银黑狐TYRP1基因CDS区鉴定及生物信息学分析

Identification and bioinformatics analysis on CDS of TYRP1 gene in silver fox

为了探明酪氨酸酶相关蛋白1（TYRP1）基因对狐狸毛色的调控机制，本研究利用RT-PCR、染色体步移和核酸测序等技术，获得了银黑狐TYRP1基因完整编码区（coding DNA sequence，CDS），利用ProtParam和PredictProtein等在线软件对该基因进行了生物信息学分析，并利用Lasergene7．1软件包进行银黑狐与其他物种间的同源性分析及系统进化树的构建。结果表明，银黑狐TYRP1基因CDS区共计1614bp，编码537个氨基酸残基，属于不稳定可溶性亲水蛋白质。亚细胞定位主要在内质网和高尔基体中，属于分泌蛋白。在第24和25个氨基酸之间存在1个裂解位点，存在信号肽。其属跨膜蛋白，包括跨膜域、胞外域和胞内域3个部分。二级结构以无规则卷曲为主（59．03％），α-螺旋占22．91％，属卷曲蛋白质。三级结构与二级结构预测结果基本一致，主要是由无规则卷曲和α-螺旋构成。银黑狐与其他10个物种间的相似度为88．6％～99．6％，说明物种之间同源性较高，TYRP1基因编码区在进化过程中保守性较强。系统进化分析提示，银黑狐与赤狐和家犬的遗传亲缘关系最近。银黑狐TYRP1基因编码区的成功获取及分析为进一步探讨其在毛色形成过程中的功能研究提供了较好的理论依据。

To explore the regulation of fox TYRP1 gene in coat color,the complete CDS region se quence of silver fox TYRP1 gene was obtained by RT-PCR,genome walking and direct sequencing methods. The bioinformation of silver fox TYRP1 gene were analyzed by online software like Prot Param,PredictProtein, etc. Meanwhile, the homology and the phylogenetic tree were analyzed and constructed for silver fox and other 10 species by LasergeneT. 1 software package as well. The coding sequence of silver fox TYRP1 gene was 1 614 bp,encoding a total of 573 amino acids. It was an unstable,soluble and hydrophilic protein. Subcellular localization of TYRP1 was in the endoplasmic reticulum and Golgi,belong to the secreted protein. The silver fox TYRP1 had obvious signal peptide,and the cleavage site of the signal peptide was between position 24 and 25. The transmem- brane segments composed of extracellular domain, transmembrane domain intracellular domain. The secondary structure of TYRP1 was mainly random coil and s-helices,random coil was 59.03% and α-helices was 22. 91%, and TYRP1 was a coil class protein. The tertiary structure was also mainly composed of random coil and α-helix, which was consistent with its secondary structure. Compared with other species,the sequence percent identity of amino acids were 88.6% to 99.6%, which suggested that TYRP1 gene coding region was conservative in the course of long-term biological evolution. The phylogenetic analysis suggested that there were the closest genetic relationship among silver fox, red fox and dog. The successful acquisition and bioinformatics analysis of the coding region of silver fox TYRP1 gene laid a good theoretical basis for further revealing its function in coat color formation process.