Expression And Effect Of Chlorella Vulgaris CvFad3 Gene In Human Breast Cancer Cell

Objective To investigate expression and effect of n-3 fatty acid desaturase gene CvFad3 of Chlorella vulgaris in breast cancer MCF-7 cells.Methods n-3 fatty acid desaturase gene CvFad3 of Chlorella vulgaris was cloned by RT-PCR from Chlorella vulgaris. The cDNA of the gene was inserted into eukaryotic expression vector pEGFP-C3 to form a recombinant, then transfected it into breast cancer cell MCF-7 by transfection reagent. The plasmid pEGFP-C3 was set as control group.The expression of CvFad3 gene was detected by RT-PCR; MTT was used to examine the prol-iferation of cells; flow cytometry was used to examine the cell cycle and cell apoptosis of MCF-7 cells which were transfected; and lipid analysis was used to detect the change of n-3 PUFAs in MCF-7 cell membrane.Results The proposed recombinant plasmid was got through DNA recombinant technique:CvFad3 gene expressed effectively in human breast cancer cells:CvFad3 gene mRNA band appeared after transfection of pEGFP-C3-n-3 48h. and CvFad3 gene mRNA band didn't appeare after transfection of pEGFP-C3. Comparing with, the control cell (pEGFP-C3-Control), MTT shew that proliferation of MCF-7 cells was markedly inhibited(21%, t=6.039,p<0.01); flow cytometry discovered that the cells of G0/G1 phase were decreased; moreover, lipid analysis shew that CvFad3 gene significantly increased the content of n-3 PUFAs in MCF-7 cell membrane, thus, CvFad3 gene decreased the ratio of n-6/n-3 PUFAs of MCF-7cell membrane.Conclusion Recombinant eukaryotic expression vector pEGFP-C3-n-3 is successfully constructed;n-3 fatty acid desaturase gene CvFad3 of Chlorella vulgaris can express in human breast cancer cells MCF-7 heterologously, meanwhile. inhibit proliferation of MCF-7 cells.and cell cycle was mainly arrested at G0/G1 phase, and CvFad3 gene also decreased the ratio of n-6/n-3 PUFAs of MCF-7cell membrane.