MicroRNAs (miRNAs) are non-coding single-stranded RNA molecules of about 19-25 nucleotides in length that regulate different biological processes, including lipid metabolism. In this study, we explored the effect of bta-miR-33a on lipid metabolism in bovine mammary epithelial cells (BMECs) of Chinese Holstein for the first time. For this purpose, the plasmids of bta-miR-33a mimic, bta-miR-33a inhibitor and bta-miR-33a negative control were constructed to overexpress or repress bta-miR-33a in BMECs. The effects of plasmid transfection were analysed by examining the mRNA and protein expression levels of ELOVL6 and the intracellular triglycerides. The results showed that bta-miR-33a directly inhibited the expression of ELOVL6 in BMECs; decreased the mRNA levels of ELOVL5 , HACD2 , CPT1A and MSMO1 ; and increased the mRNA level of ALOX15 . Sequence bta-miR-33a also increased the contents of triglycerides in the cells, presumably as a consequence of these gene expression changes. In summary, the results of the present study suggest that bta-miR-33a regulates lipid metabolism by targeting ELOVL6 , which might be a potential molecular marker of milk fat composition.