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伪狂犬病病毒通用转移载体的构建及应用
引用本文:钱平,李祥敏,陈焕春,肖少波,仇华吉,童光志. 伪狂犬病病毒通用转移载体的构建及应用[J]. 中国预防兽医学报, 2003, 25(1): 16-20
作者姓名:钱平  李祥敏  陈焕春  肖少波  仇华吉  童光志
作者单位:1. 华中农业大学畜牧兽医学院动物病毒室,湖北,武汉,430070
2. 中国农业科学院哈尔滨兽医研究所兽医生物技术国家重点实验室,黑龙江,哈尔滨,150001
摘    要:将来自质粒pFSV40的300bpBamHI/PstI片段[其中含有SV40poly(A)和部分多克隆位点]插入到质粒pUSK相应的酶切位点中,获得重组质粒USKSV40。该重组质粒中gG基因5’端编码区缺失了428bp。将来自质粒pcDNA3.1( )的946bpBglⅡEcoRI片段(其中含CMV启动子及部分多克隆位点)插入到质粒pUSKSV40的BamHIEcoRI位点,构建了通用载体pPRVCMV-uni,其中含有CMV启动子,SV40poly(A)以及NheI,Pme1,BamHI,BstXI,EcoRI,StuI,XbaI等7个单一克隆位点,将eGFP基因插入到该通用载体的BamHI和EcoRI之间,用所获得的转移载体与TK/gG-/LacZ^ PRV基因组共转染PK-15细胞,经检测eGFP基因在重组伪狂犬病病毒中获得表达,从而证实该通用载体的构建是可行的。本研究研制以伪狂犬病病毒为载体的二价或多价基因工程疫苗奠定了物质基础。

关 键 词:伪狂犬病 伪狂犬病病毒 通用转移载体 重组伪狂犬病病毒 基因工程疫苗
文章编号:1008-0589(2003)01-0016-05
修稿时间:2002-05-20

Construction and Application of a Universal Vector of Pseudoriabies Virus
QIAN Ping ,LI Xiang-min ,CHEN Huan-chun ,XIAO Shao-bo ,QIU Hua-ji ,TONG Guang-zhi. Construction and Application of a Universal Vector of Pseudoriabies Virus[J]. Chinese Journal of Preventive Veterinary Medicine, 2003, 25(1): 16-20
Authors:QIAN Ping   LI Xiang-min   CHEN Huan-chun   XIAO Shao-bo   QIU Hua-ji   TONG Guang-zhi
Affiliation:QIAN Ping 1,LI Xiang-min 1,CHEN Huan-chun 1*,XIAO Shao-bo 1,QIU Hua-ji 2,TONG Guang-zhi 2
Abstract:A 300bp fragment containing SV40 poly(A) signal and part of nulitiple cloning sites(MCS),was inserted into Bam HI Pst I of pUSK,resulting in a recombinant pUSKSV40,which gave rise to a deleton of 428bp in the 5'-end of PRV gE gene.A 946bp Bgl II Eco RI fragment from pcDNA3.1(+) inchuding human cytomegalovirus(hCMV) promoter and partial multiple cloning sites was cloned into the Ban HI Eco RI sites of the pUSKSV40,resulting a universal transfer vector,which contained the hCMV promoter,SV40 poly(A) signal and 7 nultiple cloning sites including Nhe I, Pme l, Bam HI, Bst XI, Eco RI, Stu I and Xba I.To test the applicability of the universal vactor,e GFP was selected as a reporter gene and inserted into the Bam HI Eco RI Sites of the pPPVCMV-uni to obtain a recombinant plasmid pPRVCMV-eGFP.The PK-15 cell cultures were cotransfect with pPRVCMV-eGFP and the genome of a TK -/gG -/LacZ + PRV,generating a recombinant PRV,which was confirmed to express eGFP.The results indicated that the constructed pPRVCMV-uni was feasible.It will be useful for developing the recombinant PRV exzpressing foreign gene(s).
Keywords:Pseudoriabies virus  Universal vector  eGFP  Recombinant pseudoriabies virus
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