A Protocol for High Frequency Regeneration Through Nodal Explant Cultures and ex vitro Rooting of Plumbago rosea L.

Abstract

A rapid clonal multiplication scheme comprising direct multiple shoot initiation and downsizing of the node with buds proliferated upon during subculture was developed for Plumbago rosea. Sixty five per cent of the nodes (approximately 2.0 cm) dissected out of young shoots from field grown plants and cultured in MS agar medium containing 3% sucrose and 15.4 microM BAP remained contamination free and responded at 95% rate with callusing at basal cut end and axillary bud break in 5 days followed by the formation of 2.41 +/- 0.14 shoots of 0.87 +/- 0.14 cm length in 3 weeks. Though differences in frequency and number of buds formed between nodes of 1-5 positions from the young shoots was negligible, the shoots emanated from the youngest node were shorter (0.92 +/- 0.19 cm) than those (2.3 +/- 0.50 cm) of the mature 5th node. Synergistic influence of BAP and auxins on caulogenesis was absent. Bud emergence in shorter (approximately 0.5 cm) nodes was delayed up to 3 weeks and extensive callus proliferation from the cut basal end overlapped the 8.2 +/- 0.37 axillary shoots/buds formed after 7 weeks. Reduction in the size (downsized) of the 2.0 cm node with buds to 1.0 cm by dissecting out the basal internodal segment having the callus and subculture of them (approximately 1.0 cm) with buds in contact with the medium for 3 weeks contributed to maximum multiplication of 42.1 +/- 5.40 shoot buds. Division of the shoot cluster and transfer of 2-3 shoots (0.5-1.5 cm) in a clump to MS basal liquid medium induced elongation of the shoots to 4.1 +/- 0.18 cm in 2 weeks. Shoots of 3.0-4.2 cm length were rooted within 3 weeks at 100% efficiency in vitro or ex vitro without hardening. In vitro rhizogenesis in presence of 0.49 microM IBA is recommended for enhanced rooting and high yield of commercially important tuberous roots during cultivation in the field.