锦鲫Clock基因表达量分析中的内参基因稳定性比较.docVIP

锦鲫Clock基因表达量分析中的内参基因稳定性比较 　　摘 要 为研究锦鲫Clock基因表达量分析中的内参基因稳定性，采用qRTPCR技术进行实时荧光定量分析，并用Ge Norm 和 Norm Finder软件对5个内参基因（RPL13，GAPDH，18 S rRNA，βactin和RPS29）进行稳定性评估，筛选出不同组织中最适合的内参基因，以准确定量Clock 基因的相对表达水平.实验结果表明，5个内参基因中，18 S rRNA在锦鲫的肌肉、心脏、肝脏中表达最稳定，βactin在肠中表达最稳定，RPL13在肾中表达最稳定；根据筛选的内参基因定量Clock基因的相对表达水平发现，Clock基因在锦鲫肌肉中相对表达量最高，其次依次是肝脏、肾、肠，心脏中最低.本研究准确定量 　　Clock基因， 为锦鲫生物钟节律机制的下一步研究奠定了理论基础. 　　关键词 内参基因；qRTPCR；锦鲫；Ge Norm程序；Norm Finder程序 　　中图分类号 S9174文献标识码 A文章编号2016 　　Stability Comparison of Reference Genes on Expression 　　Analysis of Clock Gene in Carassius Auratus 　　PENG Zhitao1，2， JIANG Guomin3， ZOU Li3， LI Jinlong3， CHENG Xiaofei3， FANG Lijuan4， WANG Xiaoqing1*， LIU Li1，3* 　　（1. College of Animal Science and Technology， Hunan Agriculture University， Changsha 410128， China； 　　2. Aquatic Products Seed Stock Station in Hunan Province， Changsha 410153， China； 　　3. Fisheries Research Institute of Hunan Province， Changsha 410153， China； 　　4. College of Basic Medicine， Central South University， Changsha 410012， China） 　　Abstract The aim of this study is to compare the stability difference of reference genes from the expression analysis of Clock gene in Carassius auratus. The mRNA expression of five candidate reference genes such as RPL13， GAPDH 18 S rRNA， βactin， and RPS29 were detected by using the qRTPCR method， and Ge Norm and Norm Finder software packages. The optimal reference genes selected by analyzing and evaluating their mRNA expression stability were applied to accurately quantify the relative expression level of the Clock gene in different tissues of Carassius auratus. Our results show that one of the most stable reference genes was 18 S rRNA from three tissues， including muscle， heart， and liver， among the five reference genes， whereas that found in intestines was βactin and in kidney was RPL13. When these optimal reference genes were selected in five tissues， we found that the relative expression level of the Clock gene was the highest in th