Robledo Renato, Beggs William R, Bender Patrick K
Department of Biology, University of Cagliari, Italy.
Methods Mol Biol. 2005;311:165-76. doi: 10.1385/1-59259-957-5:165.
The 5' fluorogenic (TaqMan) assay has been successfully used in screening for single-nucleotide polymorphisms; the very few steps required and the ability to automate each step allow for high-throughput screening. Insertion/deletion polymorphisms are an important class of markers that can be studied for different applications, such as diagnostics, genome variation, and species identification. Polymerase chain reaction (PCR) and post-PCR analysis are required to score the insertion or the deletion allele. In this chapter, we describe an expansion of the TaqMan technology for a rapid, high-throughput, screening for insertion/deletion polymorphisms in which the exact endpoints are known. The method requires minimal post-PCR analysis and can be applied to polymorphisms of any size.
5'荧光(TaqMan)分析法已成功用于单核苷酸多态性的筛选;所需步骤极少且能够实现每个步骤的自动化,这使得高通量筛选成为可能。插入/缺失多态性是一类重要的标记,可用于不同的应用研究,如诊断、基因组变异和物种鉴定。需要通过聚合酶链反应(PCR)和PCR后分析来确定插入或缺失等位基因。在本章中,我们描述了TaqMan技术的扩展应用,用于快速、高通量筛选已知确切终点的插入/缺失多态性。该方法所需的PCR后分析极少,可应用于任何大小的多态性。