牦牛MITF-M基因序列分析及其在皮肤组织中表达水平

Sequence Analysis and Expression Level Detection of MITF-M Gene in Different Color Skin Tissues of Yak

研究牦牛MITF-M基因编码区序列及其编码蛋白的结构,以及其在皮肤组织中mRNA的表达水平,探讨MITF-M基因与牦牛毛色形成相关性,以期为揭示牦牛毛色形成分子机理奠定基础。采用RT-PCR技术扩增,成功获得了牦牛MITF-M基因编码区序列。采用生物信息学方法,预测分析MITF-M基因及其编码蛋白的基本理化性质、二级结构等;采用半定量PCR检测技术,检测出MITF-M基因mRNA在牦牛各组织器官中表达水平;通过荧光定量PCR技术检测出在牦牛不同颜色皮肤组织中MITF-M基因mRNA表达水平。结果表明,扩增得到的MITF-M基因的编码区是一条长为1 460 bp的DNA序列。将牦牛的MITF-M基因序列命名为YAK MITF-M,并且在NCBI数据库中注册该基因的登录号为KM985448。牦牛MITF-M序列基因含有一个长度为1 242 bp的开放性阅读框,编码413个氨基酸,二级结构主要以α螺旋和无规则卷曲为主,β折叠和延伸链较少。MITF-M基因编码产物氨基酸邻接系统树表明,牦牛MITF-M与黄牛、绵羊等物种的MITF-M氨基酸具有高度相似性。在牦牛各组织器官中,MITF-M基因mRNA仅在皮肤组织中特异性表达,且在不同毛色牦牛皮肤组织中,MITF-M在黑色被毛皮肤组织中mRNA表达量显著高于白色被毛皮肤组织(P<0.01)。

The aim of this present study are the investigation of the MITF-M gene coding region sequences,the prediction of its translated protein structures and the dictation of its transcription mRNA expression level in skin tissues of yaks with black and white skin color. This study was aim to reveal of the molecular mechanism of skin color formation in yak. In this study,a coding region sequence of yak MITF-M was cloned by RT-PCR technique. Characters of the MITF-M gene encoded protein sequences were predicted and analyzed by the methods of bioinformatics in basically chemical properties and secondary structure. The expression levels of MITF-M mRNA in skin tissues of yak were detected by semi-quantitative PCR. Quantitative PCR was employed to examine the expression levels of MITF-M mRNA in different color yak skin tissues. As results,MITF-M gene was cloned,which contains a complete ORF of 1 242 bp. This ORF encoding a peptides chain which contain 413 amino acids. The second structure of this peptides chain was contain helix,coli and extended strand. And the expression level of MITF-M mRNA in skin tissues between dark yak and white yak were extremely remarkable（ P〈0. 01）.