奶牛乳腺炎无乳链球菌菌毛岛屿PI-2a骨架蛋白BP基因的克隆及其序列分析  

Cloning and sequencing of a gene that codes for an auxiliary pilus backbone protein(PI-2a)from Streptococcus agalactiae causing bovine mastitis

作　　者：白文丽[1] 吴金花[1] 锡林高娃[1] 布日额[1] 王华[1] BAI Wen-li WU Jin-hua XI LiIN Gao-wa BU Ri-e WANG Hua(CollegeofLife Science, InnerMon- golia University for Nationalities, Inner Mongolia Autonomous Region Engineering Technology Research Center for Prevention and Control of Pathogenic Bacteria in Milk, Research Institute for Pathogens in Milk, 028043, Tongliao, Inner Mongolia, Chin)

机构地区：[1]内蒙古民族大学生命科学学院,内蒙古自治区乳源性致病菌防控工程技术研究中心,内蒙古民族大学乳源性致病菌研究所,内蒙古通辽028043

出　　处：《中国病原生物学杂志》2017年第6期505-508,共4页Journal of Pathogen Biology

基　　金：国家自然科学基金项目(No.31560689);内蒙古自治区乳源性致病菌防控工程技术研究中心开放课题(No.MDK2016036)

摘　　要：目的获得通辽地区奶牛乳腺炎致病菌无乳链球菌相关菌毛岛屿BP基因序列并进行比对分析。方法根据DNAMAN分析基因库中牛源无乳链球菌菌毛岛屿PI-2a的BP基因序列设计1对特异性引物,以无乳链球菌通辽分离株总DNA为模板,PCR扩增BP基因片段并进行序列比对分析。结果获得了菌毛岛屿PI-2a的BP基因序列,目的基因为609bp,编码203个氨基酸,其基因序列与GenBank上公布的牛源无乳链球菌菌毛岛屿PI-2a的BP基因(EU930008)同源性为100%。BP蛋白二级结构的α螺旋在N端分布较多,β折叠较丰富且分布C端较多,无规则卷曲分布较均匀。BP蛋白的亲水区分布较广,遍布于整个蛋白区,属于亲水性蛋白。BP蛋白分子表面可及性较高,区域占比达57.0%,且具有抗原性。结论获得了奶牛乳腺炎无乳链球菌通辽分离株菌毛岛屿PI-2a骨架蛋白BP基因序列,该基因序列具有高度保守性,其编码蛋白具有抗原表位,可作为奶牛无乳链球菌重组基因工程疫苗候选亚单位抗原。Objective To clone and sequence a gene that codes for a pilus backbone protein （PI-2a） of Streptococcus agalactiae causing bovine mastitis in the City of Tongliao in Inner Mongolia. Methods The bioinformatic software DNASTAR was used to analyze the sequence of the PI-2a gene of S. agalactiae published in GenBank, the software Primer 5.0 was used to design a pair of specific primers, and total DNA from clinical isolates of S. agalactiae was used as a template to amplify the PI-2a gene fragment. Results The sequence of the PI-2a gene was 609 bp in length and it en- coded 203 amino acid residues. The sequence had 100% similarity to the sequence of a PI-2a gene （EU930008） of S. aga- lactiae that codes for a pilus backbone protein published in GenBank. a-helices of the secondary structure of the protein were located more at the N-terminus, B-folds were located more at the C-terminus, and random curls were distributed throughout. The backbone protein had hydrophilic regions widely distributed throughout the protein, indicating that the protein is hydrophilic. The protein molecules had a high level of surface accessibility, accounting for as much as 57.0% of the surface, and the protein was antigenic. Surface-accessible areas of the BP protein were located at amino acids 2--5, 15 --20, 24--30, 33--38, 69--78, 108--121, 123--138, 158--164, 168-172, and 206 207. Conclusion The se- quence of the PI--2a gene was obtained from isolates of S. agalactiae causing bovine mastitis in the Tongliao area. The gene sequence was highly conserved and the protein had epitopes. The protein can be used as a potential subunit antigen with which to genetically engineer a recombinant vaccine against S. agalactiae.

关 键 词：无乳链球菌 菌毛岛屿 BP基因 克隆 序列分析 

分 类 号：R378.12[医药卫生—病原生物学]