Laboratoire de Microbiologie et Génétique Moléculaire, Université Paul Sabatier-Centre National de la Recherche Scientifique, 118 Route de Narbonne, 31062 Toulouse Cedex, France.
Appl Environ Microbiol. 2010 Feb;76(4):1095-102. doi: 10.1128/AEM.02123-09. Epub 2009 Dec 18.
The bacterium Burkholderia cenocepacia is pathogenic for sufferers from cystic fibrosis (CF) and certain immunocompromised conditions. The B. cenocepacia strain most frequently isolated from CF patients, and which serves as the reference for CF epidemiology, is J2315. The J2315 genome is split into three chromosomes and one plasmid. The strain was sequenced several years ago, and its annotation has been released recently. This information should allow genetic experimentation with J2315, but two major impediments appear: the poor potential of J2315 to act as a recipient in transformation and conjugation and the high level of resistance it mounts to nearly all antibiotics. Here, we describe modifications to the standard electroporation procedure that allow routine transformation of J2315 by DNA. In addition, we show that deletion of an efflux pump gene and addition of spermine to the medium enhance the sensitivity of J2315 to certain commonly used antibiotics and so allow a wider range of antibiotic resistance genes to be used for selection.
洋葱伯克霍尔德菌是囊性纤维化 (CF) 和某些免疫功能低下患者的病原体。从 CF 患者中分离出的最常见的洋葱伯克霍尔德菌菌株，被用作 CF 流行病学的参考菌株，是 J2315。J2315 基因组分为三个染色体和一个质粒。该菌株几年前被测序，其注释最近发布。这些信息应该允许对 J2315 进行遗传实验，但存在两个主要障碍：J2315 作为转化和接合受体的潜力较差，以及它对几乎所有抗生素的高度耐药性。在这里，我们描述了对标准电穿孔程序的修改，这些修改允许 DNA 常规转化 J2315。此外，我们还表明，删除外排泵基因并在培养基中添加精胺可提高 J2315 对某些常用抗生素的敏感性，从而允许使用更广泛的抗生素抗性基因进行选择。
