巴马猪BECN2基因的克隆及其mRNA组织表达分析  

Molecular cloning and tissue expression analysis of BECN2 gene in Bama pig

作　　者：王玥[1] 郭晓晨[1] 姜超前 牟彦双[1] 刘忠华[1] WANG Yue;GUO Xiaochen;JIANG Chaoqian;MU Yanshuang;LIU Zhonghua(Laboratory of Animal and Cell Genetic Engineering of Heilongjiang Province,Northeast Agricultural University,Harbin 150030,China)

机构地区：[1]东北农业大学黑龙江省动物细胞与遗传工程实验室,哈尔滨150030

出　　处：《黑龙江畜牧兽医》2022年第1期60-65,70,134,135,共9页Heilongjiang Animal Science And veterinary Medicine

基　　金：农业农村部转基因新品种培育重大专项(2016ZX08009-003-006);黑龙江省自然科学基金联合引导项目(LH2020C016)。

摘　　要：为了探究巴马猪BECN2基因编码蛋白的生物学特性及其mRNA在巴马猪不同组织中表达水平,试验采用PCR方法扩增并克隆巴马猪BECN2基因CDS全长序列;通过生物信息学方法对其编码蛋白进行分析并构建系统进化树;应用实时荧光定量PCR方法检测巴马猪BECN2基因mRNA在不同组织中表达情况。结果表明:经PCR扩增巴马猪BECN2基因CDS全长序列大小为1302 bp;巴马猪BECN2蛋白是一种结构较不稳定的亲水性蛋白,由433个氨基酸组成,亮氨酸含量最高,占比为13.2%。巴马猪BECN2蛋白二级结构主要由α-螺旋、无规则卷曲、β-折叠和β-转角构成,占比分别为50.04%、34.41%、10.16%、1.39%,其氨基酸主要存在于细胞膜外;亚细胞主要定位在细胞核和线粒体中,占比为56.5%和21.7%;存在16个磷酸化位点,分别为10个丝氨酸、5个苏氨酸、1个酪氨酸位点,在第1~20个编码氨基酸位点可能存在信号肽结构。巴马猪BECN2基因核苷酸序列与江豚、海豚的同源性较高,为84.3%。巴马猪BECN2蛋白氨基酸序列与牛和羊的遗传距离较近。BECN2基因mRNA在巴马猪不同组织中均有表达,且在卵巢、肺脏、心脏、睾丸、空肠、大脑、小脑、胃、皮肤、骨骼肌组织中表达量较高。说明巴马猪BECN2基因在细胞中发挥关键作用。In order to explore the biological characteristics of BECN2 gene encoding protein and its mRNA expression level in different tissues of Bama pig,the full-length CDS sequence of BECN2 gene in Bama pig was amplified and cloned by PCR.Bioinformatics method was used to analyze the protein and construct phylogenetic tree.Real time PCR was used to detect the mRNA level of BECN2 gene in different tissues of Bama pigs.The results showed that the full-length sequence of CDS of BECN2 gene in Bama pig was 1302 bp.BECN2 protein in Bama pig was a hydrophilic protein with unstable structure,which was composed of 433 amino acids,among which leucine was the highest,accounting for 13.2%.The nucleotide sequence of BECN2 protein in Bama pig was mainly composed ofα-helix,random curl,β-folding andβ-turning,accounting for 50.04%,34.41%,10.16%and 1.39%,respectively,and mainly existed outside the cell membrane.Subcells were mainly located in the nucleus and mitochondria,accounting for 56.5%and 21.7%,respectively.There were 16 phosphorylation sites,including ten serine sites,five threonine sites and one tyrosine site,and signal peptide structure might exist at the 1 st-20 th coding amino acid sites.The nucleotide sequence of BECN2 gene in Bama pig had high homology with finless porpoise and dolphin,both reaching 84.3%.The amino acid sequence of BECN2 protein in Bama pig was close to that of cattle and sheep.BECN2 gene was expressed at mRNA level in different tissues of Bama pigs,and the expression levels were higher in ovary,lung,heart,testis,jejunum,brain,cerebellum,stomach,skin and skeletal muscle tissues.These results indicated that BECN2 gene played a key role in Bama pigs.

关 键 词：BECN2基因 巴马猪 克隆 组织表达 基因编码区 进化树 

分 类 号：S828[农业科学—畜牧学] Q785[农业科学—畜牧兽医]