Cloning and sequence analysis of growth hormone gene from donkey | 驴生长激素基因的克隆与分析

Chinese. 【目的】克隆驴生长激素基因DNA和cDNA的全序列,分析其序列和cDNA编码的蛋白序列特征及其在不同物种中的遗传差异。【方法】根据不同物种同一基因序列的同源性比对结果设计引物,应用RT-PCR和PCR 技术克隆基因,用生物信息学方法对获得的DNA和cDNA及推定的氨基酸序列进行分析。【结果】从驴肝组织和血液中分别得到驴GH基因全序列1 928 bp和包括完整的编码区的cDNA序列706 bp,两者序列比对后证明驴GH基因DNA序列由5个外显子和4个内含子组成,编码216个氨基酸的GH前体蛋白,其中包括26个氨基酸的信号肽和190个氨基酸的成熟肽。序列比较结果表明,驴GH基因的序列与马同源性最高,启动子不是哺乳动物的典型TATA盒,而是CATA盒,该基因在进化过程中是保守的。【结论】从驴肝组织和血液中克隆了GH基因DNA和cDNA,DNA序列在1 267位的C→G可能影响到驴和马生长发育的差异,为下一步驴GH基因的表达调控、进化和多态性分析奠定了基础。

English. 【Objective】 The objective of this study was to analyze the genetic differences of donkey growth hormone（GH）gene from its DNA, cDNA and deduced amino acid sequence in different species.【Method】The PCR primers were designed online according to result of gene homology comparison. Donkey GH gene DNA and cDNA sequence were cloned from liver tissue and blood by RT-PCR and PCR, and compared with GH gene sequences of different species by bioinformatics method. 【Result】 The donkey GH gene sequence was 1 928 bp, including the 706 bp cDNA sequence with the complete CDS. By comparison of DNA and cDNA sequences, it was found that the GH gene sequence included 5 exons and 4 introns, encoding 216 aa, including the signal protein of 26 aa and matured protein of 190 aa. Based on the analysis of the similarity of GH genes in different species on the level of cDNA, DNA and the deduced amino acid, there was the most homology to the horse. The GH gene of donkey was conservative in the process of evolution and its promotor was not specific TATA box, but CATA box. 【Conclusion】 DNA and cDNA sequences were obtained from liver tissue and blood, and the mutation of C→G DNA sequence in 1 267 may affect the growth and development of donkeys and horses. All researches made an essential foundation for GH regulation of gene expression, evolution, polymorphism analysis in the future.