Screening Of Marine Saccharomyces Cerevisiae Strains And Genetic Recombination Of Saccharomyces Sp. W0

The world faces the progressive depletion of its energetic resources which mainly are based on non-renewable fuels, as well as the problem of environmental pollution. One remarkable solution is the use of energy in form of biomass (bioenergy). Conversion of the energy crops and lignocellulosic residues into biofuels is an important choice for the exploitation of alternative energy. Ethanol is an important biofuel either as a fuel or as a gasoline enhancer. Main feedstocks for ethanol production are starch and the other sugar raw materials. The final ethanol production is affected by various factors, for example, the ability of ethanol-tolerance. The higher concentration of ethanol coule damage the cell membrane and functional proteins, reduce cell viability, and gradually lead to cell death during the bioethanol production by S.cerevisiae.The ocean covers 71% of the earth surface and it is important for us to find various biomass to improve the industry production. Many countries have called for appropriate exploration of the ocean to find new fuels. We have collected more than 1000 yeasts from the seas of China in our laboratory, and we found there were only 17 strains of S. cerevisiae after identification. All of them were distributed in Bohai Sea,East China Sea and South China Sea except Yellow Sea. There is no reasonable explanation for the phenomenon now. The strains that could produce higher concentration of ethanol were Saccharomyces cerevisiae (2E00400,2E00498,2E00558,2E00723,2E00724), and all of them were derived from Bohai Sea, however, none of them could produce more ethanol than Saccharomyces sp. W0.Stress-tolerance is one of the most important influencing factors for industrial S. cerevisiae strains, so the breeding of yeast for stress-toleranc is important for fermentation. In my experiment, I noticed that although Saccharomyces sp. W0 could produce the most ethanol, it was not the best one to endure ethanol. Saccharomyces sp. W0 had only 39% survival after 2h's ethanol shock treatment in 18% (v/v) ethanol while 2E00498 had 51% under the same condition.In our previous studies, it was found that Sacchromycopsis fibuligera A11 could accumulate more than 25.0% (w/w) trehalose in its cells and secrete high level of acid protease and amylases, suggesting that the yeast has very high Tps1 activity.In this study, in order to further enhance ethanol tolerance of high ethanol producing yeast Saccharomyces sp. W0, we constructed a stably recombinant Saccharomyces sp. Z8, by integrating the TPS1 gene cloned from S. fibuligera A11 into the chromosomes of Saccharomyces sp. W0. The tansformant Z8 obtained accumulated more trehalose (about 30%), had higher ethanol tolerance (especially after 3 h's ethanol shock) and trehalose-6-phosphate synthase (TPS1) activity (about 33%) and produced more ethanol than Saccharomyces sp. W0 under the same condtions. This meant that trehalose did play an important role in ethanol tolerance and ethanol production by Saccharomyces sp. W0. The final ethanol production of Z8 strain was 36(g/g), while W0 had a 27(g/g) production. We determined the trehalose content during the fermentation every 24 hours, and the trehalose content in the strain Z8 was much higher than that in the strain W0.