Cassava is an important crop as it is the source of energy in the diet for the most of the tropical countries. Field gene bank is the only viable option for the maintenance of cassava germplasm. Cryopreservation offers the long-term conservation of vegetatively propagates crops without losing its viability through an effective in vitro regeneration system. In vitro regeneration response in YTP-1 and Sree Athulya genotypes of cassava was standardized through optimization of various factors influencing organogenesis. Among the different media tested, highest shoot induction frequencies of 81.4% and 77.8% were recorded in medium supplemented with 0.5 mg/l BAP + 0.2 mg/l NAA (CMSI 2) for YTP 1 and Sree Athulya respectively in the present study. 3 to 5 multiple shoots from a single shoot apex obtained from 70-80% of explants within 50-60 days. Highest regeneration frequencies of 56.6%and 46.6% were recorded in medium supplemented with 1 mg/l IAA (CRM 2) for YTP1 and Sree Athulya. As a result, an easy in vitro regeneration method that is genotype independent was created and may be effectively used to preserve cassava germplasm in the cryogen bank.