支原体肺炎靶向染色体敲除。
Targeted chromosomal knockouts in Mycoplasma pneumoniae.
机构信息
J. Craig Venter Institute, 9704 Medical Center Drive, Rockville, MD 20850, USA.
出版信息
Appl Environ Microbiol. 2010 Aug;76(15):5297-9. doi: 10.1128/AEM.00024-10. Epub 2010 Jun 11.
Abstract
Most gene knockouts in mycoplasmas are achieved through labor-intensive transposon mutagenesis. Here, we describe a method for making targeted deletions in Mycoplasma pneumoniae by use of homologous recombination. In this method, M. pneumoniae is transformed with a plasmid carrying an antibiotic resistance marker flanked by 1-kb regions surrounding the target gene. Following selection for the antibiotic resistance, colonies are screened for double crossovers which indicate complete deletion of the target open reading frame.
摘要
大多数支原体的基因敲除都是通过繁琐的转座子诱变来实现的。在这里，我们描述了一种通过同源重组在肺炎支原体中进行靶向缺失的方法。在这种方法中，将携带抗生素抗性标记物的质粒转化到肺炎支原体中，该质粒被目标基因周围 1kb 的区域所包围。在选择抗生素抗性后，筛选出发生两次单交换的菌落，这表明目标开放阅读框已完全缺失。
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