藏绵羊FAM134B基因克隆与表达分析

doi:10.3969/j.issn.1004-1524.2017.09.08

浙江农业学报 ›› 2017, Vol. 29 ›› Issue (9): 1474-1481.DOI: 10.3969/j.issn.1004-1524.2017.09.08

藏绵羊FAM134B基因克隆与表达分析

李文丽^{1, 2}, 杜晓华^{2, 3}, 刘霞^{1, 2, *}, 高景波^{1, 2}, 申超超^{1, 2}, 张超^{1, 2}, 刘伟^{1, 2}

1.甘肃农业大学生命科学技术学院,甘肃兰州 730070;
2.甘肃省草食动物生物技术重点实验室,甘肃兰州 730070;
3.甘肃农业大学动物医学院,甘肃兰州 730070

收稿日期:2016-12-26 出版日期:2017-09-20 发布日期:2017-09-27
通讯作者: 刘霞,E-mail:liux@gsau.edu.cn
作者简介:李文丽(1991—),女,甘肃会宁人,硕士研究生,从事生物化学与分子生物学研究工作。E-mail:1406523498@qq.com
基金资助:
农业部畜禽遗传资源与种质创新重点实验室开放课题资助(nzdsys2015-1)

Molecular cloning and expression analysis of FAM134B gene in Tibetan sheep

LI Wenli^{1, 2}, DU Xiaohua^{2, 3}, LIU Xia^{1, 2, *}, GAO Jingbo^{1, 2}, SHEN Chaochao^{1, 2}, ZHANG Chao^{1, 2}, LIU Wei^{1, 2}

1.College of Life Science and Technology, Gansu Agricultural University, Lanzhou 730070, China;
2. Gansu Key Laboratory of Herbivorous Animal Biotechnology, Lanzhou 730070, China;
3. College of Veterinary Medicine, Gansu Agricultural University, Lanzhou 730070, China

Received:2016-12-26 Online:2017-09-20 Published:2017-09-27

摘要/Abstract

摘要： 采用RT-PCR技术克隆了藏绵羊FAM134B(family with sequence similarity 134, member B)基因,利用生物学软件进行相关生物信息学分析,同时用实时荧光定量PCR(real-time fluorescence quantitative PCR, qRT-PCR)研究组织表达水平。克隆获得FAM134B基因全长1 144 bp (GenBank登录号: KX580309),开放阅读框1 071 bp,编码356个氨基酸。系统进化树表明,藏绵羊FAM134B和绵羊、山羊关系较近。藏绵羊FAM134B基因编码蛋白的分子式为C₁₇₁₇H₂₆₉₆N₄₄₈O₅₇₂S₁₂,分子量为39 151.72 u,理论等电点(pI)为4.49,消光系数为24 450,不稳定系数为43,疏水指数为80.31,平均亲水性为-0.422,属不稳定可溶性酸性蛋白质。二级结构主要是α-螺旋(45.51%)、无规卷曲(41.85%)和延伸链(12.64%),属于混合类蛋白质。亚细胞定位和功能分析结果显示,FAM134B编码的蛋白在内质网、细胞质膜概率分别为30.4%和21.7%,主要在蛋白质运输和结合(77.3%)方面起作用。系统发育树表明,藏绵羊FAM134B氨基酸序列与绵羊、山羊同源性非常高。qRT-PCR结果表明,FAM134B基因网胃中表达最高,十二指肠表达量最低。研究结果可为进一步研究藏绵羊FAM134B基因结构和生理功能及其遗传特性提供理论依据。

关键词: 藏绵羊, FAM134B基因, CDS区克隆, 生物信息学分析, 表达

Abstract: The gene of FAM134B(family with sequence similarity 134, member B) was cloned by RT-PCR. The bioinformatics analysis was carried out by using biological software. The expression level of FAM134B was also studied by real-time fluorescence quantitative PCR. The full length of FAM134B gene was 1 144 bp (GenBank accession number: KX580309), open reading frame 1 071 bp, encoding 356 amino acids. The phylogenetic tree showed that the Tibetan sheep FAM134B had a close relationship with the sheep and goat. The molecular formula of FAM134B protein in Tibetan sheep was C₁₇₁₇H₂₆₉₆N₄₄₈O₅₇₂S₁₂, its molecular weight was 39151.72 u, the theory isoelectric point was 4.49, the extinction coefficient was 24 450. The instability index was 43, the aliphatic index was 80.31, and the grand average hydropathicity was -0.422. It was unstable and soluble acidic protein. The secondary structure of FAM134B was mainly composed of α-helices and random coil and extended chain, with α-helices was 45.51%, random coil was 41.85% and extension chains was 12.64%, which belongs to mixed type of protein. The results of subcellular localization and function showed that the frequencies of FAM134B protein in endoplasmic reticulum and cytoplasmic membrane were 30.4% and 21.7%. It mainly plays a role in protein transport and binding(77.3%). Phylogenetic tree showed that the amino acid sequence of FAM134B showed high homology with sheep and goat. qRT-PCR results showed that the expression of FAM134B gene was the highest in the stomach and lowest in the duodenum. The results can provide a theoretical basis for further study on the structure and physiological functions and genetic characteristics of FAM134B gene.

Key words: Tibetan sheep, FAM134B gene, CDS cloning, bioinformatics, expression

中图分类号:

S826

李文丽, 杜晓华, 刘霞, 高景波, 申超超, 张超, 刘伟. 藏绵羊FAM134B基因克隆与表达分析[J]. 浙江农业学报, 2017, 29(9): 1474-1481.

LI Wenli, DU Xiaohua, LIU Xia, GAO Jingbo, SHEN Chaochao, ZHANG Chao, LIU Wei. Molecular cloning and expression analysis of FAM134B gene in Tibetan sheep[J]. , 2017, 29(9): 1474-1481.

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藏绵羊FAM134B基因克隆与表达分析

Molecular cloning and expression analysis of FAM134B gene in Tibetan sheep

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