《中国畜牧兽医》 ›› 2019, Vol. 46 ›› Issue (5): 1299-1307.doi: 10.16431/j.cnki.1671-7236.2019.05.006

• 生物技术 • 上一篇    下一篇

黑貉TYRP1基因的克隆及生物信息学分析

郭跃跃, 张宇飞, 王天骄, 邢秀梅, 徐超, 赵家平   

  1. 中国农业科学院特产研究所, 长春 130112
  • 收稿日期:2018-09-10 出版日期:2019-05-20 发布日期:2019-05-20
  • 通讯作者: 徐超, 赵家平 E-mail:xuchao@caas.cn;zhaojiaping@caas.cn
  • 作者简介:郭跃跃(1992-),女,河北邢台人,硕士,研究方向:遗传育种,E-mail:18333643847@163.com
  • 基金资助:

    中央级公益性科研院所基本科研业务费;吉林省科技发展计划(20180520032JH、20190301048NY);中国科协青年人才托举工程(2017QNRC001)

Cloning and Bioinformatics Analysis of TYRP1 Gene in Black Raccoon Dog

GUO Yueyue, ZHANG Yufei, WANG Tianjiao, XING Xiumei, XU Chao, ZHAO Jiaping   

  1. Institute of Special Animal and Plant Science of CAAS, Changchun 130112, China
  • Received:2018-09-10 Online:2019-05-20 Published:2019-05-20

摘要:

为探明黑貉酪氨酸酶相关蛋白1(tyrosinase-related protein 1,TYRP1)基因CDS序列并对其进行生物信息学分析,试验采集黑貉背部皮肤组织提取RNA后反转录为cDNA,根据GenBank中犬科TYRP1基因mRNA序列(登录号:NM_001194966.1)设计1对特异性引物,通过RT-PCR扩增TYRP1基因CDS区,将其插入到克隆载体中,对阳性菌落经PCR验证和序列测定后进行生物信息学分析。结果显示,黑貉TYRP1基因CDS序列全长为1 614 bp,编码537个氨基酸,所编码蛋白属于不稳定可溶性的亲水蛋白质。TYRP1基因与狐的遗传距离较近,说明该基因在不同物种及进化过程中具有高度保守性。TYRP1蛋白二级结构中,无规则卷曲所占比例最高,为54.0%;α-螺旋次之,为29.8%。TYRP1蛋白三级结构主要由大量无规则卷曲和α-螺旋构成,与二级结构相一致。TYRP1蛋白具有氧化还原酶活性,参与DNA修复和复制、蛋白质合成和降解,可与TYR、DCT、TP53、TRPC1等多种蛋白相互作用。本试验结果为进一步探索TYRP1基因在哺乳动物毛色遗传机制中的功能提供了理论依据。

关键词: 黑貉; TYRP1基因; 克隆; 生物信息学分析

Abstract:

To identify CDS sequence of tyrosinase-related protein 1 (TYRP1) gene in Black raccoon dog and perform bioinformatics analysis,the back skin tissue of Black raccoon dog was collected to extract RNA in this study,and RNA was reverse transcribed into cDNA.One pair of specific primers was designed according to the canine TYRP1 gene mRNA sequence (accession No.:NM_001194966.1) in GenBank.The TYRP1 gene CDS was amplified by RT-PCR,PCR products were inserted into the cloning vector,and verified by positive colony PCR and sequenced.The results showed that the CDS sequence of TYRP1 gene in Black raccoon dog was 1 614 bp in length and encoded 537 amino acids,the TYRP1 protein was an unstable soluble hydrophilic protein.The genetic distance was closer between Black raccoon dog and fox,indicating that TYRP1 gene was highly conserved in different species and evolution processes.The proportion of random coil in the secondary structures of TYRP1 protein was the highest (54.0%),followed by alpha helix,which was 29.8%.The tertiary structure of TYRP1 protein was mainly composed of a large number of random coil and alpha helix,which was consistent with the secondary structure.TYRP1 protein had oxidoreductase activity,which was involved in DNA repair and replication,protein synthesis and degradation,and interacted with various proteins such as TYR,DCT,TP53 and TRPC1.The results of this experiment provided a theoretical basis for further exploration of its function in mammalian hair color genetic mechanism of TYRP1 gene in Black raccoon dog.

Key words: Black raccoon dog, TYRP1 gene, cloning, bioinformatics analysis

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