Melon breeding programs can highly benefit from biotechnological tools, However, a breeding program associated with biotechnological tools depends upon the development of an efficient in vitro plant regeneration system. Organogenesis is used for new breeding purposes in biotechnology programs. The aim of the current study was to examine the various growth regulations and explants on direct organogenesis of Persian melon cultivar ‘Khatooni’ using induction of regeneration in cotyledon leaf explants. For this purpose, leaf cotyledons in vitro explant ( three-, five-, seven-, 10-, and 12-day-old seedlings) were cultured in Murashige and Skoog Medium (MS) supplement with 0, 0.5, and 1 mg.l-1 6-benzylaminopurine (BAP). Adventitious buds were identified by the development of the shoot meristem tip and the beginning of the leaf. The results showed that in vitro organogenesis in melon was the highest multiplication rate (5.3) and shoot regeneration (63.7%) when cotyledon leaves from seven-day-old seedlings were cultured on MS medium supplement with 1 mg.l-1 BAP. regenerated plantlets were transferred to MS medium supplement 0.1 mg.l-1 BAP and 1 mg.l-1 Gibelleric acid (GA3) to elongate shoots. Then, by transferring to MS medium supplement with 1 mg.l-1 Indole butyric acid (IBA), regenerated plantlets with developed roots were obtained. According to the results of this research, we can start propagating and producing in vitro melon plants faster and healthier. Using this method in gene transfer programs can increase the production efficiency of transgenic plants.