含有尿嘧啶DNA糖苷酶基因表达载体的构建

Construction of an Expression Vector Containing Uracil DNA Glycosylase (UNG) Gene

采用自行设计的带酶切位点的上下游引物，用PCR技术，从大肠杆菌ATCC23743的DNA中，获得了含有编码尿嘧啶DNA糖苷酶（UNG）基因的PCR产物，以该PCR产物构建了重组克隆质粒PGEM／UNG，DNA序列分析结果确证其中含有完整的UNG基因．利用该重组质粒，进一步构建了重组表达质料pBV／UNG．

Using a pair of specific primers (PⅠ and PⅡ), a PCR fragment containing the gene encodingfor uracil DNA glycosylase (UNG) was obtained from the total DNA of the Escherichia coli K-12 strain,ATCC23743. The PCR fragment was cloned into plasmid pGEM-7Zf(+) after the latter had been digested with restrictive enzymes, Eco R I and Nsi I. The nucleotide sequence of the cloned fragment in the recombinant plasmid confirmed that the cloned UNG gene was correct and intact. Thereafter, the UNG genefragment was subcloned into an expression plasmid pBV220, located downstream of the bacteriophage PLpromoter. The UNG gene in pBV/UNG was shown to be intact.