酵母双杂交筛选与胞内劳森菌LI0902互作宿主蛋白  

Screening of proteins interacted with Lawsonia intercellularis LI0902 protein by the yeast two-hybrid system

作　　者：张兰桥 赖崇德[1] 戴益民[2] 张庆华[1] 钟其旺[1] 赖芬菊[1] ZHANG Lan-qiao;LAI Chong-de;DAI Yi-min;ZHANG Qing-hua;ZHONG Qi-wang;LAI Fen-ju(School of Biology Science and Engineering,Jiangxi Agricultural University;School of Animal Science and Technology,Jiangxi Agricultural University)

机构地区：[1]i.江西农业大学生物科学与工程学院,江西南昌330045 [2]江西农业大学动物科学技术学院,江西南昌330045

出　　处：《中国兽医学报》2020年第1期72-76,共5页Chinese Journal of Veterinary Science

基　　金：国家自然科学基金资助项目(31360617);江西省自然科学基金资助项目(20161BAB204183);江西农业大学研究生创新专项资金资助项目(NDYC2018-S004)

摘　　要：为了鉴定胞内劳森菌(Lawsonia intercellularis,LI)外膜蛋白LI0902在细菌感染中的作用,本试验以胞内劳森菌疫苗DNA为模板,设计特异性引物进行PCR扩增,并成功构建诱饵表达载体pGBKT7-LI0902。将通过菌落PCR和测序验证的质粒转化酵母菌株Y2H Gold后,发现LI0902能在酵母细胞中正确表达,对酵母细胞无毒性,且无自激活活性。将含有pGBKT7-LI0902的酵母菌株Y2H Gold与猪肠上皮细胞系IPEC-1 cDNA文库菌株Y187进行杂交,筛选、验证、测序后得到3个与LI0902相互作用的宿主蛋白。本试验研究了胞内劳森菌与猪肠上皮细胞相互作用,为揭示增生性肠炎的致病机理提供了线索。To identify the role of outer membrane protein LI0902 of Lawsonia intercellularis in bacterial infection,The DNA of Lawsonia intercellularis vaccine was used as a template,specific primers were designed to be amplified by PCR and bait plasmids pGBKT7-LI0902 was constructed.Plasmids verified by colony PCR and sequencing were transformed into yeast strain Y2 H Gold.The well expression of LI0902 was confirmed by Western blot,and showed no cytotoxicity and self-activation.The yeast strains Y2 H Gold which contained bait vector pGBKT7-LI0902 was mated with the IPEC-1 cDNA library strain Y187.3 candidate interacting proteins of LI0902 were verified.These results provided good clues for interpreting Lawsonia intercellularis host-pathogen interactions and revealing the pathogenesis of proliferative enteritis.

关 键 词：胞内劳森菌 IPHC-1 酵母双杂交 互作蛋白 筛选 

分 类 号：S852.65[农业科学—基础兽医学]