This study demonstrates a protocol for in vitro regeneration of Gerbera jamesonii cv. ‘Sunglow’ developed by culturing leaf segments on Murashige and Skoog (MS)medium supplemented with 1.0, 1.5, 2.0, 2.5 and 3.0 mg L-1 indole-3-butyric acid (IBA), naphthalene acetic acid (NAA), 2,4-dichlorophenoxy acid (2,4-D) and 1.0, 2.0, 3.0, 4.0 and 5.0 mg L-1 benzyl adenine (BAP) and kinetin (Kin). The treatment, 1.5 mg l-1 2,4-D gave 100% callus induction with a friable, nodular and creamish white callus. BAP gave fair callus growth with compact and brownish callus. However, Kin failed to produce callus. Shoot regeneration was assayed with 1.0, 2.0, 3.0, 4.0 and 5.0 mg L-1BAP and Kin. BAP at 3.0 mg l-1 showed 76.67% shoot regeneration, 4 shoots per explant and shoot length of 9 cm, whereas same concentration of Kin gave only 16.67% regenerated shoots, 1.97 shoots per explant and length of 5.167 cm. In vitroroot induction was determined by using 0.5, 1.0, 1.5 and 2.0 mg L-1 indole-3-acetic acid (IAA) and NAA. IAA at 1.5 mg L-1 exhibited a rooting percentage of 97.67%, whereas similar concentration of NAA gave relatively less rooting percentage of 60.67%. IAA gave thick roots with maximum root number (7.567/per explant) and length (7.33 cm), on the contrary, NAA gave relatively thin roots having less number of 3.567 roots with a length of 4.667 cm. Key words: Gerbera jamesonii, in vitro regeneration, leaf segments, culture.