《中国畜牧兽医》 ›› 2017, Vol. 44 ›› Issue (10): 2829-2836.doi: 10.16431/j.cnki.1671-7236.2017.10.001

• 生物技术 •    下一篇

水牛FADS2基因的电子克隆及序列分析

马小娅, 庞春英, 梁莎莎, 陆杏蓉, 朱鹏, 段安琴, 梁贤威, 邓廷贤   

  1. 中国农业科学院广西水牛研究所, 农业部(广西)水牛遗传繁育重点实验室, 南宁 530001
  • 收稿日期:2017-04-05 出版日期:2017-10-20 发布日期:2017-10-20
  • 通讯作者: 邓廷贤 E-mail:dtx282000@163.com
  • 作者简介:马小娅(1989-),女,甘肃白银人,硕士,研究实习员,研究方向:水牛分子遗传学,E-mail:761195467@qq.com
  • 基金资助:

    广西科技重大专项(桂科AA16450002);广西科技公关国合项目(桂科合1504001-3);广西自然科学基金青年项目(2015GXNSFBA139103);广西水产畜牧科技推广应用项目(桂渔牧科201633009)

Cloning and Bioinformatics Analysis of FADS2 Gene in Buffalo

MA Xiao-ya, PANG Chun-ying, LIANG Sha-sha, LU Xing-rong, ZHU Peng, DUAN An-qin, LIANG Xian-wei, DENG Ting-xian   

  1. Key Laboratory of Buffalo Genetics, Breeding Reproduction Technology, Ministry of Agriculture(Guangxi), Guangxi Buffalo Research Institute, Chinese Academy of Agricultural Sciences, Nanning 530001, China
  • Received:2017-04-05 Online:2017-10-20 Published:2017-10-20

摘要:

试验旨在利用电子克隆法对水牛Δ6脂肪酸脱氢酶(Δ6-fatty acid desaturases,FADS2)基因进行克隆和生物信息学分析,为探究FADS2基因对水牛泌乳性能的作用机制奠定基础。以奶牛FADS2基因序列(GenBank登录号:NM_001083444.1)为探针设计引物,利用电子克隆法克隆水牛FADS2基因,并通过RT-PCR验证,对FADS2基因的序列特征进行生物信息学分析。测序结果表明,水牛FADS2基因序列全长为36 600 bp,由12个外显子和11个内含子组成,包含一个长1 335 bp的开放阅读框,可编码444个氨基酸。序列同源性分析显示,水牛FADS2基因编码序列与牦牛、黄牛、人、猪、家兔、虎鲸和褐家鼠序列的同源性分别为98.88%、98.88%、89.66%、90.79%、90.85%、92.35%和87.11%。蛋白质预测分析表明,水牛FADS2蛋白分子质量为52.51 ku,理论等电点(pI)为8.75,呈弱碱性,属于亲水性蛋白,无信号肽。系统进化树分析结果表明,FADS2基因在不同物种及进化的过程中具有高度保守性,其中水牛与牦牛、黄牛亲缘关系较近,与褐家鼠亲缘关系较远。水牛FADS2基因的成功克隆为今后阐明水牛泌乳性能的作用机制奠定了基础。

关键词: 水牛; FADS2基因; 电子克隆; 生物信息学分析

Abstract:

This study was aimed to clone the buffalo Δ6-fatty acid desaturases (FADS2) gene using in-Silico cloning and analyze its genetic struction with bioinformatics, which provide a foundation for investigating the milk performance in buffaloes. Primers were designed according the sequence of FADS2 gene in dairy cow (GenBank accession No.:NM_001083444.1). The FADS2 gene was amplified by RT-PCR, and its sequence was analyzed by bioinformatics. Sequence analysis revealed that the buffalo FADS2 gene had 36 600 bp in length and consisted of 12 exons and 11 introns, containing an open reading frame (ORF) of 1 335 bp which encoding 444 amino acids. Sequence homology analysis indicated that the buffalo FADS2 protein gene showed 98.88%, 98.88%, 89.66%, 90.79%, 90.85%, 92.35% and 87.11% identity with that of Bos mutus, Bos taurus, Homo sapiens, Sus scrofa, Oryctolagus cuniculus, Orcinus orca and Rattus norvegicus, respectively. Protein prediction analysis showed that the molecular weight and isoelectric point (pI) of buffalo FADS2 were 52.51 ku and 8.75, respectively, and the FADS2 protein was weak alkali and the hydrophobicity protein without signal peptide. Phylogenetic tree analysis showed that FADS2 gene was highly conserved in different species and evolutionary processes,buffalo was close to Bos mutus and Bos taurus, and was far from Rattus norvegicus. This study suggested that FADS2 gene was successfully cloned in buffalo, which laid a foundation for clarifying the mechanism of milk performance in buffalo.

Key words: buffalo; FADS2 gene; in-Silico cloning; bioinformatics analysis

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