鲮生长激素基因cDNA的分子克隆和序列分析

Molecular cloning and sequence analysis of growth hormone cDNA from Cirrhina molitorela

根据生长激素(growthhormone,GH)基因的保守性序列设计1对引物,利用RT PCR技术从鲮(Cirrhinamolitorella)脑垂体组织中克隆出鲮生长激素成熟肽cDNA片段,将纯化的DNA片段克隆到pUCm T载体上。克隆的鲮GHcDNA开放阅读框全长567bp,编码由188氨基酸残基组成的生长激素成熟肽。序列分析结果表明,鲮GH成熟肽氨基酸序列与另2种鲮—印鲮(Cirrhinamrigal)和南亚野鲮(Labeorohita)的成熟肽氨基酸序列同源性分别为87%和83%,与鲤的成熟肽氨基酸序列同源性为96%,并对9个科的17种鱼进行了分子系统进化树分析,结果与根据传统的形态学和生化特征分类进化地位基本一致。本研究所克隆的基因序列和推测的蛋白质序列已登录入GenBank、EMBL和DDBJ基因库,序列号分别为AF458105和AAL51107 1。

The total RNA was isolated from the pituitary of Cirrhina molitorella, and the cDNA encoding growth hormone (GH) peptide was amplified by reverse transcription polymerase chain reaction (RTPCR) strategy using total RNA as template. The amplified cDNA fragment was inserted to the vector pUCmT. The cloned cDNA was composed of 567 bases and encoded 188 amino acid residues. The sequence analysis indicates that the sequence homogeneity in mature GH peptide between C. irrhina molitorella and C. mrigal is 87%; between C. irrhina molitroella and Labeo rohita is 83%, and between C.irrhina molitorella and Cypeinus carpio is 96%. Also, the molecular phylogenesis tree of 17 species of fishes belonging to nine families was established to analyze their homogeneity and evolution, and the results are similar to those analyzed by traditional morphology and biochemistry in their evolution status. The gene sequence cloned in this experiment and the deduced protein sequence have been recorded in GenBank, EMBL and DDBJ, whose numbers are AF458105 and AAL 51107.1, respectively.