We developed an efficient protocol for in vitro shoot organogenesis and plant regeneration from Cymbalaria muralis leaf explant cultures. Leaf explants were cultured in media containing different concentrations of cytokinins. After determining the most effective cytokinin and cytokinin concentration, we tested the effects of various auxins at different concentrations on the regeneration efficiency and growth of shoots from excised leaf segments. Among the cytokinins tested, zeatin supplementation resulted in the highest shoot initiation efficiency, largest number of shoots per explant, and longest shoots after 6 weeks of culture. In this study, zeatin in combination with an auxin, naphthalene acetic acid, was highly effective for inducing shoot regeneration. Among the cytokinin/auxin combinations tested, supplementation of the culture medium with 1 mg/L zeatin and 0.1 mg/L naphthalene acetic acid resulted in the greatest number of shoots per explant (8.3), highest regeneration efficiency (93%), and longest shoot length (27 mm). Zeatin with 0.5 mg/L naphthalene acetic acid supplementation resulted in the second-highest values for each of these measures. The combined application of cytokinin and auxin might play a vital role in shoot organogenesis of C. muralis. [Thwe AA, Yeo SK, Chae SC, Park SU. In Vitro Shoot Organogenesis and Plant Regeneration of Cymbalaria muralis. Life Sci J 2012;9(4):878-881] (ISSN:1097-8135). http://www.lifesciencesite.com. 135