The aim of this work was to study the molecular breeding of oleaginous filamentous Mortierella alpina for high production of linoleic (LA) or oleic acid (OA). Heterologous expression of the Delta 12-desaturase (DS) gene derived from Coprinopsis cinerea in the Delta 6DS activity-defective mutant of M. alpina increased the LA production rate as to total fatty acid to 5 times that in the wild strain. By suppressing the endogenous Delta 6I gene expression by RNAi in the Delta 12DS activity-defective mutant of M. alpina, the OA accumulation rate as to total fatty acid reached 68.0%. The production of LA and OA in these transformants reached 1.44 and 2.76 g/L, respectively, on the 5th day. The Delta 6I transcriptional levels of the RNAi-treated strains were suppressed to 1/10th that in the parent strain. The amount of Delta 6II RNA in the Delta 6I RNAi-treated strain increased to 8 times that in the wild strain. (C) 2017 Elsevier Ltd. All rights reserved.