Transformation of Plasmid Recombinant Carrying Diacylglycerol Acyltransferase (DGA1) Gene in Rhodosporidium toruloides InaCC Y908

The increasing energy consumption and carbon emissions in Indonesia, which are still predominantly derived from fossil fuels, highlight the urgent need for the development of sustainable and environmentally friendly renewable energy sources. This study aimed to enhance lipid production through genetic engineering of the oleaginous yeast Rhodosporidium toruloides . The natural lipid yield of R. toruloides remains suboptimal, necessitating optimization through genetic modification. Genetic engineering was performed by integrating the gene encoding diacylglycerol acyltransferase ( DGA1 ), a key enzyme involved in triacylglycerol (TAG)-synthesizing gene, diacylglycerol acyltransferase (DGA1), using the pKLAC2 plasmid via electrophoration-based transformation. Successful integration was confirmed by colony PCR, which revealed a DNA fragment corresponding to the expected size of the DGA1 gene (554 bp). Fermentation experiments were conducted using recombinant R. toruloides and wild-type strains with glucose as the carbon source at concentrations of 30 g/L, 60 g/L, and 90 g/L. The results demonstrated that genetic recombination significantly increased lipid productivity in R. toruloides compared to the wild-type strain. Furthermore, variations in glucose concentration significantly affected lipid production in the recombinant yeast (p < 0.05), with an optimal glucose concentration of 30 g/L yielding the highest a lipid titer of 1,02 g/L and a lipid content of 58.58%. These findings indicate that recombinant R. toruloides has strong potential as an efficient host for TAG production using glucose as a carbon source through genetic engineering, thereby supporting the development of sustainable and environmentally friendly biofuels.