唐鱼生长激素基因的克隆及序列分析

Cloning and Sequence Analysis of Growth Hormone Gene from Tanichthys albonubes

[目的]为开展唐鱼转自源基因研究提供基本构架。[方法]采用RT-PCR和RACE技术分离唐鱼生长激素基因(GH)全长cD-NA序列,同时利用PCR克隆了唐鱼生长激素基因的基因组(gDNA)序列。[结果]序列分析表明:唐鱼GH cDNA的5非编码区为64bp,3非编码区为448 bp,开放阅读框(ORF)633 bp,共编码210个氨基酸,包括22个氨基酸的信号肽和188个氨基酸的成熟肽。应用MEGA 3.1软件进行序列同源性比较分析的结果显示,唐鱼GH cDNA所编码的氨基酸序列与近缘鱼种(草鱼、青鱼、鳙鱼、鲤鱼、斑马鱼等)的生长激素氨基酸序列有很高的同源性,其中与草鱼和鳙鱼的同源性高达98%。该研究获得的唐鱼生长激素基因的gDNA序列共有1403 bp,包括4个外显子和3个内含子,外显子大小分别150、117、162、204 bp;3个内含子都以GT开头、AG结尾,符合GT-AG法则。[结论]该研究为下一步构建自源GH基因构件,进行唐鱼的转自源GH基因研究,培育出生长快、体型大的唐鱼新品系奠定了基础。

[ Objective] The research aimed to provide the basic structure for the study of self transgene of Tanichthys albonubes. [ Method ] The full length cDNA encoding T. albonubes growth hormone （ TcGH ） was cloned with RT-PCR and RACE （ rapid amplification of cDNA ends）. [ Result] The TeGH cDNA consisted of an open reading frame of 633 bp, 5＇ and 3＇ untranslated regions of 64 and 448 bp respectively. The deduced amino-acid sequence of the TcGH had 210aa, including a putative signal peptide （22aa） located at its N-terminal. The homology of the TcGH amino-acid with relative species （grass carp, black carp, bighead carp, carp, zebra fish, etc. ） was high, and was as much as 98% with grass carp and bighead carp. The genomic DNA （gDNA） of T. albonubes growth hormone was obtained by PCR. The gDNA sequence consisted of four exons （150, 117, 162 and 204 bp in length, respectively） and three introns. This three introns started with GT and ended with AG, in accordance with GT-AG principle. According to amino acid of growth hormone, the molecular phylogenesis tree of 25 species of fishes belonging to nine families was established to analyze their homogeneity and evolution, and the results were similar to those ana- lyzed by traditional morphology and biochemistry in their evolution status. [ Conclusion ] This research provided the foundation for construction of GH gene elements ,the study of self transgene of T. albonubes and the new lines breeding of rapid growth , big T. albonubes.