Development of an inducible Cas9 nickase and PAM-free Cas12a platform for bacterial diagnostics

doi:10.1016/j.talanta.2023.124931

	Development of an inducible Cas9 nickase and PAM-free Cas12a platform for bacterial diagnostics
	Hu, Yuanzhao 1; Qiao, Yuefeng 1; Li, Xiu-Qing 2,3; Xiang, Zhenbo5 ; Wan, Yi 1; Wang, Peng4 ; Yang, Zhiqing 1,5
	2023-12-01
发表期刊	TALANTA
ISSN	0039-9140
卷号	265 页码:6
通讯作者	Wang, Peng(wangpeng@qdio.ac.cn) ; Yang, Zhiqing(yangzhiqing@hainanu.edu.cn)
摘要	Rapid, efficient, specific and sensitive diagnostic techniques are critical for selecting appropriate treatments for drug-resistant bacterial infections. To address this challenge, we have developed a novel diagnostic method, called the Dual-Cas Tandem Diagnostic Platform (DTDP), which combines the use of Cas9 nickase (Cas9n) and Cas12a. DTDP works by utilizing the Cas9n-sgRNA complex to create a nick in the target strand's doublestranded DNA (dsDNA). This prompts DNA polymerase to displace the single-stranded DNA (ssDNA) and leads to cycles of DNA replication through nicking, displacement, and extension. The ssDNA is then detected by the Cas12a-crRNA complex (which is PAM-free), activating trans-cleavage and generating a fluorescent signal from the fluorescent reporter. DTDP exhibits a high sensitivity (1 CFU/mL or 100 ag/& mu;L), high specificity (specifically to MRSA in nine pathogenic species), and excellent accuracy (100%). The dual RNA recognition process in our method improves diagnostic specificity by decreasing the limitations of Cas12a in detecting dsDNA protospacer adjacent motifs (PAMs) and leverages multiple advantages of multi-Cas enzymes in diagnostics. This novel approach to pathogenic microorganism detection has also great potential for clinical diagnosis.
关键词	Bacteria diagnostics CRISPR-Cas9n CRISPR-Cas12a Strand displacement
DOI	10.1016/j.talanta.2023.124931
收录类别	SCI
语种	英语
资助项目	National Key Ramp;D Program of China[2022YFD2401304] ; Hainan Province Science and Technology Special Fund[ZDYF2023XDNY074] ; Research Foundation of Hainan University ; Hainan Provincial Nat-ural Science Foundation of China[322QN228] ; Hainan Provincial Nat-ural Science Foundation of China[KYQD (ZR) 1997] ; [322MS032]
WOS研究方向	Chemistry
WOS类目	Chemistry, Analytical
WOS记录号	WOS:001048130900001
出版者	ELSEVIER
WOS关键词	INFECTIOUS-DISEASES ; LIMITATIONS ; ADVANTAGES
引用统计
文献类型	期刊论文
条目标识符	http://ir.qdio.ac.cn/handle/337002/182808
专题	海洋环境腐蚀与生物污损重点实验室
通讯作者	Wang, Peng; Yang, Zhiqing
作者单位	1.Hainan Univ, Marine Coll, State Key Lab Marine Resource Utilizat South China, Haikou 570228, Peoples R China 2.Agr & Agrifood Canada, Fredericton, NB E3B 4Z7, Canada 3.Nutra Hlth Prod & Technol Inc, Fredericton, NB E3B 6J5, Canada 4.Chinese Acad Sci, Inst Oceanol, CAS Key Lab Marine Environm Corros & Biofouling, Qingdao 266071, Peoples R China 5.Rizhao Sci & Technol Innovat Serv Ctr, 369 Jining Rd, Rizhao, Shandong, Peoples R China
通讯作者单位	中国科学院海洋研究所
推荐引用方式 GB/T 7714	Hu, Yuanzhao,Qiao, Yuefeng,Li, Xiu-Qing,et al. Development of an inducible Cas9 nickase and PAM-free Cas12a platform for bacterial diagnostics[J]. TALANTA,2023,265:6.
APA	Hu, Yuanzhao.,Qiao, Yuefeng.,Li, Xiu-Qing.,Xiang, Zhenbo.,Wan, Yi.,...&Yang, Zhiqing.(2023).Development of an inducible Cas9 nickase and PAM-free Cas12a platform for bacterial diagnostics.TALANTA,265,6.
MLA	Hu, Yuanzhao,et al."Development of an inducible Cas9 nickase and PAM-free Cas12a platform for bacterial diagnostics".TALANTA 265(2023):6.

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