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为了构建山羊痘病毒表达载体,试验利用融合PCR的方法连接4种组合的3个不同来源的基因片段,插入含有TK基因的载体中,快速构建羊痘病毒表达载体,并转染至BHK-21细胞中进行验证。结果表明:利用融合PCR技术成功融合了P11-7.5-ZSG、P11-7.5-GFP、I1L-7.5-ZSG、I1L-7.5-GFP 4个组合中的3个基因片段,产物长度均在1.2 kb左右,并插入PGEM-TK载体中,构建了4个表达载体,测序结果与理论上100%同源,转染BHK-21细胞可见绿色荧光。说明试验成功构建了4个山羊痘病毒的表达载体。
In order to construct a goat poxvirus expression vector, we cloned three different gene fragments from four different combinations by fusion PCR and inserted into the vector containing TK gene to construct the goatpox virus expression vector and transfected it into BHK-21 Cells were validated. The results showed that three fragments of P11-7.5-ZSG, P11-7.5-GFP, I1L-7.5-ZSG and I1L-7.5-GFP were successfully fused using the fusion PCR technique. , And inserted into PGEM-TK vector. Four expression vectors were constructed. The sequencing results were 100% homologous to the theory. The BHK-21 cells were transfected with green fluorescence. Explain the successful construction of four goat pox virus expression vector.