Isolation and Characterization of Plasmid DNA in Streptococcus cremoris†

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Nine industrially important strains of Streptococcus cremoris (HP, AM2, ML1, WC, C3, R1, E8, KH, and Wg2) were shown to possess a diversity of plasmid molecules. Molecular weights of plasmids were determined from their relative mobilities after agarose gel electrophoresis and via electron microscopy. To illustrate the varied plasmid sizes, strain HP contained plasmids of 26, 18, 8.5, 3.3, and 2 megadaltons (Mdal); strain ML1 contained plasmids of 29, 18, 9, 4, 2.2, and 1.8 Mdal; and strain AM2 had plasmids of 42, 27, 16, and 8.4 Mdal. The numbers of plasmids observed in the other strains were 6, 5, 5, 7, 5, and 4 for C3, E8, KH, R1, WC, and Wg2, respectively. A spontaneous proteinase-negative (Prt−) mutant of HP was missing the 8.5-Mdal plasmid, which suggests that in this strain proteinase activity could be linked to this particular plasmid. A lactose-negative (Lac−) Prt− mutant of ML1 lacked the 2.2-Mdal plasmid. Under the conditions employed, antibiotic sensitivity and heavy-metal susceptibility did not correlate with the missing plasmid in Prt− HP or in the Lac− Prt− ML1. Curing experiments with AM2, using acridine dyes and elevated temperatures, did not yield Lac− variants. AM2 was also cultured at high dilution rates in a chemostat for 168 h by using a buffered milk or lactic broth at 18 or 32°C with no selection of Lac− derivatives. The inability to obtain Lac− variants under conditions known to facilitate plasmid elimination suggests that lactose metabolism is not plasmid-mediated in AM2.

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