Comparative evaluation of different citrus cultivars for adventitious regeneration

Direct shoot regeneration (adventitious regeneration) with no intervening callus phase is preferred over indirect regeneration in tissue culture of plants owing to no/ low genetic variations. We report the development of a reproducible in vitro adventitious regeneration protocol for citrus cultivars; Kinnow, sweet lime, lemon and rough lemon. Inter nodal stem segments isolated from in vitro- grown citrus seedlings were cultured on MS medium fortified with varying BAP, NAA and kinetin concentrations. Data for days to shoot initiation, frequency of shoot induction and rooting frequency were recorded. Sweet lime exhibited the earliest shoot induction while; Kinnow explants showed delayed regeneration response. The MS + BAP 1mg/l proved the best medium for early shoot induction in all citrus cultivars. All four cultivars exhibited varied responses to different shoot induction media. Kinnow was the least responsive and sweet lime showed the highest regeneration response with 29.8% and 55.6% mean shoot induction frequencies, respectively. Single shoots were transferred to MS- based rooting medium fortified with 1, 1.5, 2 and 2.5 mg/l of NAA. Rooting frequency ranged from 7.0 to 73% in Kinnow and sweet lime, respectively. The best rooting medium for shoots of all citrus cultivars was MS+ 2mg/l NAA. Well-rooted plantlets were shifted to the potting mixture with 85% successful adaptation. LSD test at 5% significance level was used to calculate the statistical significance of recorded data. This optimized reproducible and efficient adventitious regeneration system has applications in citrus micropropagation, genetic transformation, protoplast fusion and germplasm conservation.