Selection And Mutation Of Saccharomyces Cerevisiae Strains With High-yield CLA, Purification Of Its Linoleate Isomerase

Conjugated linoleic acid (CLA), is a kind of unsaturated fatty acid. The CLA gained by food not only does it have the function of anti-cancer and anti-atherogenic, but also it helps to lower cholesterol, promote bone growth and so on. Recently, the CLA synthesized by industrial method always have numerous isomers, in addition, there are still some side effects during the process of it. So, transporting CLA through micro-organisms has become the research focus because of its advantage, including single product and simple procedures.In order to obtain micro-organisms which can product high quantity of CLA, which will lay foundation to further industrial production of high purity, high activity CLA used to add into the food. The paper selected from the yogurt strains of conjugated linoleic acid, after identified by 18SrDNA. linoleic acid isomerase were isolated and purified by utilizing salting, Sephadex G-100, DEAE-cellulose ion exchange chromatography, then use the enzyme activity and SDS-PAGE to further identification. Finally, after the ion implantation of the selected strains and optimization of fermentation conditions.In conclusion :(1)Selected one strain which can product high quantity of CLA from the yogurt, finally it was proved to be the wine yeast Saccharomyces cerevisiae after the identification of 18S rDNA sequence, and the conversion rate of CLA is 3.35%;(2)After 50 to 80% ammonium sulfate fractionation, Sephadex G-100 gel filtration chromatography, DEAE-cellulose ion exchange chromatography purification, examination of enzyme activity and determination of SDS-PAGE molecular weight, we concluded that we gained purified acid isomerase, and its molecular weight is about 68.5kDa.(3)Employ the selected wine yeast strain as the starting materials, after the ion beam mutagenesis, screening, positive and negative mutant strains, then subculture them in order to gain a CLA mutant with high genetic stability.(4)The optimal fermentation conditions for mutant strain is:42h for incubation time, pH 5.0 for starting pH value, 4% inoculation, and 40℃for incubation temperature. Then further add 0.1% linoleic acid, in this case, the production of CLA is 0.0898mg/mL, and its transporting efficiency was 21%.