采用RT-PCR技术扩增禽呼肠孤病毒ARV S1133毒株和广西分离株R1的δ2基因,将δ2基因克隆至真核表达载体pcDNA3.1(+)载体上,对获得的重组质粒经PCR酶切以及序列分析鉴定.结果表明,插入的片段为δ2目的基因,插入的位置、大小和阅码框架均正确,成功构建了真核表达载体重组质粒peDNA-S1133-δ2、pcDNA-R1-δ2;分别免疫SPF鸡2次后,用ARV S1133进行攻毒,使用RT-PCR方法进行检测.结果表明,pcDNA-S1133-δ2、pcDNA-R1-δ2 DNA疫苗免疫组检出率与对照组差异显著(p0.05).使用ARVδ2基因构建的真核表达质粒来免疫SPF鸡,鸡只获得了较好的免疫保护作用.%The U2 gene in ARV S1133 & R1 of avian reovirus, isolated from CJuangxi, was amplified by using RT-PCR.The purified RT-Pσ product (σ2 gene) was inserted into pcDNA3.1 (+) vector. The recomhinant plasmid was identified by restriction endonuclease analysis and Pσ. It was proved by DNA sequencing that the acquired recombinant plasmid contains complete U2 gene, the inserting location, size and open reading frame (ORF) were correct, therefore the recombinant DNA vaccine plasmid of pcDNA-S1133-σ2, pcDNA-RI-σ2 was constructed. SPF chickens were immunized with plasmids pcDNA-S1133-σ2 and pcDNA-RI-σ2 twice, then they were challenged with ARV S1133. and then by using RT-Pσ to detect the chickens, the results showed that the positive rate of chickens immunized with pcDNA-S1133-σ2 and pcDNA-R1-σ2 had significant difference with that of control (P＜0.05), and no significant difference with chickens immunized with inactivated vaccines or σ2 protein. The result showed that the SPF chickens immunized with pcDNA-S1133-σ2 and pcDNA-RI-σ2 could obtain hetter immune protection.
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