pUC19K质粒的构建及其在布鲁氏菌突变株构建中的应用

突变株的构建是细菌基因功能研究的前提。本研究构建了一个可用于布鲁氏菌突变株构建的自杀质粒。在pUC19质粒的多克隆位点插入卡那霉素抗性基因，在该基因两侧添加多个酶切位点，构建成为pUC19K。利用该质粒，我们构建了布鲁氏菌外膜蛋白Omp25基因的突变株。结果表明，利用该自杀质粒，通过一轮筛选即可得到目标基因被抗性基因替换的突变株。pUC19K质粒的构建及成功应用，为布鲁氏菌突变株的构建提供了一个快速有效的手段，也为布鲁氏菌的基因功能研究奠定了基础。

Construction of Plasmid pUC19K and Its Application in Brucella Mutant

Mutant construction is a prerequisite work in bacterial gene function studies.A new suicide plasmid was constructed to produce Brucella mutant.A kanamycin gene was inserted into the multi-cloning site of pUC19,and on either side of the resistance gene,several enzyme sites were inserted,resulting in the new plasmid pUC19K.Using this plasmid,a replacement mutant of Brucella omp25 gene was constructed.As the results showed,the mutant could be generated by only one round of selection with high efficiency.Therefore,the construction and application of pUC19K provides a new suicide plasmid for Brucella mutant construction and would accelerate gene function analysis of Brucella.