山羊痘病毒晚转录因子VLTF-1的原核表达及生物信息分析  

Prokaryotic expression and bioinformatics analysis of late transcription factor VLTF-1 of goat pox virus

作　　者：张成 王玉婷 杨勇飞 柳璇 吴锦艳[3] 尚佑军[3] 李有文[1,2] Zhang Cheng;Wang Yuting;Yang Yongfei;Liu Xuan;Wu Jinyan;Shang Youjun;Li Youwen(Key Laboratory of Tarim Animal Science and Technology Group,Xinjiang Production and Construction Group,College of Animal Science,Tarim University,Xinjiang Alar 843300;Engineering Laboratory for Diagnosis and Control of Animal Diseases in Southern Xinjiang of Xinjiang Production and Construction Corps,College of Animal Science,Tarim University,Xinjiang Alar 843300;State Key Laboratory of Veterinary Etiological Biology,Lanzhou Veterinary Research Institute,Chinese Academy of Agricultural Sciences,Gansu Lanzhou 730046)

机构地区：[1]塔里木大学动物科学学院新疆生产建设兵团塔里木畜牧科技兵团重点实验室,新疆阿拉尔843300 [2]塔里木大学动物科学学院新疆生产建设兵团南疆动物疫病诊断与防控工程实验室,新疆阿拉尔843300 [3]中国农业科学院兰州兽医研究所家畜疫病病原生物学国家重点实验室,甘肃兰州730046

出　　处：《现代畜牧兽医》2022年第3期20-25,共6页Modern Journal of Animal Husbandry and Veterinary Medicine

基　　金：国家重点研发计划(2018YFD0502006);国家自然科学基金资助项目(31360616、31760741)。

摘　　要：试验旨在研究山羊痘病毒晚转录因子VLTF-1的特性。研究以山羊痘病毒SS基因组株为模板,采用PCR方法扩增晚转录因子VLTF-1基因,采用基因克隆常规方法与pET-42b表达载体相连接构建原核表达载体,经菌液PCR、双酶切和测序鉴定,阳性克隆转化大肠杆菌BL21,以0.1 mmol/L IPTG诱导蛋白表达,SDS-PAGE电泳和Western Blot检测表达情况;同时对VLTF-1基因及其蛋白特性进行生物信息学分析。结果显示,PCR扩增的目的基因全长783 bp,与参考株Pellor同源性为100%,推测其编码261个氨基酸。遗传进化树分析表明,羊痘病毒属的3个种各分成一个分支,目的基因与山羊痘毒株位于同一分支。测序结果显示,目的基因表达载体构建正确,SDS-PAGE检测表达了27 kDa的蛋白,Western Blot鉴定表明目的蛋白得到表达。生物信息学分析结果显示,SS株的VLTF-1基因与山羊痘病毒聚于一支,该蛋白是一个跨膜亲水蛋白,具有26个磷酸化位点,二级结构是其结构连接5个α-螺旋。研究表明,试验成功克隆晚转录因子VLTF-1基因,并诱导表达蛋白,预测该蛋白的生物学信息。The purpose of the experiment was to study the properties of goat pox virus late transcription factor VLTF-1.In the study,the late transcription factor VLTF-1 gene was amplified by PCR using goat poxvirus SS gene strain as template,and the prokaryotic expression vector was constructed by linking the gene cloning with peT-42b expression vector by conventional gene cloning method.The positive clone was transformed into EScherichia coli BL21 by PCR,double enzyme digestion and sequencing.The protein expression was induced by 0.1 mmol/L IPTG,and the expression was detected by SDS-PAGE and Western Blot.At the same time,the characteristics of VLTF-1 gene and its protein were analyzed by bioinformatics.The results showed that the full length of the target gene amplified by PCR was 783 bp,and the homology with the reference strain Pellor was 100%,and it was speculated that it encoded 261 amino acids.The target gene was in the same branch as the goat pox strain.Sequencing results showed that the target gene expression vector was constructed correctly,SDS-PAGE detected the expressed protein of 27 kDa,and Western Blot identification showed that the target protein was expressed.The results of bioinformatics analysis showed that the VLTF-1 gene of the SS strain was clustered with goat pox virus.The protein was a transmembrane hydrophilic protein with 26 phosphorylation sites,and its secondary structure was connected with 5α-helix.The study indicates that the experiment successfully cloned the late transcription factor VLTF-1 gene,induced the expression of the protein,and predicted the biological information of the protein.

关 键 词：山羊痘病毒 晚转录因子VLTF-1基因 原核表达 生物信息学分析 

分 类 号：S855.3[农业科学—临床兽医学]