qRT-PCR分析鳜鱼内参基因的筛选  

Screening of Reference Genes in Siniperca chuatsi for qRT-PCR Analysis

作　　者：李迪[1] 吴萍[2] 何美凤[1] 李伟[1] 肖调义[1] 褚武英[1] 

机构地区：[1]湖南农业大学动物科学技术学院,中国湖南长沙410128 [2]湖南大学生物学院,中国湖南长沙410082

出　　处：《生命科学研究》2016年第3期214-217,共4页Life Science Research

基　　金：湖南省教育厅重点实验室开放基金项目(15K011);国家自然科学基金项目(31472256)

摘　　要：选择合适的内参基因是提高实时荧光定量PCR分析(quantitative real-time polymerase chain reaction,qRT-PCR)准确性的首要条件。本实验采取鳜鱼8个不同胚胎发育阶段、5个胚后发育时期和8个成鱼组织为研究对象,应用qRT-PCR技术,分析了RPL13、RPL19、EF1a、RPL13a、B2M、hprt1和rps29七个内参基因的表达稳定情况。经GeNorm软件分析发现,B2M和RPL13a在鳜鱼成鱼不同组织中表达最稳定;在胚后不同时期中表达最稳定的是EF1a和RPL13a;EF1a和B2M是在不同胚胎发育阶段中表达最稳定的两个基因。根据内参基因标准化因子的配对差异分析V_(n/n+1),在鳜鱼不同组织和不同发育阶段中,均使用两个最稳定表达的内参基因即可达到准确校正的目的。因此,该实验结果为鳜鱼基因表达分析时内参基因的选择提供了参考。Selection of stable reference genes is a prerequisite for accurate application of real-time quantitative PCR （qRT-PCR） analysis in Siniperca chuatsi. Seven reference genes, including RPL13, RPLI19, EFIa, RPL13a, B2M, hprtl and rps29 were chosen to analyze expression stability by qRT-PCR in different adt, h tissues and developmental stages. The results showed that B2M and RPL13a were the most stable reference genes in different adult tissues. However, in different embryonic developmental stages, EF! a and B2M were the most stable reference genes, and in different post-embryonic developmental stages, EFIa and RPLI3a were the most stable genes. On the basis of the normalization factor Vn/n＋1, the optimum number of reference genes was two either in different adult tissues or in different developmental stages. The present study provided a useful method of choosing suitable reference genes in gene expression analysis in Siniperca chuatsi.

关 键 词：鳜鱼 内参基因 基因筛选 实时荧光定量 PCR Ge NORM 软件 

分 类 号：Q786[生物学—分子生物学]