用于在天然不可转化、低GC含量的革兰氏阳性细菌中进行高效等位基因替换的新型载体。
New vector for efficient allelic replacement in naturally nontransformable, low-GC-content, gram-positive bacteria.
作者信息
Arnaud Maryvonne, Chastanet Arnaud, Débarbouillé Michel
机构信息
Unité de Biologie des Bactéries Pathogènes à Gram Positif, Institut Pasteur, Paris, France.
出版信息
Appl Environ Microbiol. 2004 Nov;70(11):6887-91. doi: 10.1128/AEM.70.11.6887-6891.2004.
Abstract
A shuttle vector designated pMAD was constructed for quickly generating gene inactivation mutants in naturally nontransformable gram-positive bacteria. This vector allows, on X-Gal (5-bromo-4-chloro-3-indolyl-beta-D-galactopyranoside) plates, a quick colorimetric blue-white discrimination of bacteria which have lost the plasmid, greatly facilitating clone identification during mutagenesis. The plasmid was used in Staphylococcus aureus, Listeria monocytogenes, and Bacillus cereus to efficiently construct mutants with or without an associated antibiotic resistance gene.
摘要
构建了一种名为pMAD的穿梭载体，用于在天然不可转化的革兰氏阳性细菌中快速产生基因失活突变体。该载体在X-Gal（5-溴-4-氯-3-吲哚基-β-D-吡喃半乳糖苷）平板上，能够对丢失质粒的细菌进行快速的比色蓝白鉴别，极大地便于诱变过程中的克隆鉴定。该质粒用于金黄色葡萄球菌、单核细胞增生李斯特菌和蜡状芽孢杆菌，以高效构建带有或不带有相关抗生素抗性基因的突变体。
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