一种获得重组传染性法氏囊病病毒的新方法  

Generation of Recombinant Infectious Bursal Disease Virus by Transfection of Chicken Embryo Fibroblasts with cDNA of IBDV Genome

作　　者：曹永长[1] 毕英佐[1] 刘丽[1] 

机构地区：[1]华南农业大学动物科学系,广东广州510642

出　　处：《中国兽医学报》2002年第1期19-21,共3页Chinese Journal of Veterinary Science

基　　金：国家自然科学基金资助项目 (3 9870 5 5 0 )

摘　　要：采用 RT- PCR技术 ,从传染性法氏囊病病毒 (IBDV)细胞适应株 GZ911中扩增了 A片段的 2个片段 GA5和 GA3,从中国分离的 IBDV超强毒株 L X中扩增了 B片段的 2个片段 L B5和 L B3。将 GA5和 GA3克隆到质粒p Bss K的 Eco R / Kpn 位点 ,将 L B5和 L B3克隆到 p Bss K的 Eco R / Xba 位点 ,获得携带 GZ911完整 A片段基因的质粒 GA- p Bss K和携带 L X完整 B片段基因的质粒 L B- p Bss K。然后将 GZ911的 A片段 c DNA和 L X的 B片段c DNA分别插入带有巨细胞病毒立即早期加强子 /启动子的质粒 p AL TER- MAX中 ,获得重组质粒 GA- p AL TER和L B- p AL TER。用 GA- p AL TER和 L B- p AL TER共转染鸡胚成纤维细胞 (CEF) ,获得具有感染活性的重组 IBDV,命名为 r IBDV- GZ+L X。该方法的建立为在体外对 IBDV的基因组进行遗传操作 ,进而彻底了解 IBDV基因的结构与功能的关系打下了基础。By using reverse transcription polymerase chain reaction technique, two cDNA fragments, GA5 and GA3, were amplified from the genome of GZ911, a CEF adapted strain of infectious bursal disease virus. Two fragments, LB5 and LB3, were also amplified from the genome of LX, a very virulent IBDV strain isolated in China. Fragments GA5 and GA3 were inserted into the EcoRⅠ/KpnⅠ site of pBssK to obtain the plasmid GA pBssK containing full length cDNA of segment A of GZ911 strain. Fragments LB5 and LB3 were cloned into the EcoRⅠ/XbaⅠ site of pBssK to obtain the plasmid LB pBssK containing full length cDNA of segment B of LX strain. The full length cDNA fragment of segment A of GZ911 and the full length cDNA fragment of segment B of LX strain were then subcloned into the expression vector pALTER MAX carrying cytomegalovirus immediate early inhancer/promoter to obtain plasmids GA pALTER and LB pALTER. A recombinant IBDV capable of infecting chicken embryo fibroblasts, named rIBDV GA+LX, was obtained after transfection of plasmids GA pALTER and LB pALTER into CEF. This approach may be used to manipulate the genome of IBDV and display a role in the study of the relation between structure and functions of the gene of IBDV.

关 键 词：传染性法氏囊病病毒 转染 重组病毒 

分 类 号：S852.65[农业科学—基础兽医学]