Engelke D R, Hoener P A, Collins F S
Department of Biological Chemistry, University of Michigan Medical School, Ann Arbor 48109.
Proc Natl Acad Sci U S A. 1988 Jan;85(2):544-8. doi: 10.1073/pnas.85.2.544.
The polymerase chain reaction is a recently described technique that uses flanking oligonucleotide primers and repeated cycles of enzymatic primer extension to amplify a short segment of DNA by greater than 100,000-fold. By use of sequencing primers located internal to the amplification primers, direct genomic sequence was obtained from enzymatically amplified DNA by using the dideoxynucleotide chain-termination method. The method is relatively simple and offers significant advantages in identifying mutations in genes for which the normal sequence is known. Heterozygous and homozygous mutations in the human beta- and gamma-globin loci were unambiguously identified in 3 days with less than 1 microgram of genomic DNA.
聚合酶链反应是一种最近描述的技术，它使用侧翼寡核苷酸引物和酶促引物延伸的重复循环，将一小段DNA扩增超过100,000倍。通过使用位于扩增引物内部的测序引物，利用双脱氧核苷酸链终止法从酶促扩增的DNA中获得直接的基因组序列。该方法相对简单，在鉴定已知正常序列的基因中的突变方面具有显著优势。用不到1微克的基因组DNA在3天内就能明确鉴定出人类β和γ珠蛋白基因座中的杂合和纯合突变。
