绵羊性腺褪黑素受体基因MT2的克隆及其生物信息学分析

Cloning and primary bioinformatics analysis of MT2 gene from ovine gonads

采用RT-PCR技术,以绵羊性腺(睾丸和卵巢)cDNA为模板扩增褪黑素受体MT2基因,经连接转化,蓝白斑筛选,挑取阳性克隆测序。结果显示,该试验成功克隆了MT2基因(282bp);通过Blast比对,克隆的MT2基因与已知序列(GenBank ID:NM_001130938)的同源性达到100%。生物信息学分析表明,克隆的MT2基因序列属于MT2第二外显子的一部分,富含GC(69.5%);整个MT2总共编码376个氨基酸的肽序列,其中丙氨酸(Ala)比例最高(13.03%);通过对绵羊MT2编码产物的二级结构、亲水性/疏水性、信号肽、亚细胞定位和跨膜结构域等进行预测和分析,发现绵羊MT2基因的编码蛋白是一种脂溶性较强的跨膜蛋白,与山羊MT2的分子进化距离最近,亲缘关系最为密切。MT2基因的成功克隆及其生物信息学分析为进一步研究它与绵羊繁殖季节性的相关性奠定了基础。

Using ovine gonads（testis and ovary） cDNA as a template and RT-PCR technique,MT2 gene sequence was amplified, cloned and sequenced. In result, the cloned sequence（282 bp） had 100% similarity to the ovine MT2 gene previously published in the GenBank（ID：NM_001130938） ,indicating that the gene was successfully cloned. Bioinformatics analysis showed that the cloned sequence belonged to the part of MT2 exon 2 with a high GC content（69. 5%）. MT2 gene codes for a polypeptide of 376 amino acids,in which alanine（Ala） has the highest percentage（13.03%）. The ovine MT2 protein was a hydrophobicity transmembrane protein,based on the results of secondary structure, hydrophobicity, signal peptide, subcellular localization and transmembrane structure prediction. In conclusion, ovine MT2 gene had close evolutionary relationship to goat MT2 based on the analysis of molecular evolution. Cloning of MT2 gene from ovine gonads and the function analysis of the deduced MT2 protein have laid foundations for further studies of MT2 gene＇s functions in ovine seasonal breeding regulation.