表达猫瘟病毒VP2蛋白重组减毒鼠伤寒沙门菌SL1344ΔcrpΔasd(pYA3493-VP2)构建及特性研究  

Construction and characterization of recombinant attenuated Salmonella typhimurium SL1344ΔcrpΔasd(pYA3493)expressing VP2 protein of feline panleukopenia virus

作　　者：王文婕 杜付熙 郁川 尚珂 余祖华[1,2] 李静 贾艳艳[1,2] 廖成水 丁轲[1,2] 张春杰 程相朝[1,2] 陈松彪 WANG Wen-jie;DU Fu-xi;YU Chuan;SHANG Ke;YU Zu-hua;LI Jing;JIA Yan-yan;LIAO Cheng-shui;DING Ke;ZHANG Chun-jie;CHENG Xiang-chao;CHEN Song-biao(Luoyang Key Laboratory of Live Carrier Biomaterial and Animal Disease Prevention and Control,Luoyang 471003,China;Laboratory of Functional Microbiology and Animal Health,Henan University of Science and Technology,Luoyang 471003,China;Animal Diseases and Public Health Engineering Research Center of Henan Province,Luoyang 471099,China)

机构地区：[1]洛阳市活载体生物材料与动物疫病防控重点实验室,河南洛阳日471003 [2]河南科技大学功能微生物与畜禽健康实验室,河南洛阳471003 [3]河南省动物疫病与公共卫生工程研究中心,河南洛阳471099

出　　处：《中国兽医科学》2023年第2期189-197,共9页Chinese Veterinary Science

基　　金：河南科技大学博士研究启动专项基金(13480104);河南省自然科学基金项目(182300410078)。

摘　　要：为构建稳定表达猫瘟病毒(FPV)免疫原性蛋白VP2减毒鼠伤寒沙门菌重组菌株,以洛阳分离株基因组为模板扩增VP2基因,并克隆至原核表达载体pYA3493,然后电转化至减毒鼠伤寒沙门菌SL1344ΔcrpΔasd,构建重组菌株SL1344ΔcrpΔasd(pYA3493-VP2),并对其表型及生长特性、遗传稳定性、生物被膜、运动性以及毒力等进行分析。结果显示,携带FPVVP2基因重组减毒沙门菌SL1344ΔcrpΔasd(pYA3493-VP2)重组菌构建成功并表达出外源蛋白;SL1344ΔcrpΔasd(pYA3493-VP2)的生化、生长特性与空载体互补菌SL1344ΔcrpΔasd(pYA3493)相似,较亲本菌株SL1344失去了利用麦芽糖、蔗糖、甘露醇、山梨醇等碳源能力,生长速度、运动性、生物膜形成能力相较亲本株SL1344均发生了显著降低,但小鼠LD50较亲本株提高126倍。结果表明,成功构建了稳定携带FPV VP2蛋白新型减毒鼠伤寒沙门菌SL1344ΔcrpΔasd(pYA3493-VP2),该重组菌能够有效递呈所携带外源基因并稳定表达,为进一步预防FPV感染和新型疫苗研发提供理论依据。To construct a recombinant strain of Salmonella typhimurium that stably expressed the immunogenic protein VP2 of feline panleukopenia virus(FPV).Firstly,VP2 gene was amplified by using the genome of FPV Luoyang isolate as a template,and cloned into prokaryotic expression vector p YA3493.Then,the recombinant vector p YA3493-VP2 was electrically transformed into Salmonella typhimurium SL1344ΔcrpΔasd and obtained recombinant strain SL1344ΔcrpΔasd(p YA3493-VP2).Lastly,the recombinant strain phenotype,growth characteristics,genetic stability,biofilm,motility and virulence were analyzed.The results showed that recombinant strain SL1344ΔcrpΔasd(p YA3493-VP2)was successfully constructed and expressed foreign protein.The biochemical and growth characteristics of SL1344ΔcrpΔasd(p YA3493-VP2)were similar to those of the empty vector complementary strain SL1344ΔcrpΔasd(p YA3493).Compared with the parental strain SL1344,the strain lost the ability to utilize maltose,sucrose,mannitol,sorbitol and other carbon sources,and its growth rate motility and biofilm formation ability of the recombinant strain were significantly decreased,but the mouse LD50strain was 126 times higher than the parental strain.These results indicated that the novel attenuated Salmonella typhimurium strain SL1344ΔcrpΔasd(p YA3493-VP2)stably carrying FPV VP2 protein was successfully constructed.The recombinant bacteria can effectively present expressionthe foreign genes,which provides a new idea for further prevention of FPV infection and the development of novel vaccines.

关 键 词：猫瘟病毒 VP2 减毒鼠伤寒沙门菌 特性 

分 类 号：S852.659.2[农业科学—基础兽医学]