核酸
清脆的
DNA
亚硫酸氢盐
DNA甲基化
核糖核酸
计算生物学
亚硫酸氢盐测序
分子诊断学
生物
分子生物学
生物化学
遗传学
基因
基因表达
作者
Linxian Li,Shiyuan Li,Na Wu,Jiacheng Wu,Gang Wang,Guoping Zhao,Jin Wang
标识
DOI:10.1021/acssynbio.9b00209
摘要
The next-generation CRISPR-based molecular diagnostics has the merits of rapidness, accuracy, and portability. We discovered the Cas12a trans-cleavage activity against collateral single-stranded DNA (ssDNA) and employed the activity to develop a rapid nucleic acid detection system, namely HOLMES (one-hour low-cost multipurpose highly efficient system). Here, with the employment of thermophilic CRISPR-Cas12b, we create HOLMESv2 for four different applications: (1) specifically discriminating single nucleotide polymorphism (SNP); (2) simply detecting virus RNA, human cell mRNA and circular RNA; (3) conveniently quantitating target nucleic acids with a one-step system combined with LAMP amplification in a constant temperature, thus avoiding cross-contamination; (4) accurately quantitating target DNA methylation degree with the combination of Cas12b detection and bisulfite treatment. These results highlight the potential of HOLMESv2 as a promising platform for both molecular diagnostics and epigenetics applications.
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