草鱼原肌球蛋白基因的克隆及其组织表达  

Molecular cloning and expression analysis of tropomyosin gene from Grass carp

作　　者：孙念[1] 陈开建[1] 褚武英[2] 肖调义[1] 李迪[1] 

机构地区：[1]湖南农业大学动物科学学院,湖南长沙410003 [2]长沙学院生物工程与环境科学系,湖南长沙410003

出　　处：《湖南文理学院学报（自然科学版）》2015年第2期51-56,共6页Journal of Hunan University of Arts and Science(Science and Technology)

摘　　要：为揭示原肌球蛋白基因在草鱼肌肉中的作用,利用RT-PCR和RACE技术克隆获得了草鱼原肌球蛋白基因c DNA,并对该基因在普通草鱼和脆肉鲩不同组织中的表达情况进行研究分析。结果表明原肌球蛋白基因c DNA全长序列为1 705 bp,包含387 bp的5′UTR序列,1 307 bp的3′UTR序列和855 bp开放阅读框(ORF)。其ORF编码284个氨基酸。系统进化分析表明普通草鱼与斑马鱼、墨西哥脂鲤的原肌球蛋白基因核苷酸同源性分别是93%和87%,氨基酸同源性分别是96%和93%。在聚类上普通草鱼原肌球蛋白基因与其他鲤科鱼类同源性较高,表明亲缘关系最近,与传统分类相一致。Real time-PCR结果表明原肌球蛋白基因在所检测的普通草鱼和脆肉鲩7个组织中均有表达,原肌球蛋白基因在普通草鱼腹肌中表达最高,其次为前肠。原肌球蛋白基因在脆肉鲩腹肌中的表达低于普通草鱼,而脆肉鲩中肌肉、肝脏、肾脏、前肠、后肠中原肌球蛋白基因表达量大于普通草鱼相对应组织,但差异不显著。To reveal the original role of TPM gene in grass carp muscle, using rapid amplification of eDNA ends （RACE）, the full length of TPM eDNA sequence in grass carp is cloned, and the expression of different tissues in common grass carp and embrittlement of grass carp are analyzed by Real-time PCR. The sequence analysis shows that the TPM eDNA had 1 705 bp in length with a 3＇-untranslated region （1 307 bp）, a 5＇-untranslated region （387 bp）, and a 855 bp open reading frame （ORF）. Bioinformatics analysis predicts that the ORF of TPM4a eDNA encoded a protein of 284 amino acid residuals with a structure of protein tertiary model with 277 alpha helix and 7 random curl. The phylogenetic analysis shows that the amino acid sequence of common grass carp TPM possessed 96% and 93% identity with the TPMs of Daniorerio and Astyanax mexicanus, respectively. Real-time PCR results demonstrated that the abdominal muscle of common grass carp had the highest expression of TPM, and next the foregut. Comparing with embrittlement of grass carp, TPM shows lower expression inabdominal muscle of common grass carp, while the higher expression of embrittlement of grass carp tissues were muscle, liver, spleen, foregut and hindgut. The results indicate that TPM can be a new candidate target gene for investigating embrittlement mechanism of grass carp embrittlement and improving healthy level of culture.

关 键 词：普通草鱼 原肌球蛋白 基因表达 脆肉鲩 

分 类 号：TS254.1[轻工技术与工程—水产品加工及贮藏工程]