Oligonucleotide inhibitors of Taq DNA polymerase facilitate detection of low copy number targets by PCR
Oligonucleotide inhibitors of Taq DNA polymerase facilitate detection of low copy number targets by PCR
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DOI：
10.1006/jmbi.1996.0640
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发表时间：
1996-11-29
影响因子：
5.6
通讯作者：
Jayasena, SD
中科院分区：
文献类型：
--
作者：
Dang, C;Jayasena, SD
通过筛选单链DNA随机序列文库，分离出对水热菌DNA聚合酶（Tag pol）高亲和力的序列，Tag pol是一种常见于聚合酶链反应（PCR）的耐热酶。所选择的寡核苷酸序列与Taq pol结合，解离常数在低皮摩尔范围内，在室温（20 ~ 25℃）下有效抑制聚合酶活性，但在40℃以上的温度下没有抑制作用，并且抑制作用是热可逆的。一种称为“热启动”PCR的过程通常用于在扩增低拷贝数目标时防止非特异性PCR产物。我们表明，添加寡核苷酸抑制剂消除了对“热启动”条件的需要，并提高了PCR中低拷贝数目标的检测效率。(C) 1996学术出版社有限公司
A random sequence Library of single stranded DNA was screened to isolate sequences with high affinity for Thermus aquaticus DNA polymerase (Tag pol), a thermostable enzyme commonly used in the polymerase chain reaction (PCR). Selected oligonucleotide sequences bound Taq pol with dissociation constants in the low picomolar range, and efficiently inhibited polymerase activity at room temperature (20 to 25 degrees C), but did not inhibit at temperatures above 40 degrees C. Moreover, inhibition was thermally reversible. A process called ''hot start-'' PCR is commonly used to prevent non-specific PCR products in amplification of low copy number targets. We show that the addition of oligonucleotide inhibitors eliminated the need for ''hot start'' conditions and improved the efficiency of detection of a low copy number target in PCR. (C) 1996 Academic Press Limited