传染性喉气管炎病毒gD基因重组禽痘病毒的制备、纯化及初步鉴定

目的制备传染性喉气管炎病毒(ILTV)gD基因重组禽痘病毒,并进行纯化及初步鉴定。方法将含有ILTV河南长葛株gD基因的禽痘病毒转移载体转染禽痘病毒感染的鸡胚成纤维细胞,采用蓝色蚀斑筛选纯化重组禽痘病毒,并经PCR及间接免疫荧光试验(IFA)进行鉴定。结果经过5轮蚀斑筛选纯化,获得1株重组禽痘病毒rFPV-ILTV-gD,其PCR产物经琼脂糖凝胶电泳,可见约1400bp的ILTVgD特异性条带。IFA检测显示重组禽痘病毒中的ILTVgD基因得到了表达。结论已成功制备并纯化了ILTVgD基因重组禽痘病毒,为预防ILT提供了新的途径。

Preparation, Purification and Preliminary Identification of Recombinant Fowl Pox Virus Expressing Infectious Laryngotracheitis Virus Glycoprotein D Gene

Objective To prepare,purify and preliminarily identify the recombinant fowl pox virus(FPV)expressing infectious laryngotracheitis virus(ILTV)glycoprotein D(gD)gene. Methods The recombinant fowl pox virus transfer vector pSY681-gD-LacZ carrying gD gene of ILTV HNCG strain was transfected to chick embryo fibroblast(CEF)infected with FPV-017 strain,then recombinant FPV was purified by blue plaque screening and identified by PCR and indirect IFA. Results After purification by 5 cycles of blue plaque screening,a recombinant FPV strain rFPV-ILTV-gD was obtained. The specific ILTV gD band at a length of about 1 400 bp was observed on the agarose gel electrophoretic profile of PCR product of rFPV-ILTV-gD,and IFA showed the expression of ILTV gD. Conclusion The recombinant FPV expressing ILTV gD gene was successfully prepared and purified,which provided a novel route for the prevention of ILT.