表明：方法对5 pmol/L的病毒RNA模板具有良好的区分度，最低可检测出长度仅为40 nt的病毒RNA所携带的点突变。

关键词: 连接酶检测反应, 新冠病毒, 碎片化, D614G突变

Abstract: The D614 mutation of the S gene of the new coronavirus (SARS-CoV-2) significantly increases the virus' infectivity, and is also the main source of the delta mutant strain. How to quickly type such high-risk SNPs in viral nucleic acid detection is a technical issue that researchers are very concerned about. However, the new coronavirus is a single-stranded RNA virus, and the genome is easily degraded. The existing point mutation identification technology is not suitable for highly degraded fragmented viral nucleic acid. In this study, a ligase detection reaction (LDR) technology combined with fluorescent quantitative PCR was used to establish a method to detect the fragmented new coronavirus S gene D614G. The results showed that this method had a good degree of discrimination for 5 pmol/L viral RNA templates, and the minimum point mutations carried by viral RNAs with a length of only 40 nt could be detected.

Key words: LDR, SARS-CoV-2, fragmentation, D614G mutation

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