法氏囊病毒B87株B节段cDNA定点突变及真核表达载体的构建

The cDNA site- directed mutagenesis and the construction of the eukaryotic expression vector for the segment B of IBDV B87 strain

为了探索构建具有分子标记的传染性法氏囊病毒(IBDV)B87株B节段真核表达载体,试验在不改变载体氨基酸序列的基础上,采用PCR方法将新的酶切标记位点引入表达载体上,通过酶切鉴定、测序鉴定及转染对载体表达情况进行观察和分析。结果表明:目的片段与载体片段连接正确,构建的绿色荧光蛋白真核表达载体pEGFP-NB能在Vero细胞内表达。说明成功构建了含分子标记的突变真核表达载体pEGFP-NB。

To explore and construct the eukaryotic expression vector with molecular markers of genomic segment B of infectious bursal disease virus （IBDV） vaecine strain B87,a new enzyme locus was induced into the expression vector using PCR method on the basis of without changing the amino acid sequence of the vector. The vector expression was observed and analyzed using restriction enzyme digestion, sequencing and the transfection. The results showed that the connection that the target fragment ligated with the vector fragment was correct, and the constructed eukaryotic expression vector pEGFP - NB with green fluorescent protein, could be expressed in Veto cells. The results indicate the mutant eukaryotic expression vector pEGFP - NB with molecular markers is successfully constructed.