拟构建表达鸡传染性支气管炎病毒(IBV)QS10株S1蛋白的重组复制缺陷型人5型腺病毒,并在本动物上进行初步的免疫学试验.通过RT-PCR获得S1基因,插入腺病毒表达系统穿梭质粒,构建重组穿梭质粒pac-Ad5 CMV-S1,转染293AD细胞获得表达S1蛋白的重组腺病毒.PCR检测显示,S1基因已重组到腺病毒基因组；间接免疫荧光和Western blot检测证明该重组病毒在293AD细胞中真实表达了具有免疫反应性的IBV S1糖蛋白.重组病毒培养滴度可达到107 TCID50·mL-1.免疫接种SPF鸡后,通过ELISA检测,免疫鸡产生了针对IBV的特异性抗体.作者成功构建了表达IBV S1蛋白的重组腺病毒,该重组病毒可在SPF鸡体内诱导产生IBV特异性抗体.%To construct human adenovirus type 5 vectored recombinant virus expressing S1 glyco-protein gene of avian infectious bronchitis virus (IBV), S1 gene of IBV QS10 strain obtained through RT-PCR was inserted into shuttle plasmid of the adenoviral expression system, forming pacAd5 CMV-S1, as well as the backbone plasmid, which was linearized with Pac I and trans-fected into 293AD cells. Detection with PCR revealed that the recombinant virus was successfully packaged with a titer up to 107TCID50 o ml-1. Furthermore, The results of Western blotting and indirect immunofluorescence assay (IFA) indicated that the SI gene was significantly expressed. Sera antibodies against IBV of the SPF chicken immunized with the recombinant virus were detected by enzyme-linked immunosorbent assay (ELISA). The recombinant adenovirus expressing SI gene of IBV was successfully constructed with potent capacity to induce the specific antibodies in chickens and had the good immunogenicity, which laid the foundation for further development of recombinant avian infectious bronchitis vaccine.
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