Study on xylose-fermentation capacity of recombinant Saccharomyces cerevisiae.

In order to establish the xylose-fermentation pathway in Saccharomyces cerevisiae,two key genes for xylose-metabolizing,xylose reductase xyl 1 and xylitol dehydrogenase xyl 2,were amplified from Pichia stipitis by polymerase chain reaction(PCR)technique and cloned into YEp 24,a high-copy expression vector in yeast,respectively and simultaneously,and then they were constructed into S.cerevisiae by transformation using lithium acetate.In three recombinant strains,the recombinant S.cerevisiae NLR04 could grow and ferment in media with xylose as sole carbon source while the two genes xyl 1 and xyl 2 were transferred simultaneously into an auxotrophic mutant of S.cerevisiae,and it showed an ineffective capacity of fermenting xylose to ethanol under trace oxygen or anaerobic conditions and its ethanol yield could reach 37.0 % of the theoretical value.These recombinant yeast strains enriched the strains library for the xylose-metabolic engineering.Detective results confirm that there was some difference between P.stipitis and the recombinant S.cerevisiae NLR04 in the gene xyl 1 and the gene xyl 2 expression mechanism,the gene xyl 1 is constitutely expressed in the latter but xylose-inducible in the former.However,the gene xyl 2 is reversed as it is constitutely expressed in the former but xylose-inducible to some extent.On the xylitol-dehydrogenase relative enzyme activity,the recombinant S.cerevisiae NLR04 is markedly lower than P.stiptis,which maybe the one of bottlenecks for xylose-utilization in recombinant strains.