文献类型： 外文期刊
作者： Meng, Qingzhou 1 ; Wang, Xinjie 1 ; Wang, Yanqun 1 ; Dang, Lu 2 ; Liu, Xinyi 5 ; Ma, Xiaodong 4 ; Chi, Tian 6 ; Wang, Xia 1 ;
作者机构： 1.Guangzhou Med Univ, Natl Clin Res Ctr Resp Dis, Natl Ctr Resp Med,Guangzhou Inst Resp Hlth, Guangzhou Lab,State Key Lab Resp Dis,Affiliated H, Guangzhou 510120, Peoples R China
2.Guangzhou Med Univ, Affiliated Canc Hosp & Inst, Guangzhou, Peoples R China
3.ShanghaiTech Univ, Shanghai Inst Adv Immunochem Studies, Shanghai, Peoples R China
4.South China Normal Univ, Ctr Studies Psychol Applicat, Inst Brain Res & Rehabil, Guangdong Key Lab Mental Hlth & Cognit Sci, Guangzhou, Peoples R China
5.Chinese Acad Agr Sci, Agr Genom Inst Shenzhen, Shenzhen, Peoples R China
6.ShanghaiTech Univ, Sch Life Sci & Technol, Shanghai, Peoples R China
7.Northwest A&F Univ, Coll Vet Med, Dept Prevent Vet Med, Yangling, Shaanxi, Peoples R China
关键词： COVID-19; CRISPR; Cas12a; CRISPR-based detection; D614G; SARS-CoV-2
期刊名称：BIOTECHNOLOGY JOURNAL （ 影响因子：4.677； 五年影响因子：4.596 ）
ISSN： 1860-6768
年卷期：
页码：
收录情况： SCI
摘要： Detection of pathogens with single-nucleotide variations is indispensable for the disease tracing, but remains technically challenging. The D614G mutation in the SARS-CoV-2 spike protein is known to markedly enhance viral infectivity but is difficult to detect. Here, we report an effective approach called "synthetic mismatch integrated crRNA guided Cas12a detection" (symRNA-Cas12a) to detect the D614 and G614 variants effectively. Using this method, we systemically screened a pool of crRNAs that contain all the possible nucleotide substitutions covering the -2 to +2 positions around the mutation and identify one crRNA that can efficiently increase the detection specificity by 13-fold over the ancestral crRNA. With this selected crRNA, the symRNA-Cas12a assay can detect as low as 10 copies of synthetic mutant RNA and the results are confirmed to be accurate by Sanger sequencing. Overall, we have developed the symRNA-Cas12a method to specifically, sensitively and rapidly detect the SARS-CoV-2 D614G mutation.
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