布氏杆菌S2弱毒疫苗株的S2JY3基因生物信息学分析及原核表达  

Bioinformatic analysis and prokaryotic expression of the S2JY3 gene from the attenuated Brucella S2 vaccine

作　　者：苏倩 白丽艳[1] 孙柯 林洋 呼和巴特尔[1] 王文龙[1] SU Qian;BAI Liyan;SUN Ke;LIN Yang;HUHE Bateer;WANG Wenlong(College of Veterinary Medicine/Key Laboratory of Clinical Diagnosis and Treatment Technology in Animal Disease,Ministry of Agriculture and Rural Affairs,Inner Mongolia Agricultural University,Hohhot 010018,China)

机构地区：[1]内蒙古农业大学兽医学院/农业农村部动物疾病临床诊疗技术重点实验室,内蒙古呼和浩特010018

出　　处：《畜牧与兽医》2020年第6期76-80,共5页Animal Husbandry & Veterinary Medicine

基　　金：内蒙古自治区科技计划项目(201502071,20140174,20120244)。

摘　　要：为分析布氏杆菌S2弱毒疫苗特有的S2JY3基因的功能和作为诊断抗原的可能性,应用PCR技术扩增S2JY3基因,用限制性内切酶对目的片段与pET30a(+)原核表达载体进行双酶切,构建重组表达载体,用IPTG诱导表达,蛋白纯化,并进行Western blot检测;利用生物信息学软件对该基因理化性质、结构特点进行分析。结果表明,S2JY3基因全长1257 bp,编码418个氨基酸,蛋白质理论大小值为43 kDa,理论等电点为9.73,蛋白质不稳定系数为36.31,是稳定蛋白,总平均亲水性为-0.577,为亲水性蛋白;跨膜结构和信号肽分析表明S2JY3蛋白无跨膜区,无信号肽区域。本研究成功构建了重组表达蛋白rS2JY3,Western blot结果表明rS2JY3与布氏杆菌S2疫苗免疫羊血清发生特异性结合反应,不与自然感染羊血清反应,具有良好的抗原性和特异性。该研究为羊布氏杆菌S2疫苗免疫和自然感染快速检测和鉴定等提供理论依据。In order to determine the function of the S2 JY3 gene and the possibility of its being used as a diagnostic antigen,the gene was amplified by PCR,and the target fragment and the pET30 a(+)prokaryotic expression vector were subjected to double enzyme digestion with restriction enzyme.The recombinant expression vector was constructed,induced by IPTG,purified by protein and detected by Western blot.The physicochemical properties and structural characteristics of the gene were analyzed by a bioinformatics software.The results showed that S2 JY3 was 1257 bp in length and that it encoded 418 amino acids.The theoretical size of the protein was 43 kDa,the theoretical isoelectric point was 9.73,the instability coefficient of the protein was 36.31 indicating it to be a stable protein,and the total average hydrophilicity was-0.577 indicating it to be a hydrophilic protein.Analysis of its transmembrane structure and analysis of the signal peptide indicated that the S2 JY3 protein had no transmembrane region and no signal peptide region.The recombinant expression protein rS2 JY3 was successfully constructed in this study.The results of Western blot showed that rS2 JY3 specifically reacted with the immune serum of the Brucella S2 vaccine but did not react with the naturally infected sheep serum,and that the protein had good antigenicity and specificity.This study provided a theoretical basis for the rapid detection and identification of Brucella S2 vaccine infection and natural infection.

关 键 词：布氏杆菌 S2JY3基因 原核表达 生物信息学分析 

分 类 号：S855.1[农业科学—临床兽医学]