Development of high efficiency micropropagation protocol of an adult tree-wrightia tomentosa

Abstract

Highly efficient and reproducible micropropagation protocol for Wrightia tomentosa using sexually adult material has been developed. Multiple shoots were induced from nodal shoot segments through forced axillary branching in vitro. Nature and management of the donor tree, season of collecting explant and their orientation on the medium strongly influenced the initial establishment of cultures. Explants collected in April-June period and placed vertically on the MS medium containing 2 mgl-1 BAP produced shoots from axillary nodes in vitro. Management of donor tree by serial harvesting of explants every fortnight was necessary to obtain vigorous growth of shoots in vitro. Explants of the fifth flush (F5) were found most suitable to obtain more than 7 shoots per node on the above medium. The rate of multiplication in subsequent subcultures was a little more than two and half-fold. Incorporation of phloroglucinol (100 mgl-1) into the multiplication medium containing BAP (2 mgl-1) accelerated the rate of multiplication to 3-fold per subculture. Similar response could be obtained by using 10 mM thidiazuron (TDZ) alone in the multiplication medium. Nodal segments from in vitro raised shoots were also used to initiate a new culture cycle. The shoots could be multiplied for at least 24 months without loss of vigor. More than sixty per cent shoots obtained after sixth subculture developed roots when treated with pre-autoclaved indole-3-butyric acid solution (100 mgl-1) for 10 min and implanted on modified MS medium (major salts reduced to 1/4 strength and 400 mgl -1 activated charcoal). Successfully rooted plants were hardened in vitro in glass bottles containing Soilrite™ irrigated with 1/4 strength MS salt solution (pH 5.0). More than 5,000 plantlets were successfully hardened in vitro and transferred to greenhouse for acclimatization. The survival rate of the plants during hardening was more than 95 per cent. © 2005 Springer Science + Business Media, Inc. All rights reserved.