Co-expression Of Elongase And Desaturase In Transgenetic Yeast

The DHA plays an important role in keeping the healthy of people, and clinical studies have shown that DHA has many positive effects on diseases such as hypertension, depression and cancers. so the demand for DHA as a dietary additive has been increased.In order to produce DHA via transgenetic technique, the recombinant plasmid pYTFD5-FAD4, including genes of delta-5 Elongase (FTD5), delta-4-Desaturase (FAD4), was constructed and transformed into Saccharomyces cerevisiae INVScl to construct recombinant strain. The genes were induced by the addition of galactose to 2% (w/v).The yeast medium was supplemented with 0.3mmol/L exogenous fatty acid subtrate Eicosapentaenoicacid (EPA, C20:5n-3) in the presence of 1% NP-40. The total fatty acids were extracted from the induced cells and subjected to methyl-esterification. The fatty acid methyl esters (FAME) were analyzed by GC-MS detection. Two novel peaks corresponding to Docosapentaenoic acid (DPA,22:5n-3) and Docosahexaenoic acid (DHA, C22:6n-3) methyl ester standards were detected with the same retention time which were absent in the control. The transformation efficiency of EPA to DHA is 12.47%. The results revealed that delta-5-Elongase and delta-4-Desaturase could specifically catalyze EPA into DPA and DHA, and DHA can be produced by transgenetic technique in yeast.To improve the transformation efficiency, the recombinant plasmid pYTFD5-FAD4-B5, including genes of delta-5 Elongase (FTD5), delta-4-Desaturase (FAD4) and cyb5, was constructed and transformed into Saccharomyces cerevisiae INVScl to construct recombinant strain. The genes were induced in the same method, but the transformation efficiency of DPA to DHA is not significant increase. The results revealed that promoter is the most importantt factor to affect transformation efficiency.