Ogryz'ko E P, Nikiforov V G
Genetika. 1988 Oct;24(10):1894-6.
A multicopy plasmid pLMN1 expressing a wild type rpoB gene encoding Escherichia coli RNA polymerase beta subunit gene was constructed. Introduction of this plasmid into rifampicin-resistant RpoB mutants makes them rifampicin-sensitive. Rifampicin-resistant clones appear in such strains with frequencies up to 10(-3), due to recombinational (recA-dependent) transfer of rif-r mutations from chromosome to pLMN1. This provides a simple selection procedure for transfer of any rpoB mutation, together with a rif-r mutation from a chromosome to pLMN1. In this way, we transferred rpoB22 amber mutation to pLMN1 for localization of the mutant codon by DNA sequencing.
构建了一个表达编码大肠杆菌RNA聚合酶β亚基基因的野生型rpoB基因的多拷贝质粒pLMN1。将该质粒导入耐利福平的RpoB突变体中可使其对利福平敏感。由于耐利福平（rif-r）突变从染色体到pLMN1的重组（recA依赖性）转移，在这些菌株中耐利福平的克隆出现频率高达10^(-3)。这为将任何rpoB突变以及耐利福平突变从染色体转移到pLMN1提供了一个简单的选择程序。通过这种方式，我们将rpoB22琥珀突变转移到pLMN1，以便通过DNA测序定位突变密码子。
