通过 II 类内含子在土拉弗朗西斯菌中进行靶向基因敲除。
Targeted gene disruption in Francisella tularensis by group II introns.
机构信息
South Texas Center for Emerging Infectious Diseases and Department of Biology, University of Texas San Antonio, San Antonio, TX 78249, USA.
出版信息
Methods. 2009 Nov;49(3):270-4. doi: 10.1016/j.ymeth.2009.04.011. Epub 2009 May 4.
Abstract
Francisella tularensis is a highly infectious Gram-negative bacterium that is the causative agent of tularemia. Very little is known about the molecular mechanisms responsible for F. tularensis virulence, in part due to the paucity of genetic tools available for the study of F. tularensis. We have developed a gene knockout system for F. tularensis that utilizes retargeted mobile group II introns, or "targetrons". These targetrons disrupt both single and duplicated target genes at high efficiency in three different F. tularensis subspecies. Here we describe in detail the targetron-based method for insertional mutagenesis of F. tularensis genes, which should facilitate a better understanding of F. tularensis pathogenesis. Group II introns can be adapted to inactivate genes in bacteria for which few genetic tools exist, thus providing a powerful tool to study the genetic basis of bacterial pathogenesis.
摘要
土拉弗朗西斯菌是一种高度传染性的革兰氏阴性细菌，是土拉热的病原体。由于缺乏用于研究土拉弗朗西斯菌的遗传工具，因此对于导致土拉弗朗西斯菌毒力的分子机制知之甚少。我们已经开发了一种用于土拉弗朗西斯菌的基因敲除系统，该系统利用了经重新靶向的移动组 II 内含子，或“targetrons”。这些 targetrons 可高效地破坏三种不同土拉弗朗西斯菌亚种中单拷贝和双拷贝的靶基因。在这里，我们详细描述了基于 targetron 的土拉弗朗西斯菌基因插入诱变方法，这将有助于更好地理解土拉弗朗西斯菌的发病机制。组 II 内含子可适应于遗传工具很少的细菌中的基因失活，从而为研究细菌发病机制的遗传基础提供了强大的工具。
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