A PAM-free CRISPR/Cas12a ultra-specific activation mode based on toehold-mediated strand displacement and branch migration

作者全名："Wu, You; Luo, Wang; Weng, Zhi; Guo, Yongcan; Yu, Hongyan; Zhao, Rong; Zhang, Li; Zhao, Jie; Bai, Dan; Zhou, Xi; Song, Lin; Chen, Kena; Li, Junjie; Yang, Yujun; Xie, Guoming"

作者地址："[Wu, You; Luo, Wang; Weng, Zhi; Yu, Hongyan; Zhao, Rong; Zhang, Li; Bai, Dan; Zhou, Xi; Song, Lin; Chen, Kena; Li, Junjie; Yang, Yujun; Xie, Guoming] Chongqing Med Univ, Coll Lab Med, Chongqing Med Lab Microfluid, Key Lab Clin Lab Diagnost,Chinese Minist Educ, Chongqing 400016, Peoples R China; [Wu, You; Luo, Wang; Weng, Zhi; Yu, Hongyan; Zhao, Rong; Zhang, Li; Bai, Dan; Zhou, Xi; Song, Lin; Chen, Kena; Li, Junjie; Yang, Yujun; Xie, Guoming] Chongqing Med Univ, SPRi Engn Res Ctr, Chongqing 400016, Peoples R China; [Guo, Yongcan] Southwest Med Univ, Clin Lab, Tradit Chinese Med Hosp, Luzhou 646000, Peoples R China; [Zhao, Jie] Chongqing Med Univ, Clin Mol Med Testing Ctr, First Hosp, 1 Youyi Rd, Chongqing 400016, Peoples R China"

通信作者："Li, JJ; Yang, YJ; Xie, GM (通讯作者)，Chongqing Med Univ, Coll Lab Med, Chongqing Med Lab Microfluid, Key Lab Clin Lab Diagnost,Chinese Minist Educ, Chongqing 400016, Peoples R China.; Li, JJ; Yang, YJ; Xie, GM (通讯作者)，Chongqing Med Univ, SPRi Engn Res Ctr, Chongqing 400016, Peoples R China."

来源：NUCLEIC ACIDS RESEARCH

ESI学科分类：BIOLOGY & BIOCHEMISTRY

WOS号：WOS:000877183900001

JCR分区：Q1

影响因子：14.9

年份：2022

卷号：　

期号：　

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结束页：　

文献类型：Article; Early Access

关键词：　

摘要："CRISPR (clustered regularly interspaced short palindromic repeats) technology has achieved great breakthroughs in terms of convenience and sensitivity; it is becoming the most promising molecular tool. However, only two CRISPR activation modes (single and double stranded) are available, and they have specificity and universality bottlenecks that limit the application of CRISPR technology in high-precision molecular recognition. Herein, we proposed a novel CRISPR/Cas12a unrestricted activation mode to greatly improve its performance. The new mode totally eliminates the need for a protospacer adjacent motif and accurately activates Cas12a through toehold-mediated strand displacement and branch migration, which is highly universal and ultra-specific. With this mode, we discriminated all mismatch types and detected the EGFR T790M and L858R mutations in very low abundance. Taken together, our activation mode is deeply incorporated with DNA nanotechnology and extensively broadens the application boundaries of CRISPR technology in biomedical and molecular reaction networks."

基金机构："National Natural Science Foundation of China [82172369, 81972025]; Natural Science Foundation of Chongqing, China [cstc2021jcyj-msxmX0326, CYB22219]"

基金资助正文："This research work was financially supported by the National Natural Science Foundation of China [82172369 and 81972025]; the Natural Science Foundation of Chongqing, China [cstc2021jcyj-msxmX0326]; and theChongqing Postgraduate Research Innovation Project [CYB22219]."