High Yield 3 - The Acetoin Bacillus Subtilis Breeding

This paper takes a bacillus subtilis strain YD-1 preserved in our laboratory as the original strain. The optimal carbon source is glucose and the optimal nitrogen source is syrup and yeast powder. Metal ions Mn2+ are need in the fermentation process. The strain was well produced with the initial pH7.2 of liquid medium under the temperature 37℃,35mL outfit fluid quantity,10h seed,5.7% vaccination quantity,180r/min conditions in shaking table for 3 days.The acetoin yield can reach up to 15.60 g/LWith YD-1 for the original strain,Mutate the strain by UV. After the primary screening and secondary screening, its acetoin production was improved from 15.60g/L to 17.78g/L. Then mutated the strain with DES. After the primary screening and secondary screening, acetoin production was 19.02g/L, which was 21.92% higher than YD-1.Take genetic stability experiment to the mutation strain S-10. Cultive the S-10 for 20 times in the beveled medium. Taking the 1,5,10,15,20 generations for fermentation. Assaying the acetion production of the strain.Result proves that the genetic stability is well, and the strain has the potential of to be production strain.Take preliminarily reserache on the physiological and biochemical characteristics of S-10. The death temperature incred 1℃of S-10 compared with the original strain YD-1. The color of fermentation fluid are also changed. When observe under microscope,we find that the mutated strain are more easily growth with each other. Other physiological indexes have no change.Eeperiments revel that the the best carbon sourceis is glucose and the best nitrogen source is yeast powder and corn slurry mixture. Studies on divalent metal ions Mg2+, Zn2+ Mn2+ and trivalent metal ions Fe3+ to the function of fermentation fluid show that four kinds of ions have promotive affect. Among the four metal ions, Mn+ has the most promotive affection. After optimization, acetoin production reach up to 19.88 g/L.Optimized fermentation conditions of S-10. Its best fermentation conditions is pH 7.0, temperature 36℃, outfit fluid quantity 30mL,8h seed, vaccination quantity 5%, shaking table speed 180r/min,84h fermentation time. After optimization, the production of acetion reach up to 20.50 g/L.Bacillus subtilis YD-1 and S-10 were analyzed with SDS-PAGE. Result show that there is no difference between the two strains. The genetic substances of the two strains were analyzed with RAPD in further experiment. Results showed a different electrophoretic band. Thus we judg that the mutation strain has a slightc change in the DNA level. YD-1 is a bacillus subtilis new strains.