新疆农业科学2017，Vol.54Issue(3)：564-573,10.DOI:10.6048/j.issn.1001-4330.2017.03.022
新疆羊种布鲁氏菌分离株的鉴定与L7/L12蛋白的原核表达和生物信息学分析
Identification of Isolate Strain of Brucella Melitensis in Xingjiang and Prokaryotic Expression and Bioinformatics Analysis of Its L7/L12 Protein
摘要
Abstract
[Objective] To isolate and identify Xinjiang Brucella melitensis,this project focuses on prokaryotic expression of the L7/L12 protein of the bacteria,detection of its reactionogenicity and and partial biological analysis.[Method]Isolation and identification of Brucella melitensis by using bacterial streak culture,morphological observation,biochemical test and PCR detection.Using conventional and molecular biological methods to express and purify the L7/L12 protein of this Brucella melitensis.Expression and purification isolate strain L7/L12 protein by using the conventional molecular biological methods,and analysis of the fusion protein reactionogenicity by Western Blot.Using bioinformatics software to analyze some functions of this protein.[Result]After identification,the strain was identified as Brucella melitensis.After enzyme digestion and sequencing,the expression vector pET-28a-L7/L12 was correctly constructed.SDS-PAGE tests showed that the purified L7/L12 protein was a single band.The fusion protein had good reactionogenicity by Western Blot detection.Bioinformatics analysis showed the protein had no trans-membrane domain and no signal peptide.Its secondary structure was mainlyα-helix.And the three-dimensional structure of the protein was constructed by Phyre2 Server.[Conclusion]The isolated strain was identified successfully and L7/L12 fusion protein of this isolate strain was expressed and purified.Blot Western test proved that the protein had a good reactionogenicity,which laid the foundation for the protein follow-up research of the subunit vaccine.关键词
羊种布鲁氏菌/L7/L12/蛋白纯化/反应原性/生物学Key words
Brucella melitensis/L7/L12/protein purification/reactionogenicity/biological analysis分类
农业科技引用本文复制引用
刘升,江雅丽,付强,史慧君,李爽,孟露萍,郭飞,张辉,陈创夫..新疆羊种布鲁氏菌分离株的鉴定与L7/L12蛋白的原核表达和生物信息学分析[J].新疆农业科学,2017,54(3):564-573,10.基金项目
国家自然科学基金面上项目"TceSR和TcfSR在布鲁氏菌致病过程中的分子调控机制研究"(31572491)General Programs of the National Natural Science Foundation of China"Study on the molecular regulation of TceSR and TcfSR in the pathogenesis of Brucella" (31572491). （31572491）
