Effect of cytokinins and lines on plant regeneration from long-term callus and suspension cultures of Allium cepa L.

Abstract

Suspension cultures were initiated from callus cultures of Allium cepa, which had been precultured on a solidified medium for 7 months. For another 3 months the 83 callus lines were kept in suspension culture. Each line is derived from a single zygotic embryo, 24 lines from onion and 59 lines from shallot. Of these, 20 suspension lines showed adequate growth and were used to test the effect of cytokinins on plant regeneration. On average, 1.25% of the calli produced shoots on a growth regulator-free medium (MS30: MS basal medium supplemented with 30 g/l sucrose). Thus plant regeneration decreased significantly with time because the overall average plant regeneration frequency was 35.5% after 3 months of culture on solidified medium. Plant regeneration after 3 months proved not to be a reliable predictor of plant regeneration after 10 months in vitro culture. Overall only 0.25–0.88% of the calli produced shoots when the regeneration medium was supplemented with different types and concentrations of cytokinins. Contrary to expectations, the type and concentration of cytokinins could not increase the shoot regeneration capacity. However as expected, the increasing cytokinin concentration (especially TDZ at the range of 1–5 mg/l) led to a decrease in root formation. Plant regeneration proved to be highly dependent upon the line used. The best line was atm24 with an overall shoot regeneration capacity of 4.62% (among the 13 treatments); it had its highest shoot regeneration on MS30 treatment with 12.5%. The results obtained show that for the development of a reliable transformation protocol only young callus material (< 3 months), which has still a high regeneration potential, can be used.