Nisin생산 Streptococcus lactis ATCC 11454, IFO 12007과 Lactobacillus casei3012의 plasmid 분포와 세포융합에 관한 연구

Studies on the plasmid pattern and protoplast fusion of nisin producting Streptococcus lactis ATCC 11454, IFO 12007 and Lactobacillus casei 3012

Antibiotic resistant mutants of Streptococcus lactis ATCC l1454, IFO 12007 and Lactobacillus casei 3012 were obtained by acriflavin treatment. Their plasmid DNA was isolated and characterized by agarose gel electrophoresis. Experiments on protoplast generation, regeneration and cell fusion were carried out; the following results were obtained. 1. Three mepicillin and hostacillin resistant mutants of S. lactic CU 3023, CU 3024, CU 2022: two lincocin resistant mutants of S. lactis CU 2022 and L. casei 3012 were obtained. Mutants which have resistance against mepicillin and hostacillin lost their lactose and sucrose fermenting ability. 2. Streptococcus lactis ATCC l1454 and S. lactis CU 3021 had 90 M d single plasmid, whereas S. lactis CU 3023, CU3024 had 9.5 M d single plasmid and S. lacis IFO 12007 contained 40 M d single plasmid. There observed no plasmid band in S. lactis CU 3022 and in L. casei 3012. 3. Cell wall of S. lactis IFO 12007 lysed and protoplast was produced by treatment with 1.0mg-ml of lysozyme at 37C for 30 minutes, whereas L. casei 3012 lysed to produce protoplasts treating with 6.3 ug-ml of mutanolysin under the same conditions. 4. Frequency ocurring osmoticaly stable cells after cell wall lysis ranged from 3.7×10⁻³ in S. lactis CU 3022 to 2.2×1⁻² in S. lactis CU 3021. Protoplasts regenerated with the frequency of 3.1×10⁻³ in S. lactis CU 3022, 2.8×1⁻² in L. casei 3012. Protoplasts between S. lactic CU 3022 and CU 3023 fused and regenerated at 3.12×10⁻³, 1.2×1⁻⁵ between S. lactis CU 3022 and L. casei 3012 under the presence of PEG 4000.