Modified plasmid isolation method for Clostridium perfringens and Clostridium absonum

Abstract

A rapid plasmid isolation procedure for Clostridium perfringens and C. absonum is described. The ratio of culture volume to lysis buffer volume was found to be crucial for efficient plasmid isolation. The method can be scaled up, without difficulty, for large-scale plasmid preparation.

Plasmid-Encoded Degradation ofp-Nitrophenol byPseudomonas cepacia

“…The ability of PNP 0 derivatives to revert to growth on PNP was determined by inoculating approximately 10 10 -10 11 cells from a washed, overnight nutrient broth culture onto MM agar containing PNP. Plasmid DNA isolations were carried out as described previously (1,2,18,24,25,26,33), but the method of O'Sullivan and Klaenhammer (24) was found to be most appropriate and was routinely used. Purification of DNA by Cesium Chloride-Ethidium Bromide (CsCl-EtBr) density gradients was carried out as described earlier (27).…”

Molecular genetics and pathogenesis of Clostridium perfringens.

“…perfringens plasmids (3,13,14,18,70,92,113,118,149,158,162,167,168,192). Plasmid profiling also has been used for strain differentiation in C. perfringens (118,149).…”

Development of a new shuttle plasmid system for Escherichia coli and Clostridium perfringens

“…Plasmid pJU122 (13) was isolated from C. perfringens VPI 12502 by using a previously described method for obtaining a cleared lysate (10). The lysate was electrophoresed on a 0.7% agarose gel, and the plasmid was electroeluted from the appropriate gel segment.…”

“…Plasmid isolation from C. perfringens L-13, transformed by plasmid pHR106 (the plasmid constructed in this study), was done by incubation at 37°C in BHIS-Cam to an optical density at 650 nm of about 0.55 (8 to 10 h). Plasmid DNA was isolated from a 600-ml culture volume as described previously (10) by using a treatment ratio of culture volume to lysis volume, as defined therein, of 6; lysozyme was omitted. This step was followed by purification on cesium chloride-ethidium bromide gradients.…”