Gene Cloning of Omega ۶ Fatty acid as an Effective Dietary Supplement in Cancer Prevention in Yeast host

Background: γ-Linolenic acid (GLA), is a prominent omega۶ polyunsaturated fatty acid that hasa structural role in lipid membranes ingredients. In humans, GLA is metabolized to produceprostaglandins and eicosanoids such as leukotrienes which have many health and medicinal rolesin cardiovascular disorders, cancers, inflammatory disorders, diabetes, and some other diseasesby regulating the levels of expression in various genes. Some microorganism such Mucor rouxiiis a typical oleaginous filamentous fungus has been widely used to investigate GLA production.The production of GLA in yeas is cost-effective than fungi as the present study focused onoverexpression of delta-۶ desaturase gene from M. rouxii to Pichia pastoris.Materials and Methods: Fungus RNA was extracted by RNX- PLUS Kit and cDNA wassynthetized. Purified products were ligated into pTZ۵۷R/T vector according to themanufacturer's instructions and transformed using the heat shock method into E. coli DH۵αcompetent cells prepared by chemical CaCl۲ method. After TA cloning gene was transformed toexpression vector pPICZC.Results: After transformation into E. coli DH۵α, positive clones were selected with directcolony PCR screening. Finally, pPICZC - delta-۶ desaturase plasmid was additionally confirmedby restriction enzyme digestion and sequencing. The GC assay of lipid combination was alsoevaluated and revealed ۴۶% lipid production with ۷۲.۳ mg GLA in recombinant.Conclusion: Recombinant yeast may provide an opportunity for the development of the methodfor industrial-scale GLA manufacturing.

کلیدواژه ها:

Omega ۶ ، Cancer ، Cloning ، yeast