应用荧光定量比较Ct值法测定基因相对表达量.docVIP

  应用荧光定量比较 Ct 值法测定基因相对表 达量# 魏洁书，杨锦芬** 5 10 （广州中医药大学中药资源科学与工程研究中心，岭南中药资源教育部重点实验室（广州中 医药大学），广州 510006） 摘要：实时荧光定量 PCR 是一种准确快速的核酸定量分析技术，具有特异性强、灵敏度高、 重复性好、定量准确、速度快、全封闭反应等优点。比较 Ct 值法是利用荧光定量 PCR 测定 基因相对表达量常用的方法，运用比较 Ct 值法进行基因相对表达量的分析，可在确定扩增 效率的前提下，对所要测定的样品的目的基因及内参基因同时进行扩增，大大减少实验工作 量，且得到的实验结果比较可靠，是一种经济实用，准确合理的实验方法。本文通过实验操 作的具体说明，介绍运用比较 Ct 值法测定基因相对表达量的关键步骤和技术细节。 关键词：荧光定量 PCR；比较 Ct 值法；相对定量；基因表达量 中图分类号：Q789 15 Application of real-time fluorescent quantitative polymerase chain reaction based on the Ct value comparison method to determine the relative genes expression WEI Jieshu, YANG Jinfen 20 25 30 (Research Center of Chinese Herbal Resource Science and Engineering, Guangzhou University of Chinese Medicine, Key Laboratory of Chinese Medicinal Resource from Lingnan (Guangzhou University of Chinese Medicine ), Ministry of Education, GuangZhou 510006) Abstract: Real-time fluorescent quantitative polymerase chain reaction is one of the nucleic acid quantification techniques, with the advantages of high specificity, high sensitivity, good repeatability, accurate quantization, fast speed and entire blocking. Ct value comparison method is the commonly used method of fluorescent quantitative PCR determination for relative genes expression. The target gene(s) and reference gene(s) can be amplified simultaneously by using Ct value comparison method to analyze relative genes expression, thus the workload of the experiment will be greatly reduced, and the experimental results are more reliable. It is an economical, practical, accurate and reasonable experimental method. In this paper, the key steps and technical details of the Ct value comparison method for the determination of the relative gene expression will be introduced through the description of concrete operations. Keywords: Fluorescent quantitative PCR; Ct value comparison method; relative quantitative; gene expression 35 0 引言 实时荧光定量 PCR (real-time fluorescent quantitative polymerase chain reaction，FQ-PCR ) 是 20 世纪 90 年代发展