Adventitious shoots

Present study was carried out to standardize a protocol for high efficiency in vitro adventitious shoot regeneration from Murraya koenigii using cotyledon and hypocotyl explants. Adventitious regeneration, which is a pre-requisite in most genetic transformation studies using Agrobacterium and ballistics, needs to be developed as a protocol for micropropagation of M. koenigii. Direct adventitious shoot proliferation was achieved from intact seedling on Murashige and Skoog (MS) medium supplemented with various concentrations of 6-benzyleaminopurine (BAP) 2.64 µM to 22.21 µM, Kinetin 2.34 to 13.96 µM and Adenine sulphate (ADS) 40.72 to 244.39 μM to induce in vitro multiple shoots. Percentage response of cotyledon explants was 95.00 ± 0.58 which was significantly higher than the response of hypocotyl explants (76.2 ± 0.06) explant in the MS basal medium supplemented with 12.95 µM BAP, 8.98 µM Kinetin and 152.74 µM ADS. The 35-40 mm elongated shoots were cultured to MS basal medium augmented with different concentrations of indole-3-butyric acid (IBA). The maximum percentage, 84.8 ± 0.13 of rooting was achieved on MS basal medium containing 17.26 µM IBA. In-vitro plantlets regenerated from cotyledon and hypocotyl explants were hardened for four weeks in a green house. The hardened plantlets were transferred to field conditions. Eighty percent hardened plantlets were successfully survived under natural conditions

Rapid shoot proliferation was established by adventitious shoot formation from inter-node segments. Shoots were directly regenerated in vitro without callusing. The concentration of plant growth regulators (PGRs) in MS medium exhibited a discrete role in the efficacy of adventitious shoot induction. Maximum number of shoot (82.02 ± 0.08) regeneration was achieved on modified MS medium supplemented with BAP 13.01 µM, Kinetin 8.87 µM and ADS 136.52 µM. Rooting of in vitro shoots occurred in three to four weeks on transfer to MS medium containing IBA (19.68 µM). About 88 % of in vitro-raised plantlets were survived under field conditions. The protocol development in present study for rapid adventitious shoot regeneration from inter-node segments of Murraya koenigii could be helpful in carrying out various genetic modifications in this economically important medicinal plant