Breeding Of The Yeast With High Ethanol Yield From Xylose-Fermentation

Bio-ethanol is one of the most promising renewable and clean energies. The production of ethanol from lignocelluloses is a good way to solve the energy crisis and environment pollution. For the second abondant of sugar of lignocellulose hydrolysate can not be converted to ethanol efficiently and thermotolerance of strains is weak, this paper aims at breeding a thermotolerant yeast with high ethanol yield in the fermentation of xylose.In this paper, comparisons about stress tolerance(ethanol-tolerance, thermotolerance and osmotic-tolerance), sugar fermentation ability(glucose, xylose, the mixture of glucose and xylose), high temperature fermentation ability (with xylose as carbon source)were conducted between Spathaspora passalidarum NRRL Y-27907 and Pichia stipitis NRRL Y-7124. The results show that NRRL Y-27907 performs better than NRRL Y-7124 in stress tolerant experiments. NRRL Y-27907 has lower glucose fermenting ability, but higher xylose-fermenting performance than NRRL Y-7124. Glucose repression effect has less influence on NRRL Y-27907 than NRRL Y-7124, which leads to much better fermenting performance of NRRL Y-27907 than NRRL Y-7124 with mixed sugar of glucose and xylose. According to the results of fermentation at different temperatures, NRRL Y-27907 performs better in high temperature fermentation. So Spathaspora passalidarum NRRL Y-27907 was selected as the starting strain.UV mutagenesis and enrichment at high temperature were conducted on the strain U, 200 strains were collected. After tube screening, bioscreen screening and shake flask screening, a mutant U-30 was obtained. Ethanol yield of mutant U-30 was 13.62 g/L at 37℃, which was 13.03% higher than U.Then the protoplast fusion between U-30 and Saccharomyces cerevisiae F3 (preserved in our lab) was proceeded by using single affectionate inactivated(U-30) protoplast fusion technology, we use xylose as sole carbon to screen fusants. After screening, fusant UF44 produced 20.78% more ethanol than U-30, and the productivity is 16.45 g/L at 37℃. The genetic characters of UF44 is stable.