Guo Zi-fen, Chen Lin-ling, Zhang Jia, Peng Cui-ying, Yang Xiang-dong, Zhang Xu, He Shu-ya, Liao Duan-fang, Li Kai
Department of Pharmacy and Pharmacology, Nanhua University, Hengyang, Hunan, 421001 PR China.
Zhonghua Yi Xue Yi Chuan Xue Za Zhi. 2003 Aug;20(4):328-30.
To determine whether 3'phosphorothioate-modified-2 terminal mismatched primers can turn off DNA polymerization mediated by Exo(+) polymerase.
Two-directional primer extension was performed using polymerase with and without 3' exonuclease activity. The effects of unmodified primers and 3' phosphorothioate-modified primers on primer extension were evaluated.
Exo(-) polymerase yielded products from matched and mismatched primers regardless of their modification. However, 3' phosphorothioate-modified primers with a single base mismatch at -2 position worked similarly to the terminal (-1) mismatched primers in triggering the novelly reported "off-switch" of Exo(+) polymerase.
These data suggested that the "off-switch" can be of enormous application in the diagnosis of single gene diseases and in the association studies by single nucleotide polymorphism screening.
确定3'-硫代磷酸酯修饰的2'-末端错配引物是否能关闭由具有3'-外切核酸酶活性的聚合酶介导的DNA聚合反应。
使用具有和不具有3'-外切核酸酶活性的聚合酶进行双向引物延伸。评估未修饰引物和3'-硫代磷酸酯修饰引物对引物延伸的影响。
无论引物是否修饰，无3'-外切核酸酶活性的聚合酶均能从匹配和错配引物产生产物。然而，在-2位置具有单个碱基错配的3'-硫代磷酸酯修饰引物在触发新报道的具有3'-外切核酸酶活性的聚合酶的“关闭开关”方面，与末端（-1）错配引物的作用相似。
这些数据表明，该“关闭开关”在单基因疾病诊断和单核苷酸多态性筛查的关联研究中具有巨大的应用价值。
