Research Of Abe Fermentation From Lignocellulose Using Recombinant Clostridium Acetobutylicum

Clostridium acetobutylicum is a industrial strain that extensively used for production of biobutanol. Although this microorganism contains all the genes of cellulose-degrading enzymes, it cannot grow and ferment with cellulose as feedstocks.In this study, gene cphy3367was cloned from Clostridium phytofermentans genomic DNA by PCR. Recombinant E.coli-Clostridium acetobutylicum shuttle vector pSOS95-cphy3367was constructed and was successfully transformed into Escherichia coli. Results showed that the thiolase promoter initated the expression of gene cphy3367in E.coli by using real-time quantitative PCR and activity detection of recombinant enzyme. In conclusion, the thiolase promoter can be recognized by E.coli host cell.Recombinant plasmids were transformed into ATCC824by electrotransformation, thus recombinant strains ATCC824/pSOS95-cphy3367and ATCC824/pSOS95del were constructed. ABE (Acetone-Butanol-Ethanol) fermentation experiment with filter paper cellulose as the sole carbon source was carried out. Result:ATCC824could not utilizte cellulose as the sole carbon source; ATCC824/pSOS95del could produce ethanol. it means that the empty vector could affect the metabolic pathway in C.acetobutylicum and improve the ABE fermentation; ATCC824/pSOS95-cphy3367could yield acetone, ethanol and butanol with feedstock of cellulose, and the solvent production was higher than ATCC824/pSOS95del. Gene cphy3367enhanced the ability of fermentaion of C.acetobutylicum ATCC824with cellulose feedstocks.Furthermore, ABE fermentation experiment with hydrolyzate of sugarcane bagasse as the sole carbon source was conducted. Result:ATCC824could not ferment this materials to produce ethanol; ATCC824/pSOS95del could ferment and yield ethanol, and the yield of acetone and butanol was slightly lower than ATCC824; ATCC824/pSOS95-cphy3367could ferment and produce acetone, ethanol and butanol(0.78g/L,0.97g/L,1.09g/L, respectively), the yield of them and the total amount of solvent (2.84g/L) were higher than those of ATCC824and ATCC824/pSOS95del, these demonstrated that the recombinated strains with gene cphy3367have a enhanced ability of ABE fermentation with hydrolyzate of sugarcane bagasse as feedstock.