Ceriporiopsis subvermispora, a white-rot fungus, is characterized as one of the best biopulping fungi because it can degrade lignin selectively without serious damage to cellulose. We previously demonstrated that during the early stage of wood decay, this fungus produces large amounts of linoleic acid (18:2n-6) and degrades lignin by manganese peroxidase-catalyzed lipid peroxidation. In this study, we cloned a Delta 12-fatty acid desaturase gene absolutely essential for the biosynthesis of linoleic acid as the main substrate for lipid peroxidation. This gene designated Cs-fad2 encodes a protein with three histidine-rich domains and four membrane-spanning domains characteristic of other Delta 12-fatty acid desaturases. Moreover, we heterologously expressed Cs-fad2 in Saccharomyces cerevisiae lacking Delta 12-fatty acid desaturase, and detected the de novo biosynthesis of linoleic acid by gas chromatography-mass spectrometry analysis. We also investigated transcription of Cs-fad2 under various conditions. The transcription was activated and repressed in the presence of a lignin fragment and exogenous fatty acids, respectively. These results may shed light on the molecular relationship between fatty acid metabolism and selective lignin degradation in C. subvermispora.