Influence of meta-topolin on in vitro organogenesis in Tecoma stans L., assessment of genetic fidelity and phytochemical profiling of wild and regenerated plants

Abstract

Comparative effect of meta-topolin and other cytokinins was assessed to develop an efficient and reliable regeneration protocol for Tecoma stans, using mature nodal explants. The morphogenic effect of benzyl adenine (BA), kinetin (Kin), meta- topolin (mT) and 2-iP (2-iso pentenyl adenine) at various concentrations (1.0–10 µM) was studied individually or in combination with auxins (IAA, IBA or NAA). Superior multiplication rates were achieved on MS medium supplemented with mT and NAA. Of the tested combinations, maximum shoot regeneration (95%), mean shoot number (19.6 ± 0.60) and length (5.26 ± 0.73 cm) was recorded on MS medium supplemented with 7.5 µM mT + 0.5 µM NAA after 8 weeks of incubation. Among the different auxins employed for in vitro root induction, 92.5% microshoots rooted on MS medium enriched with 1.0 µM IBA with 10.8 ± 0.20 mean root number and 5.62 ± 0.17 cm length after 4 weeks of incubation. The acclimatized plants grew well in green house with 90% survival rate. The gas chromatography–mass spectrometry (GC–MS) analysis of ethanol leaf extract of in vitro-raised plants yielded a higher number of compounds than control plant. The assessment of genetic fidelity among regenerants, using ISSR markers did not reveal any somaclonal variation. Therefore, the protocol developed appears to be simple and reliable for mass production of clones with higher diversity of secondary metabolites. Our studies based on interaction of mT with auxins for better shoot proliferation will undoubtedly be helpful in understanding many developmental processes in plant tissue culture. The regenerated plantlets survived well under field conditions and no somaclonal variation was detected. This study proved that in vitro raised plants are superior for the production of higher diversity of secondary metabolites than in vivo plant. Thus, our findings are helpful in generating an efficient micropropagation system for mass scale production of plants with higher diversity of bio-active molecules without losing their genetic stability.