Yarrowia lipolyticais a well-known oleaginous yeast that naturally accumulates lipids to more than 20% of their dry cell weight. Due to its brief doubling time and Generally Recognized as Safe (GRAS) properties,Y. lipolyticahas been exploited for the production of commercially valuable lipids. Among the genes related to the lipid synthesis, the gene YALI0E16797g (LRO1) encoding a major triacylglycerol synthase ofY. lipolyticashows a significant impact during the acylation process. Thus, in the present work, we explore the contributions of hp4d or TEFintron promoters to the response of LRO1 expression on lipid accumulation by molecular cloning technology. Results showed that over-expression of LRO1 led to higher lipid content as well as lipid yield. The one with the hp4d promoter showed the highest lipid content of 12% wt. However, such an enhancement also caused a growth defect of cells. On the other hand, the lipid content of the cells over-expressing LRO1 with TEFintron promoter revealed only a modest increase in lipid content, but it promoted cell growth. Therefore, all things considered the one with the TEFintron promoter showed the highest lipid yield.