Vanillin production in metabolically engineered Beta vulgaris hairy roots through heterologous expression of Pseudomonas fluorescens HCHL gene

Vanillin is a naturally occurring phenolic compound, which has broad spectrum importance in food, cosmetics, flavouring agent and bioprservative. Aditionally, it also imparts many therapeutic benfits in the form of antimicrobial, antimutagenic and anticarcinogenic agent. Present study demonstrated the production of vanillin from two transgenic hairy root (HR) lines of Beta vulgaris bearing HCHL gene from Pseudomonas fluorescens, encoding p-hydroxycinnamoyl-CoA hydratase/lyase under the control of CaMV 35S promoter. These HCHL expressing HR clones viz. Bv 42 and Bv 18, showed conversion of inherently available phenypropanoid precursor (ferulic acid) into vanillin. Optimization of central parameters, such as media formulation, concentration of sucrose (1-5%), pH of the media (17), culture duration in combination with accessibility to higher amount of precursor by feeding with ferulic acid (2-10 mM), was carried out in detail, which have noticeably influenced the growth as well as vanillin synthesis potential of the roots. The HR clone Bv 42 showed maximum growth (GI = 1426 ± 22.71) in MS medium with 3% sucrose at pH 5.8 within 45 days, which was two times and five times higher than that with pH 7 and pH 3 respectively. Evidently, although the maximum synthesis of vanillin (0.052% ± 0.00013) could be obtained at pH 3 without any additional precursor feeding, its overall yield was better at 5.8 pH, which may be due to retardation in biomass yield as observed at pH 3. Conspicuously, addition of 10 mM of ferulic acid as the precursor enhanced the vanillin yield upto four fold (0.209% ± 0.0021), relative to that in the cultures devoid of any added precursor. The presently established transgenic HR mediated vanillin production technology adds much significance in realising the industrial goal of meeting the global demand of this biomolecule.