Enhancing secondary metabolite biosynthesis from ethnomedicinal plant Solanum nigrum L. through cytogenetically stable mass propagation, transgenic hairy root induction, and using different LED light and culture vessels for hairy root culture

Abstract

This study aimed for stepwise enhancement of bioactive compound production in ethnomedicinal plant Solanum nigrum through novel approaches of micropropagation, hairy root induction, and treatment of hairy roots with various physical conditions. Polyamines were first-time used for in vitro shoot multiplication of S. nigrum, where highest shoot numbers (55.70 ± 0.64) were observed in Murashige and Skoog’s (MS) medium with 0.2 mg/L thiadiazuron + 20 mg/L spermine, whereas maximum root numbers (34.20 ± 0.80) were observed in ½MS medium with 0.5 mg/L indole-3-acetic acid. Pioneer studies of cytogenetical fidelity assessment through inter simple sequence repeat (ISSR) and start codon targeted (SCoT) markers, and chromosome analysis approved genetic uniformity in regenerated plants. Cent percent hairy roots were induced from in vitro S. nigrum leaves (25–30 roots/explant) using Agrobacterium rhizogenes (A4 strain). PCR of transgenes confirmed the transformed nature and absence of bacterial contamination in hairy roots. HPLC studies indicated greater secondary metabolites in hairy roots than in vivo and in vitro plants. Hairy root culture with different culture vessels revealed overall better biomass gain in tightly sealed vessels. However, hairy root cultures treated with red light showed highest biomass accumulation in fresh (2380.13 ± 16.83 mg) and dry (174.66 ± 4.68 mg) weight with maximum solasodine (3.25 ± 0.04 mg/g) and diosgenin (1.03 ± 0.02 mg/g) contents, whereas optimal production of caffeic (3.51 ± 0.05 mg/g), coumaric (2.53 ± 0.06 mg/g), ellagic (0.18 ± 0.02 mg/g), and ferulic (2.17 ± 0.03 mg/g) acid production was found in blue light. These reproducible protocols can be utilized in future bioreactor-mediated mass-culture of hairy roots for commercial-level secondary metabolite production from S. nigrum.