Regulation of sesquiterpenoid metabolism in recombinant and elicited Valeriana officinalis hairy roots Author Ricigliano, Vincent Department of Biological Sciences, St. John’s University, Jamaica, NY 11439, United States Author Kumar, Santosh Author Kinison, Scott Author Brooks, Christopher Author Nybo, S. Eric Author Chappell, Joe Author Howarth, Dianella G. text Phytochemistry 2016 2016-05-31 125 43 53 http://dx.doi.org/10.1016/j.phytochem.2016.02.011 journal article 285686 10.1016/j.phytochem.2016.02.011 5613c391-71cd-4dac-b03b-1f026c15b6e2 1873-3700 10530233 2.1. Establishment and characterization of V. officinalis hairy roots V. officinalis hairy roots were generated using Agrobacterium rhizogenes -mediated transformation of in vitro grown leaf explants. This genetic manipulation relies on conjugative transfer and integration of A. rhizogenes root-inducing plasmid DNA into the host plant nuclear genome. V. officinalis seeds were aseptically germinated and hairy roots were initiated on in vitro leaf explants. Single root segments were removed from different explants to afford independent transgenic events, which were maintained on hormone-free solid medium. Isogenic lines were used to initiate shake flask cultures and generate clonal biomass for molecular and biochemical analyses ( Fig. S1 ). As a metabolic qualification of the hairy root cultures, chemical profiles of hairy roots transformed with A. rhizogenes harboring Ri plasmid pRi15834 were compared to those of roots from wild type , soil-grown V. officinalis plants. Root biomass was powdered in liquid nitrogen, the materials fully lysed by mixing with an equal volume of acetone, followed by the addition of an equal volume of water, then partitioned into hexane. Aliquots of the hexane extracts were analyzed directly by GC–MS, as well as dried under nitrogen prior to trimethylsilyl diazomethane derivatization for GC–MS ( Fig. 2 , Fig. S7 ). The chemical profile of extracts from roots of wild type plants exhibited qualitative and quantitative differences relative to the hairy root cultures. The pRi15834 lines tended to accumulate fewer constituents than the roots from soil grown plants, but the levels of β- caryophyllene ( 5 ) and valerenal ( 7 ) were elevated while valerenadiene ( 6 ) and valerenic acid ( 8 ) were comparable ( Fig. 2 ).