Researches Of The Strains For Ethanol Frementation Of Xylose

Xylose is a kind of pentose, which is the second most sugar in the hydrolysate of lignocellulose. How to ferment xylose to ethanol efficiently is the key factor which decides the feasibility of ethanol production by lignocellulose, and finding out a effective strain is the most important part for the process. In this experiment, two aspects were studied, one is about the natural xylose-using Pachysolen tannophilus and the other is about the construction of engineering Saccharomyces cerevisiae which express the xylA gene from E.coli.Pachysolen tannophilus from our laboratory was firstly separated and several strains were obtained. All of them were tested in the pure xylose fermentation medium and the best one was selected for the further study. The optimum conditions of fermentation in shake-flask were obtained from the aspects of nitrogen, carbon, time, dissolved oxygen (DO), vitamins and inoculums size, the ethanol production is increased from 3.96g/L to 15.77g/L,4-folds than the initial strain and 68.9%of the theoretical yield. The evolutionary culture was used to stabilize the fermentation property and the growth curve was obtained.Clone and expression of xylA gene from E.coli in the Saccharomyces cerevisiae were studied. E.coli genome DNA was set as the template, devising the primer according to the xylA in the GenBank, product of 1.4kb was obtained with the method of PCR. Plasmid of pMD18-T-xylA was constructed with T-A clone. After sequencing, the results were the same as 98%in the GenBank. The plasmids of pMD18-T-xylA and pYEs were digested with restriction enzyme EcoRâ… and Hindâ…¢separately, and the production was linked after purification. The positive strains with pYEs-xylA were obtained after screen.