Research On Screening, Mutation Of High-yielding Butanol Strains And Fermentation Technology

Acetone butanol fermentation is one of the ancient fermentation industry, long before and after first war has started to common application in different countries. Butanol is the foundation of the bulk of chemical raw materials, with increasing of oil resources in short supply, people began to move to the eye of renewable resource use up, Butanol is expected to become a new generation of liquid biofuels。The decisive role in butanol fermentation is the quality of the strains,so breeding genetics stability, high yield strains are the key of success or failure for butanol fermentation. This study from jilin agricultural university soybean in soil a test-bed for screening plant high yield butanol strains, named for the X3. Bought from the Chinese academy of sciences in microbial ordinary microorganism strains of center management preservation acetone butanol fungi8011controlled plant, In order to obtain genetic traits were stable, butanol high output strains respectively, the strains X3and8011ultraviolet mutagenesis and microwave induced.Finally, using the response surface method to the Mutations8W3-4of fermentation conditions of fermentation and the optimization, make the output of strains is distinctly improved. Specific research results are as follows:1. Use of corn mash enrichment of the training of the method, From jilin agricultural university test-bed for soybean were studied in10strains of soil with corn mash can produce for fermentation strains of butanol, through the fermentation tests further after screen, by gas chromatograph method yields, get the highest yield butanol strains, named for the X3.With7%of the corn of the fermented mash, s chromatography:butanol’s production for the1.41g/L. The determination of the growth curve, and the results show that in the12h at the end of the logarithm growth. Dilute applied on a flat plate, colony morphology for:round, smooth surface, low convex, not transparent. On16SrDNA strains of sequence analysis, constructing the system growth trees, and the results show that strains the X3and bacillus subtilis strain have high similarity and the similarity of99%above, from which we can identified the bacteria for bacillus subtilis.2. To obtain high yield of butanol strain, and8011strains of X3separately carried on the uv mutagenesis, microwave mutagenesis, uv and microwave alternate mutation. Ultraviolet mutation results show that:the X3best dilution gradient strains for10-5, best mutagenesis time for60s; The best concentration gradient strain801110-4, best mutagenesis time for80s. Microwave mutation results show that:the X3best dilution gradient strains for10-5, best mutagenesis time for80s; The best concentration gradient strain801110-5, the best time for160s mutants. After three rounds of ultraviolet microwave alternate after mutagenesis, yield butanol mutations XW3-4and8W3-4, yield of4.74g/L and10.34g/L. Mutants strains of genetic stability test, five rounds ChuanDai test shows that the genetic mutations have good stability.3. Single factor experiment for Mutations8W3-4, with the response surface method are built up for fermentation composition, fermentation conditions and butanol output regression model. Investigation fermentation composition and the conditions of fermentation production to butanol influence. Through the experiment the best medium components is:(NH4)2SO40.203g/L,CaCO30.12g/L,MgSO4·7H2O0.02g/L.The best technological conditions combination is:fermentation temperature37.05℃, Training pH value5.13Inoculation amount of10.29%,Under the optimal conditions in combination, the butanol production is15.15g/L, The fermentation cultivation condition increased46.5%than the initial production butanol quantity (10.34g/L).