一种基于聚合酶链式反应检测SNP的方法

SNP是具有广泛利用潜力的第3代分子标记,本文旨在开发一种利用PCR技术快速检测SNP的方法。设计思路是:根据已知SNP位点设计2条特异正向引物,其最后一个碱基分别与已知SNP的2个碱基相同,同时在1条引物的5′端添加1段20 bp左右的其他物种的特异序列(如细菌DNA序列),然后选择1条合适的反向引物;最后同时加入3条引物,通过梯度PCR选择合适的退火温度进行PCR反应。利用这一方法成功将玉米的ZDS基因定位在玉米第7染色体短臂7.02 Bin。这种检测SNP的方法设计简单,费用低廉,尤其适合SNP标记的分子标记连锁图构建或者基因定位。

Method for detecting SNP based on PCR technique

SNP is one kind of powerful molecular markers.The purpose in this study was to establish a simple method of detecting SNP based on PCR technique.Two specific forward primers were synthesized,both were in conformity with the found SNP in the last nucleotide.A 19 bp pecicific sequence was added into one primer with other species' sequence(eg.Bacteria DNA sequence) in 5' end.The same reversed primer was selected using the software Primer 5.0.The PCR was performed with the three specific primers using an appropriate anneal temperature selected by gradient PCR.The ZDS gene has been mapped on the short arm of chromosome 7 in maize based on this method.This technique is simple,effective and cheap,and could be helpful in constructing linkage maps and map genes.