A method of genotyping single nucleotide polymorphisms (“SNP”) and point mutations in nucleic acid based on chain extension by polymerase. This invention is based on the fact that the nucleoside immediately 5′ adjacent to any SNP/point mutation site is known, and the neighboring sequence immediately 3′ adjacent to the site is also known. A primer complementary to the sequence directly adjacent to the SNP on the 3′ side in a target polynucleotide is used for chain elongation. The polymerase reaction mixture contains one chain-terminating nucleotide having a base complementary to the nucleotide directly adjacent to the SNP on the 5′ side in the target polynucleotide. An additional dNTP may be added to produce a primer with the maximum of a two-base extension. The resultant elongation/termination reaction products are analyzed for the length of chain extension of the primer, or for the amount of label incorporation from a labeled form of the terminator nucleotide.
一种基于聚合酶链延伸对核酸中单核苷酸多态性("SNP")和点突变进行
基因分型的方法。本发明基于以下事实:与任何 SNP/点突变位点紧邻 5′的核苷是已知的,与该位点紧邻 3′的邻接序列也是已知的。与目标多核苷酸 3′侧 SNP 直接相邻序列互补的引物用于核苷酸链的延伸。聚合酶反应混合物中含有一个链终止核苷酸,该核苷酸的碱基与目标多核苷酸 5′侧与 SNP 直接相邻的核苷酸互补。可添加额外的 dNTP,以产生最大延伸两个碱基的引物。对由此产生的延伸/终止反应产物进行分析,以确定引物的链延伸长度或终止核苷酸标记形式的标记结合量。