使用Primer Express®软件进行基于SYBR Green的定量PCR引物设计。

Primer design using Primer Express® for SYBR Green-based quantitative PCR.

作者信息

Singh Amarjeet, Pandey Girdhar K

机构信息

Department of Plant Molecular Biology, University of Delhi South Campus, Benito Juarez Road, Dhaula Kuan, New Delhi, 110021, India.

出版信息

Methods Mol Biol. 2015;1275:153-64. doi: 10.1007/978-1-4939-2365-6_11.

DOI:10.1007/978-1-4939-2365-6_11

PMID:25697658

Abstract

To quantitate the gene expression, real-time RT-PCR or quantitative PCR (qPCR) is one of the most sensitive, reliable, and commonly used methods in molecular biology. The reliability and success of a real-time PCR assay depend on the optimal experiment design. Primers are the most important constituents of real-time PCR experiments such as in SYBR Green-based detection assays. Designing of an appropriate and specific primer pair is extremely crucial for correct estimation of transcript abundance of any gene in a given sample. Here, we are presenting a quick, easy, and reliable method for designing target-specific primers using Primer Express(®) software for real-time PCR (qPCR) experiments.

摘要

为了定量基因表达，实时逆转录聚合酶链反应（RT-PCR）或定量聚合酶链反应（qPCR）是分子生物学中最灵敏、可靠且常用的方法之一。实时PCR检测的可靠性和成功与否取决于优化的实验设计。引物是实时PCR实验（如基于SYBR Green的检测分析）中最重要的组成部分。设计一对合适且特异的引物对于正确估计给定样品中任何基因的转录本丰度极为关键。在此，我们介绍一种使用Primer Express®软件进行实时PCR（qPCR）实验的目标特异性引物设计的快速、简便且可靠的方法。