Study On Secretion Expression Of Cellulase And One-step Fermentation With Corn-cob

The cellulose biomass widely exists in nature. Industrialized production of bioethanol using cellulose is a trend of the new energy field and it is also the effective way to produce glycerin by cellulose fermentation in food industry, medicine, light industry, and other fields. Further improving the utilization of cellulose is the current problem to be solved.In this study, an integrated secretion expression vector pHBM368was structured with induction of rDNA to deliver foreign cellulase gene into yeast Y33respectively. The exoglucanase gene (CBH, GeneBank:AY861348) from Chaetomium thermophilum, synthetic endoglucanasegene (EG, GeneBank:EU169241) and β-glucosidase gene (BG, GeneBank: AF163097) from Thailand red sandalwood (Dalbergia cochinchinensis)were connected separately to the pHBM368. After picking out the recombinants, which were restructured to the linear DNA for respectively transfering into S.cerevisiae Y33by chemical method.After comfirming the recombinants, galactose was added respectively into the fermentation medium for the induction of secretion expression of cellulase. In the condition of50℃and OD600of yeast biomass was2.0, enzyme activity of cellulase per milliliter, including exoglucanase, endoglucanase and β-glucosidase, were detected orderly by DNS as3.96U/mL,3.01U/mL and3.89U/mL.Fermentation experiments selected cheap corncob powder for natural cellulose source, which was joined into fermentation medium as the only carbon source after preliminary pretreatment. The experimental results proved that cellulose can be utilized effectively by cellulase, the one-step fermentation process was the combinative one of synchronization saccharification fermentation(SSF) and the mixed fermentation(MF) to get bioethanol and glycerine. After96hours anaerobic fermentation, the mixed bacteria module of1L bottle fermentation in the level of the ethanol everage concentration reached2.02g/L, glycerin everage concentration reached0.85g/L. In the same condition of magnifying3L bottle fermentation, ethanol reached2.13g/L, glycerin reached0.82g/L. The recombinants have the higher utilization of cellulose in contrast to original yeast.