A general solution for opening double-stranded DNA for isothermal amplification.
A general solution for opening double-stranded DNA for isothermal amplification.
复制标题
打开双链 DNA 进行等温扩增的通用解决方案
DOI：
10.1038/srep34582
复制
发表时间：
2016-09-30
影响因子：
4.6
通讯作者：
Tang Z
中科院分区：
文献类型：
--
作者：
Chen G;Dong J;Yuan Y;Li N;Huang X;Cui X;Tang Z
核酸扩增是分子生物学和基因工程的核心技术。已经开发了各种等温扩增技术作为聚合酶链反应（PCR）的替代。然而，这些方法中的大多数只能检测单链核酸。在此，我们提出了一个简单的解决方案打开双链DNA的等温检测方法。该策略利用弗罗姆.大肠杆菌（RecA）与单链DNA形成核蛋白复合物，可扫描双链模板寻找同源位点。然后，在ATP存在下，核蛋白可以侵入双链模板形成异源双链体，导致链交换。高浓度的ATP可消除ATP再生系统，而入侵链的3′-OH端可被其他DNA修饰酶如DNA聚合酶或DNA连接酶识别。此外，dATP被发现是RecA的一个更好的辅因子，这使得该系统与DNA聚合酶更相容。本文所述方法是开放dsDNA的通用解决方案，为在此基础上开发更多的真实的DNA样品等温核酸检测方法提供了平台。
Nucleic acid amplification is the core technology of molecular biology and genetic engineering. Various isothermal amplification techniques have been developed as alternatives to polymerase chain reaction (PCR). However, most of these methods can only detect single stranded nucleic acid. Herein, we put forward a simple solution for opening double-stranded DNA for isothermal detection methods. The strategy employs recombination protein fromE. coli(RecA) to form nucleoprotein complex with single-stranded DNA, which could scan double-stranded template for homologous sites. Then, the nucleoprotein can invade the double-stranded template to form heteroduplex in the presence of ATP, resulting in the strand exchange. The ATP regeneration system could be eliminated by using high concentration of ATP, and the 3′-OH terminal of the invasion strand can be recognized by other DNA modifying enzymes such as DNA polymerase or DNA ligase. Moreover, dATP was found to be a better cofactor forRecA, which make the system more compatible to DNA polymerase. The method described here is a general solution to open dsDNA, serving as a platform to develop more isothermal nucleic acids detection methods for real DNA samples based on it.
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影响因子：
1.5
作者：
POOLE, CB;WILLIAMS, SA
通讯作者：
WILLIAMS, SA
影响因子：
2.7
作者：
OHTANI, T;SHIBATA, T;ANDO, T
通讯作者：
ANDO, T
DOI：
10.1073/pnas.78.6.3433
发表时间：
1981-01-01
期刊：
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA-BIOLOGICAL SCIENCES
影响因子：
--
作者：
COX, MM;LEHMAN, IR
通讯作者：
LEHMAN, IR
影响因子：
2.9
作者：
MORRICAL, SW;LEE, J;COX, MM
通讯作者：
COX, MM
影响因子：
64.8
作者：
通讯作者：
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