Xiaohan Tong , Tianle Li , Kun Zhang , Dongming Zhao , Ying Zhang , Hao Yin
{"title":"Cas12a变异体的结构导向设计提高了核酸的检测","authors":"Xiaohan Tong , Tianle Li , Kun Zhang , Dongming Zhao , Ying Zhang , Hao Yin","doi":"10.1016/j.cellin.2025.100228","DOIUrl":null,"url":null,"abstract":"<div><div>CRISPR-Cas12a holds promising potential for pathogen detection. However, its performance is not optimal when combined with isothermal amplification. Hence, we engineered a mutant of LbCas12a (K595A) with reduced <em>cis</em>-cleavage activity, to minimize interference with isothermal amplification. Compared to wild-type Cas12a, the K595A mutant exhibited a 2–3 times faster reaction speed and a 1,000–10,000 times increase in sensitivity in a one-pot reaction. We applied this mutant for detection of African Swine Fever Virus (ASFV). This K595A mutant successfully detected all 30 ASFV samples within 20 minutes. Our study suggests a universal approach to improve the performance of Cas12a for pathogen detection.</div></div>","PeriodicalId":72541,"journal":{"name":"Cell insight","volume":"4 2","pages":"Article 100228"},"PeriodicalIF":6.4000,"publicationDate":"2025-04-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":"{\"title\":\"Structure-Guided design of Cas12a variants improves detection of nucleic acids\",\"authors\":\"Xiaohan Tong , Tianle Li , Kun Zhang , Dongming Zhao , Ying Zhang , Hao Yin\",\"doi\":\"10.1016/j.cellin.2025.100228\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"<div><div>CRISPR-Cas12a holds promising potential for pathogen detection. However, its performance is not optimal when combined with isothermal amplification. Hence, we engineered a mutant of LbCas12a (K595A) with reduced <em>cis</em>-cleavage activity, to minimize interference with isothermal amplification. Compared to wild-type Cas12a, the K595A mutant exhibited a 2–3 times faster reaction speed and a 1,000–10,000 times increase in sensitivity in a one-pot reaction. We applied this mutant for detection of African Swine Fever Virus (ASFV). This K595A mutant successfully detected all 30 ASFV samples within 20 minutes. Our study suggests a universal approach to improve the performance of Cas12a for pathogen detection.</div></div>\",\"PeriodicalId\":72541,\"journal\":{\"name\":\"Cell insight\",\"volume\":\"4 2\",\"pages\":\"Article 100228\"},\"PeriodicalIF\":6.4000,\"publicationDate\":\"2025-04-01\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"\",\"citationCount\":\"0\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"Cell insight\",\"FirstCategoryId\":\"1085\",\"ListUrlMain\":\"https://www.sciencedirect.com/science/article/pii/S2772892725000021\",\"RegionNum\":0,\"RegionCategory\":null,\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"2025/1/20 0:00:00\",\"PubModel\":\"Epub\",\"JCR\":\"\",\"JCRName\":\"\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"Cell insight","FirstCategoryId":"1085","ListUrlMain":"https://www.sciencedirect.com/science/article/pii/S2772892725000021","RegionNum":0,"RegionCategory":null,"ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"2025/1/20 0:00:00","PubModel":"Epub","JCR":"","JCRName":"","Score":null,"Total":0}
Structure-Guided design of Cas12a variants improves detection of nucleic acids
CRISPR-Cas12a holds promising potential for pathogen detection. However, its performance is not optimal when combined with isothermal amplification. Hence, we engineered a mutant of LbCas12a (K595A) with reduced cis-cleavage activity, to minimize interference with isothermal amplification. Compared to wild-type Cas12a, the K595A mutant exhibited a 2–3 times faster reaction speed and a 1,000–10,000 times increase in sensitivity in a one-pot reaction. We applied this mutant for detection of African Swine Fever Virus (ASFV). This K595A mutant successfully detected all 30 ASFV samples within 20 minutes. Our study suggests a universal approach to improve the performance of Cas12a for pathogen detection.
京公网安备 11010802042870号
京ICP备2023020795号-1
分享
求助内容：
应助结果提醒方式：
扫码关注我们
