Adenine sulphate

Murraya koenigii (L.) Spreng, commonly known locally as “curry patta” or “mitha neem” in India, is a valuable medicinal plant known for its biochemical and aromatic properties. This paper presents a protocol for the rapid and high frequency shoot regeneration from nodal and inter-node explants from matured plant and epicotyl, cotyledons, cotyledonary node (embryonic axis), juvenile leaf, hypocotyl and root segment of in vitro derived seedling via axillary and adventitious shoot formation of M. koenigii. 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. Cytokinins benzylaminopurine (BAP), Kinetin (Kin), adenine sulphate (ADS) and indole-3-acetic acid (IAA) were used for multiple shoot induction. Addition of an auxin along with cytokinin improved the shoot production capacity. Shoot buds could be initiated from all the explants tested, with epicotyl explants producing the highest average number of shoots/explant. N6-benzyle adenine (BA), kinetin, adenine sulphate (ADS) and indole-3-acetic acid (IAA) in combination were the most effective PGRs for shoot induction.

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