Banana is one of the most important and remunerative cash crops grown round the year in Ethiopia. The energy and nutritional status of banana are much higher than other common tropical and subtropical fruits. The objective of this study was to establish in vitro root development protocol in Poyo and Grand naine varieties from sucker. For root induction suckers of Poyo and Grand naine were cultured on MS basal medium that contained 30 g/l sucrose, 6 g/l agar and vitamins. Rooting of the shoots was done on full strength MS basal medium supplemented with different concentrations of IBA (0.5, 1.0, 1.5, 2.0 mg/l) and NAA in sole and in combination (0.5 mg/l IBA+ 0.5 mg/l NAA and 1.0 mg/l IBA + 1.0 mg/l NAA). In all cases, MS basal medium without PGRs was used as control. The rooted plantlets were then acclimatized under greenhouse conditions by transplanting the plantlets on moist red soil, sand soil and compost or their mixture in 2:1:1 ratio. Results showed that root formation responses were significantly (P<0.05) higher under all levels and combinations of hormones when compared with control treatments. For Poyo variety, 0.5mg/l IBA had higher response for root numbers and root length than at other treatments. Number of days to root emergence was found to be shorter at 20.mg/l NAA of hormonal level than other treatments. In the case of Grand naine variety, the concentration of 1.5mg/l IBA alone treatment had positive effects on multiple root formation and MS + 0.5 mg/l IBA for highest length of roots. The number of days for root emergence was found to be shorter at combination of 1.0 + 1.0mg/l IBA and NAA in grand naine. Compared to the control, both IBA and NAA alone or in combination showed significant positive effect on root formation of shoots of both varieties. The acclimatization experiment showed that plantlets of both varieties survived better on the sterilized mixture of moist red soil, sand and compost soil in a ratio of 2:1:1. Comparison of the performance of both varieties showed that Poyo showed superior performance compared to Grand naine variety with respect to all measured agronomic traits except field survival rate which found better in grand naine, suggesting that varietal difference for the difference in the response to the PGRs.

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