嗜热水生栖热菌DNA聚合酶催化的链式反应用于检测人乳头瘤病毒

Thermus aquaticus DNA polymerase-catalysed chain reaction for the detection of human papillomaviruses.

作者信息

Chow V T, Tham K M, Bernard H U

机构信息

Institute of Molecular and Cell Biology, National University of Singapore.

出版信息

J Virol Methods. 1990 Jan;27(1):101-12. doi: 10.1016/0166-0934(90)90150-e.

DOI:10.1016/0166-0934(90)90150-e

PMID:2155246

Abstract

To delineate the conditions for the polymerase chain reaction (PCR) using primers specific for human papillomavirus (HPV) types 6b, 16 and 18, a number of important technical features were analysed. Buffer, concentrations of magnesium, Taq polymerase, primers and DNA templates, annealing temperature, and extension time were studied by a combination of gel electrophoresis, Southern and slot-blot hybridization. Amplification of E6 gene fragments of HPV-16 and HPV-18 generated bands of 110 bp and 154 bp respectively, as predicted. However, amplification of a segment within the long control region of HPV 6b yielded an unexpected size of 340 bp. Different conditions were found for each HPV type-specific primer pair. These results, and the applications of PCR in HPV research and in an increasingly wide range of fields in medical virology are discussed.

摘要

为了确定使用针对人乳头瘤病毒（HPV）6b型、16型和18型的引物进行聚合酶链反应（PCR）的条件，分析了许多重要的技术特征。通过凝胶电泳、Southern杂交和狭缝印迹杂交相结合的方法，研究了缓冲液、镁离子浓度、Taq聚合酶、引物和DNA模板、退火温度以及延伸时间。如预期的那样，HPV-16和HPV-18的E6基因片段扩增分别产生了110 bp和154 bp的条带。然而，HPV 6b长控制区内一个片段的扩增产生了意想不到的340 bp大小。发现每种HPV型特异性引物对的条件不同。讨论了这些结果以及PCR在HPV研究和医学病毒学越来越广泛领域中的应用。