利用不稳定质粒和 I-SceI 内切酶在土拉弗朗西斯菌中常规生成无标记缺失突变体。
Utilization of an unstable plasmid and the I-SceI endonuclease to generate routine markerless deletion mutants in Francisella tularensis.
机构信息
Department of Microbiology and Molecular Genetics, University of Pittsburgh School of Medicine, Pittsburgh, PA 15261, USA.
出版信息
J Microbiol Methods. 2010 Jan;80(1):106-8. doi: 10.1016/j.mimet.2009.10.013. Epub 2009 Oct 29.
Abstract
We engineered an efficient system to make Francisella tularensis deletion mutations using an unstable, poorly maintained plasmid to enhance the likelihood of homologous recombination. For counterselection, we adapted a strategy using I-SceI, which causes a double-stranded break in the integrated suicide vector, forcing a second recombination to mediate allelic replacement.
摘要
我们设计了一个高效的系统，使用不稳定、维护不良的质粒来制造土拉弗朗西斯菌缺失突变，以提高同源重组的可能性。为了进行反向选择，我们采用了一种使用 I-SceI 的策略，该策略会在整合的自杀载体中造成双链断裂，迫使第二次重组介导等位基因替换。
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