OBJECTIVE To analyze berberine and sanguinarine biosynthetic capacities of both in vitro shoot and root cultures of Argemone mexicana and tissues from entire plants at different developmental stages. RESULTS Berberine and sanguinarine were equally distributed in roots and aerial tissues of developing plantlet whereas, in juvenile plants, sanguinarine was only detected in roots. This alkaloid distribution was consistent with that of biosynthetic transcripts in juvenile plants. However, lower transcript abundance in plantlets´ leaves suggests that alkaloids were mainly formed in roots and then mobilized to this tissue. In vitro root cultures maintained similar alkaloid profiles to those from intact seedlings and plantlets. However, in addition to berberine, rootless shoot cultures accumulated high levels of sanguinarine and biosynthetic transcripts. CONCLUSIONS In vitro shoot cultures of A. mexicana can synthesize sanguinarine in addition to berberine. This represent a convenient system for the production of both alkaloids.