Improvement of single nucleotide polymorphism genotyping by allele-specific PCR using primers modified with an ENA residue
Improvement of single nucleotide polymorphism genotyping by allele-specific PCR using primers modified with an ENA residue
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DOI：
10.1016/j.ab.2005.02.029
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发表时间：
2005-05-15
影响因子：
2.9
通讯作者：
Kasuya, A
中科院分区：
文献类型：
--
作者：
Koizumi, M;Morita, K;Kasuya, A
当我们在引物的3 '端或n-1、n-2或n-3位插入ENA残基时，只有在n-2位含有ENA残基的引物才能被Tay DNA聚合酶读取，其中n-1、n-2或n-3位在3 '端含有单核苷酸多态性（SNP）位点。ENA引物的使用避免了不期望的短产物的产生，所述短产物被认为源自引物二聚体。与常用于等位基因特异性聚合酶链反应（AS-PCR）的相应未修饰的DNA引物相比，观察到这些含有ENA残基的引物对SNP位点的更大区分。这种改进可能是由于在修饰的引物-模板双链体中通过Taq DNA聚合酶将核苷酸掺入错配的ENA引物中的困难。这些结果表明，基于ENA引物的AS-PCR将能够实现SNP基因分型的快速和可靠的技术。(c)2005年爱思唯尔公司All rights reserved.
When we placed an ENA residue into primers at the 3 ' end, or the n-1, n-2, or n-3 position, Which included a single nucleotide polymorphism (SNP) site at the 3 ' end, only primers containing the ENA residue at the n-2 position were read by Tay DNA polymerase for amplification. The use of the ENA primers avoided the generation of undesired short products, which are thought to be derived from primer-dimers. A greater discrimination of the SNP site by these primers containing the ENA residue was observed compared with that of the corresponding unmodified DNA primers that are often used for allele-specific polymerase chain reaction (AS-PCR). This improvement is probably due to the difficulty of incorporating a nucleotide into the mismatched ENA primer by Taq DNA polymerase in the modified primer-template duplex. These results demonstrate that ENA primer-based AS-PCR would enable a rapid and reliable technique for SNP genotyping. (c) 2005 Elsevier Inc. All rights reserved.