Background Alkylresorcinols (ARs) are compounds belonging to the class of phenolic lipids. A rich source of ARs are cereal grains such as rye, wheat, triticale or barley. ARs found in plants are characterized by a variety of biological properties such as antimicrobial, antifungal and cytotoxic activity. Moreover, they are proven to have a positive influence on human health. Here, we aimed to find and characterize the gene with ARs synthase activity in the species Secale cereale. Methods and results Using BAC library screening, two BAC clones containing the gene candidate were isolated and sequenced. Bioinformatic analyses of the resulting contigs were used to examine the structure and other features of the gene, including promoter, intron, 3’UTR and 5’UTR. Mapping using the FISH procedure located the gene on the 4R chromosome. Comparative analysis showed that the gene is highly similar to sequences coding for type III polyketide synthase. The level of gene expression in various parts of the plant was investigated, and the biochemical function of the gene was confirmed by heterologous expression in yeast. Conclusions The conducted analyses contributed to a better understanding of the processes related to ARs synthesis. Although the research concerned the rye model, the knowledge gained may help in understanding the genetic basis of ARs biosynthesis in other species of the Poaceae family as well.