Genetic studies in Borrelia burgdorferi have been hindered by the lack of a nonborrelial selectable marker. Currently, the only selectable marker is gyrBr , a mutated form of the chromosomal gyrB gene that encodes the B subunit of DNA gyrase and confers resistance to the antibiotic coumermycin A1. The utility of the coumermycin-resistantgyrBr gene for targeted gene disruption is limited by a high frequency of recombination with the endogenousgyrB gene. A kanamycin resistance gene (kan) was introduced into B. burgdorferi, and its use as a selectable marker was explored in an effort to improve the genetic manipulation of this pathogen. B. burgdorferi transformants with the kan gene expressed from its native promoter were susceptible to kanamycin. In striking contrast, transformants with thekan gene expressed from either the B. burgdorferi flaB or flgB promoter were resistant to high levels of kanamycin. The kanamycin resistance marker allows efficient direct selection of mutants in B. burgdorferi and hence is a significant improvement in the ability to construct isogenic mutant strains in this pathogen.
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机译：缺乏疏螺旋选择标记阻碍了勃氏疏螺旋体的遗传研究。目前，唯一的选择标记是 gyrB r ，这是一种染色体 gyrB 基因的突变形式，编码DNA回旋酶的B亚基并具有抗性抗生素coumermycin A 1 。耐香豆霉素的 gyrB r 基因在靶向基因破坏中的应用受到与内源性 gyrB 基因重组的高频率的限制。将卡那霉素抗性基因（ kan ）引入 B。探索了burgdorferi 及其作为选择标记的用途，以期改进对该病原体的遗传控制。 B。从其天然启动子表达 kan 基因的burgdorferi 转化子对卡那霉素敏感。与之形成鲜明对比的是，具有 kan 基因的转化子是从 B表达的。 burgdorferi flaB 或 flgB 启动子对高水平的卡那霉素具有抗性。卡那霉素抗性标记允许有效地直接选择 B中的突变体。 burgdorferi ，因此在该病原体中构建等基因突变菌株的能力有了显着提高。
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