The effect of different levels of Benzyl adenine (BA) and Naphthalene acetic acid (NAA) on in vitro regeneration of Spunta and Mondial potato cultivars were studied, in addition, the effect of different levels of colchicine administered either as part of MS media (0.0, 10 μ M, 100 μ M and 1 mM) or by soaking potato buds (0.0, 1, 2 and 3 mM) on in vitro regeneration of two potato cultivars was studied in the Faculty of Agriculture and Veterinary Medicine, An-Najah National University, Tulkarm, Palestine. Higher shoot number was obtained when BA at 1 and 2 ppm were combined with 0.5 ppm NAA or without NAA, with variation between the two cultivars. Root number was significantly affected with the hormone levels, higher root number per bud in Spunta cultivar was obtained when NAA was utilized exclusively at 1 and 0.5 ppm respectively, however with Mondial cultivar, the higher number was observed at 1 ppm NAA. Significant effect of colchicine on shoot multiplication in both cultivars was observed, higher average shoot number for both cultivars (3.25 and 3.07) was recorded when higher level of colchicine ( 1mM and 100 μ M) was added to the media for “Spunta’ and “Mondial” respectively. When potato buds were soaked in colchicine, no significant effect on regeneration ability of the two cultivars was observed. Our results indicated a positive effect of colchicine on in vitro regeneration of two potato cultivars, therefore, protocols of polypolidization of potato through in vitro culture could be implemented.