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qRT-PCR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,β-actin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,β-actin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.
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,β-actin, and RPS29 were detected by using the qRT-PCR method, and Ge Norm and Norm Finder software packages. The optimal reference genes selected by analyzing and evaluating their mRNA ex-pression stability were applied to accurately quantify the relative expression level of the Clock gene in different tis-sues 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 wasβ-actin 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 the muscle tissue, followed by liver, kidney, and intestines, and the lowest was in heart. In summary, this study has accurately quantified the relative expression levels of the Clock gene in tissues, which should serve as the theoretical basis for the future investigation of the Clock gene rhythm system.
作者
彭治桃
蒋国民
邹利
李金龙
程小飞
房丽娟
王晓清
刘丽
PENG Zhi-tao1 2 JIANG Guo-min3 ZOU Li3 LI Jin-long3 CHENG Xiao-fei3 FANG Li-juan4 WANG Xiao-qing1＂ LIU Li~＇3 ~ 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
出处
《晓庄学院自然科学学报》
CAS
北大核心
2016年第6期37-42,共6页
Journal of Natural Science of Hunan Normal University
基金
国家自然科学基金资助项目(31372530)
