Construction Of Fat-1 Transgenic Mice

Polyunsaturated fatty acids(PUFAs)are present in all groups of organisms and play a key role in the maintenance of the proper structure and functioning of biological membanes. The amount ofω-3 PUFAs is insufficient in most people, whereas the level ofω-6 PUFAs is relatively too high, the balance of these two classes of PUFAs is important for homeostasis and normal development. Besidesω-3 PUFAs have shown to exert many preventive and therapeutic actions besides their important role in maintenances human health and normal development. However most animals have to obtainω-3 PUFAs from diet, because they can not synthesize them by themselves. The protein encoded by Fat-1 gene isω-3 fatty acid desaturase, which is an important fatty acid desaturase in the biosynthesis of PUFAs.This enzyme can catalyze the conversion ofω-6 PUFAs toω-3 PUFAs. In this study , based on the published seqence of the C.elegans Fat-1 gene in the Genebank and the regulation of the codons usage of the bovine milk protein with the bias, we optimized the codons of Fat-1 gene and synthesized the optimized gene, and inserted the restriction enzyme digestion sites XhoI to the 5'and 3'ends of sequende. After the whole modified sequence was synthesized by the Shanghai Sangon Biological Engineering Technology and Service Co,Ltd. Then we reconstruct the gene vector pBC1-Fat-1 based on the pBC1. After that we transferred this Fat-1 gene into mice by the method of microinjection. Then we proved that foreign genes has been integated within the mice genome by the PCR and Southern blot.Meanwhile we analysis theω-3/ω-6 PUFAs of the transgenic mice milk, the result demonstrated clearly thatω-6 PUFAs in milk of the transgenic mice are reduced,indicating thatω-6 PUFAs have been converted toω-3 PUFAs,causing the ratio ofω-3/ω-6 PUFAs to increased.