非洲猪瘟病毒(Malawi LIL 20/1株)k8R基因的表达与鉴定  

EXPRESSION AND CHARACTERIZATION OF THE AFRICAN SWINE FEVER VIRUS (MALAWI LIL 20/1) GENE k8R

作　　者：孙怀昌[1] Linda K Dixon Parkhouse R M E 

机构地区：[1]江苏农学院动物科学系

出　　处：《江苏农学院学报》1998年第2期1-7,共7页Jiangsu Agricultural Research

基　　金：英国生物技术及生物科学研究委员会资助;农业部资助

摘　　要：蛋白多肽二级结构的电脑预测表明，非洲猪瘟病毒（ＭａｌａｗｉＬＩＬ２０／１株）ｋ８Ｒ基因编码带有多个疏水氨基酸小区的２７ｋＤａ蛋白质。该基因的ＰＣＲ产物克隆入质粒ｐＧＥＸ－２Ｔ后，在大肠杆菌中表达４２～５４ｋＤａ不溶性ＧＳＴ－ｋ８Ｒ融合蛋白。此表达蛋白能被针对不同非洲猪瘟病毒株的猪免疫血清识别。在非洲猪瘟病毒细胞适应株感染的细胞中，针对ｋ８Ｒ大肠杆菌表达产物的单抗能检测出２７ｋＤａ特异病毒蛋白。进一步鉴定表明，ｋ８Ｒ基因编码的蛋白质为病毒非结构蛋白，不发生糖基化，出现于病毒感染周期的晚期，主要集中于靠近细胞核的病毒复制部位。用大肠杆菌表达的ＧＳＴ－ｋ８Ｒ融合蛋白免疫猪未能抵抗ＭａｌａｗｉＬＩＬ２０／１强毒株攻击。African swine fever virus(ASFV) (Malawi LIL 20/1) gene k8R was predicted by computing to encode a 27 kDa protein with multiple small stretches of hydrophobic amino acids The k8R gene was amplifed by PCR, of which product was cloned into plasmid pGEX 2T for expression in E coli After IPTG induction, the gene expressed GST k8R fusion proteins with moleculer weights ranging from 42 to 54 kDa The affinity purified proteins were recognized by pig antisera to different ASFV strains In the virally infected cells a 27 kDa protein was detected by monolonal antibodies raised against the E coli expressed k8R protein Further characterization showed that the k8R gene encoded a late non structural protein, which was non glycosylated and associated with the viral assembly sites Immunization with the E coli expressed GST k8R fusion protein failed to protein pigs from challenge with the virulent Malawi LIl 20/1 Strain

关 键 词：猪 基因 表达 鉴定/非洲猪瘟病毒 

分 类 号：S858.285.3[农业科学—临床兽医学]