CRISPR-Cas12a-assisted nucleic acid detection.
CRISPR-Cas12a-assisted nucleic acid detection.
复制标题
CRISPR-Cas12a辅助核酸检测
DOI：
10.1038/s41421-018-0028-z
复制
发表时间：
2018
期刊：
影响因子：
33.5
通讯作者：
Wang J
中科院分区：
文献类型：
--
作者：
Li SY;Cheng QX;Wang JM;Li XY;Zhang ZL;Gao S;Cao RB;Zhao GP;Wang J
今天，在人类基因分型和病原体检测等领域，对时间有效和成本有效的核酸检测方法的需求仍在增长。使用合成的生物分子组分，已经开发了许多用于快速核酸检测的方法1-3;然而，它们可能无法同时满足特异性、灵敏度、速度、成本和简单性。最近，建立了一种非常有前途的基于CRISPR的诊断（CRISPR-Dx）（即SHERRP 4），其基于RNA引导和RNA靶向CRISPR效应子Cas 13 a 4的附带效应。该方法具有灵敏度高、特异性强、操作简便等优点。然而，为了检测DNA序列，必须在SHERPRK测试之前进行DNA到RNA的体外转录，这可能是不方便的。在最近的一项研究中，我们发现Cas 12 a，属于2类VA CRISPR-Cas系统5，在形成Cas 12 a/crRNA/靶DNA三元复合物6时，对非靶向ssDNA进行侧切。在此，利用该特征，我们使用淬灭荧光ssDNA报告基因（例如，补充表S1中的HEX-N12-BHQ 1）作为探针，并开发了HOLMES（一种一小时低成本多用途高效系统），其可用于快速检测靶DNA和靶RNA。在HOLMES中，如果反应体系中存在靶DNA，则Cas 12 a/crRNA二元复合物与靶DNA形成三元复合物，然后将
Dear Editor, Today, the need for time-effective and cost-effective nucleic acid detection methods is still growing in fields such as human genotyping and pathogen detection. Using synthetic biomolecular components, many methods have been developed for fast nucleic acid detection 1–3; however, they may not be able to satisfy specificity, sensitivity, speed, cost and simplicity at the same time. Recently, a very promising CRISPR-based diagnostic (CRISPR-Dx)(namely SHERLOCK) was established, which was based on the collateral effect of an RNA-guided and RNA-targeting CRISPR effector, Cas13a 4. SHERLOCK is of high sensitivity and specificity, and is very convenient in detection of target RNA. However, to detect DNA sequences, in vitro transcription of DNA to RNA must be conducted prior to the SHERLOCK test, which could be inconvenient.In a recent study, we found that Cas12a, which belongs to the class 2 type VA CRISPR-Cas system 5, performed collateral cleavage on non-targeted ssDNAs upon the formation of the Cas12a/crRNA/target DNA ternary complex 6. Here, with the employment of this feature, we used a quenched fluorescent ssDNA reporter (eg, HEX-N12-BHQ1 in Supplementary Table S1) as the probe, and developed HOLMES (an one-HOur Low-cost Multipurpose highly Efficient System), which could be used for fast detection of target DNA as well as target RNA. In HOLMES, if a target DNA exists in the reaction system, the Cas12a/crRNA binary complex forms a ternary complex with the target DNA, which will then
登录
查看更多内容
影响因子：
14.9
作者：
Lei C;Li SY;Liu JK;Zheng X;Zhao GP;Wang J
通讯作者：
Wang J
影响因子：
64.5
作者：
Pardee K;Green AA;Ferrante T;Cameron DE;DaleyKeyser A;Yin P;Collins JJ
通讯作者：
Collins JJ
影响因子：
64.5
作者：
Yamano T;Nishimasu H;Zetsche B;Hirano H;Slaymaker IM;Li Y;Fedorova I;Nakane T;Makarova KS;Koonin EV;Ishitani R;Zhang F;Nureki O
通讯作者：
Nureki O
影响因子：
64.5
作者：
Zetsche B;Gootenberg JS;Abudayyeh OO;Slaymaker IM;Makarova KS;Essletzbichler P;Volz SE;Joung J;van der Oost J;Regev A;Koonin EV;Zhang F
通讯作者：
Zhang F
影响因子：
64.5
作者：
Pardee, Keith;Green, Alexander A.;Collins, James J.
通讯作者：
Collins, James J.