Combe M, Lemeland J, Pestel-Caron M, Pons J
G.R.A.M. (Groupe de Recherche sur les Antimicrobiens et les Microorganismes, EA 2656), U.F.R. Médecine-Pharmacie de Rouen, 22 Boulevard Gambettta, 76183, Rouen, France.
FEMS Microbiol Lett. 2000 Apr 15;185(2):169-74. doi: 10.1111/j.1574-6968.2000.tb09057.x.
An optimized multilocus enzyme electrophoresis method, which involves polyacrylamide-agarose gel electrophoresis followed by electrophoretic transfers on nitrocellulose sheets, was developed for the analysis of enzyme polymorphism in several aerobic and anaerobic bacterial species including Staphylococcus aureus, Streptococcus pneumoniae, S. agalactiae, Klebsiella pneumoniae and K. oxytoca, Clostridium bifermentans and C. sordellii, and Prevotella bivia. Serial electrophoretic transfers (during 5-15 min each) from a single polyacrylamide gel could be achieved for most enzymes studied, and allowed an increased definition of enzyme bands on nitrocellulose as compared to migration gels. Four enzymes, which could not be blotted in such conditions, could still be stained in gels after blotting. Thus, the method allowed the combined analysis of several enzymes after a single gel electrophoresis separation. The analysis of enzyme polymorphism in the various species studied raised the interest of polymorphic loci such as esterase or glutamic-oxaloacetic transaminase for epidemiologic studies. The method characterized a genetic diversity of enzyme loci of S. pneumoniae higher than previously reported, and is thus convenient for the analysis of genetic relationships between related isolates. Since the present method reduces the tediousness of multilocus enzyme electrophoresis and requires experimental conditions that are not specific for the bacterial population studied, it may be proposed for rapid population genetics analysis of a wide variety of bacteria.
开发了一种优化的多位点酶电泳方法，该方法包括聚丙烯酰胺 - 琼脂糖凝胶电泳，随后在硝酸纤维素膜上进行电泳转移，用于分析几种需氧和厌氧细菌物种中的酶多态性，这些细菌包括金黄色葡萄球菌、肺炎链球菌、无乳链球菌、肺炎克雷伯菌和产酸克雷伯菌、双发酵梭菌和索氏梭菌，以及二路普雷沃菌。对于大多数所研究的酶，可以从单个聚丙烯酰胺凝胶进行连续电泳转移（每次5 - 15分钟），与迁移凝胶相比，这使得硝酸纤维素膜上的酶带清晰度增加。在这种条件下无法印迹的四种酶，在印迹后仍可在凝胶中染色。因此，该方法允许在单次凝胶电泳分离后对几种酶进行联合分析。对所研究的各种物种中的酶多态性分析引起了对酯酶或谷草转氨酶等多态性位点在流行病学研究中的关注。该方法表征了肺炎链球菌酶位点的遗传多样性高于先前报道，因此便于分析相关分离株之间的遗传关系。由于本方法减少了多位点酶电泳的繁琐性，并且所需的实验条件并非所研究细菌群体特有的，因此可用于多种细菌的快速群体遗传学分析。
