Solanum dulcamara L. (bittersweet) is a medicinal plant that has been used to treat skin diseases, warts, tumors, felons, arthritis, rheumatism, bronchial congestion, heart ailments, ulcerative colitis, eye inflammations, jaundice and pneumonia. A reliable in vitro culture protocol for bittersweet was established. Explants (leaf and petiole segments) were cultured on Murashige and Skoog minimal organics (MSMO) medium with various plant growth regulator combinations. Leaf explants formed more shoots than petiole explants. Plant regeneration was observed through indirect organogenesis with both explants. Best shoot proliferation was obtained from leaf explants with 3 mg/l BA (benzyladenine) and 0.5 mg/l IAA (indole-3-acetic acid). Regenerated shoots were transferred to rooting media containing different levels of IAA (indole-3-acetic acid), IBA (indole-3-butyric acid), NAA (naphthalene acetic acid) or 2,4-D (2,4 dichlorophenoxyacetic acid). Most shoots developed roots on medium with 0.5 mg/l IBA. Rooted explants were transferred to vermiculate in Magenta containers for acclimatization and after 2 weeks, they were planted in plastic pots containing potting soil and maintained in the plant growth room.