刀豆蛋白A纯化猪流行性腹泻病毒的研究  

Research on the purification of porcine epidemic diarrhea virus by Concanavalin A

作　　者：李兰[1] 张浩明[1] 杨利[1] 程海卫[1] 侯立婷[1] 张元鹏[1] 侯继波[1] 陈瑾[1] 郑其升[1,2,3] LI Lan;ZHANG Haoming;YANG Li;CHENG Haiwei;HOU Liting;ZHANG Yuanpeng;HOU Jibo;CHEN Jin;ZHENG Qisheng(National Research Center of Engineering and Technology for Veterinary Biologicals,Jiangsu Academy of Agricultural Sciences,Nanjing 210014,China;Cultivation Base of State Key Laboratory for Food Quality and Safety of Jiangsu Province,Nanjing 210014,China;GuoTai(Taizhou)Center of Technology Innovation for Veterinary Biologicals,Taizhou 225300,China)

机构地区：[1]江苏省农业科学院动物免疫工程研究所,南京210014 [2]江苏省食品质量安全重点实验室国家重点实验室培育基地,南京210014 [3]兽用生物制品(泰州)国泰技术创新中心,泰州225300

出　　处：《中国农业大学学报》2023年第11期125-130,共6页Journal of China Agricultural University

基　　金：国家自然科学基金项目(32102690);江苏省农业科技自主创新资金(CX(21)3135)。

摘　　要：为开发一种新型高效、低成本的猪流行性腹泻病毒(Porcine epidemic diarrhea virus,PEDV)抗原纯化方法,本研究尝试了一种基于凝集素(刀豆蛋白A,Con A)的PEDV浓缩纯化方法。通过Con A与PEDV病毒粒子的特异性结合,形成Con A-PEDV复合物,然后通过低速离心和竞争性解离即可实现病毒纯化。结果表明:1)当Con A的使用浓度为80μg/mL,室温作用2 h,4000 r/min离心15 min时,纯化效果较佳;2)利用Tris-HCl缓冲液(pH 7,0.01 mol/L,内含200 mmol/L甲基-α-D-吡喃甘露糖苷和50 mmol/L NaCl)进行解离时,较PBS缓冲液有更高的解离效率;3)该方法病毒回收率大约为80%,总蛋白去除率可达90%。综上,本研究利用Con A纯化PEDV是可行的,为高质量PEDV灭活疫苗的研发奠定基础。In order to develop a new efficient and low-cost method for antigen purification of porcine epidemic diarrhea virus(PEDV),apurification method based on the lectin(Concanavalin A,Con A)was explored.By specifically binding Con A with PEDV virus particles,a Con A-PEDV complex was formed,and the virus was purified through lowspeed centrifugation and competitive dissociation.The results showed that:1)The suitable concentration of Con A was 80μg/mL,the action time between Con A and PEDV was for 2hours,the centrifugal condition was 4000r/min for 15min;2)The solution contained methyl-α-D-mannopyranoside and NaCl at the concentration of 200mmol/L and 50mmol/L respectively had higher dissociation efficiency compared with PBS buffer;3)The recovery rate of the purification method was approximately 80%,and the total protein removal rate could reach 90%.In conclusion,it is feasible to purify PEDV by Con A,which can lay a foundation for the development of high-quality inactivated vaccine for PEDV.

关 键 词：刀豆蛋白A 猪流行性腹泻病毒 纯化 

分 类 号：S855.3[农业科学—临床兽医学]