Effect of purified vermicast extract with plant growth regulators on indirect regeneration of Naregamia alata Wight & Arn from leaf explants and genetic fidelity analysis of regenerants
2022
Raja, H David | Srinivasan, P. | Tamilvanan, R.
An efficient indirect organogenesis protocol for Naregamia alata was standardized in the present investigation. Purified form of vermicast extract (VCE) was used in combination with various plant growth regulators for callus induction, shoot regeneration, and root induction. Leaf explants were inoculated on MS medium with various concentrations (1.0–2.5 mg L⁻¹) of 2, 4-D and NAA combined with 20%, 30% and 40% of purified VCE for callus induction. Among the various concentrations tested, 2.0 mg L⁻¹ 2, 4-D, and 20% VCE produced maximum callus induction (96.2%) with green nodular callus. Then, the calli were transferred to MS medium fortified with different concentrations (1.0–2.5 mg L⁻¹) of TDZ and BA in combination with purified VCE (20%, 30% and 40%). The highest percentage of adventitious shoot regeneration (100%) was observed on MS medium supplemented with 1.5 mg L⁻¹ TDZ and 30% purified VCE with respect to the number of shoots (20.7 ± 0.28 shoots/callus) and shoot length (11.4 ± 0.21 cm). The well-elongated shoots were rooted in MS medium with IBA and IAA (1.0–2.5 mg L⁻¹) with purified VCE (20%, 30% and 40%). The maximum percentage of root formation (99%), number of roots (9.6 roots /shoot) and root length (5.9 cm) were observed in 2.0 mg L⁻¹ IBA with 30% VCE combination. Healthy rooted plants were successfully acclimatized with a survival rate of 98% in field conditions. The genetic fidelity analysis of in vitro raised plants of N. alata was evaluated using RAPD and ISSR markers and they were found to be true genetic traits. The present work suggests the addition of purified VCE with the plant growth regulators would enhance the regeneration ability of N. alata from leaf explants resulting in regenerants with genetic stability.
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