The Breeding And Optimizatio Of L-histidine High Productive Strain

L-histidine is an alkalescency amino acid with an imidazole group. In order to realizing the industrialization production of L-histidine, In this paper, Screening of strains from nature and laboratories. The high-yield strains was screening from producing strains after physical induction, chemical induction and filtrating. Study on the optimization of fermentation conditions of the high-yield strains, in order to obtain high-yield strains which have stable transmissibility. The contents and results are as follows.1.Studied the method of determining L-histidine. Research and comparison qualitative determination of L-histidine with paper chromatography and rapid quantitative determination with colorimetric method.2.This experiment mainly screened L-histidine producing strains from Soil, medium-temperature starter, high temperature Daqu starter and the laboratory. During the primary screening, qualitative analysis and determination with paper chromatography and secondary screening with Pauly colorimetric method.Finally obtained a high-yield strain-Bacillus SubtilisY-7,as the original stains for next mutation breeding,which have stable transmissibility.3. Bacillus subtilis Y1 was used as original strain.With 90 units, the per unit injection amount of which is 2.5*1013/cm3of N+implantation and cooperatively treated by UV and LiCl, and then by DES and LiCl as mutating agent over again. Expecting endow the strains with 6-mercaptopurine resistance and acquire shikimic acid auxotrophy Mutant and deficient mutants of histidase. After screening and flask shaking fermentation chose the high-yielding mutant stain named Y-217 (6-MPR, transketolase’, histidase")whose yield of L-histidine acid is 2.57g/L, which is higher than the initial amount of 1.76 g/L.Through 6th passage culture and flask shaking fermentation experiments, The results show that the yield of L-histidine acid was not increase and proof the quite good genetic stability.4. Optimization of fermentation conditions of Y-217:By the single factor experiment and orthogonal experiments, we studied the factors that could impact on the yield of acid, response surface analysis was applied for further optimization.The optimum condition was as follows:soybean meal 14g/L, glucose 10 g/L, ammonium sulfate 1.5 g/L, corn flour 11.2 g/L, potassium dihydrogen phosphate 1.5 g/L, magnesium sulfate heptahydrate 1.8 g/L, calcium carbonate 5 g/L. After optimization, the production of acid was increased to 4.61 g/L.