Abstract
Horseradish hairy root cultures are suitable plant tissue organs to study the glucosinolate–myrosinase–isothiocyanate system and also to produce the biologically active isothiocyanates and horseradish peroxidase, widely used in molecular biology. Fifty hairy root clones were isolated after Agrobacterium rhizogenes infection of surface sterilized Armoracia rusticana petioles and leaf blades, from which 21 were viable after antibiotic treatment. Biomass properties (e.g., dry weight %, daily growth index), glucosinolate content (analyzed by liquid chromatography-electronspray ionization-mass spectrometry (LC-ESI-MS/MS)), isothiocyanate and nitrile content (analyzed by gas chromatography-mass spectrometry (GC-MS)), myrosinase (on-gel detection) and horseradish peroxidase enzyme patterns (on-gel detection and spectrophotometry), and morphological features were examined with multi-variable statistical analysis. In addition to the several positive and negative correlations, the most outstanding phenomenon was many parameters of the hairy root clones showed dependence on the organ of origin. Among others, the daily growth index, sinigrin, glucobrassicin, 3-phenylpropionitrile, indole-3-acetonitrile and horseradish peroxidase values showed significantly higher levels in horseradish hairy root cultures initiated from leaf blades.
Highlights
Strains T37, 15837 and 8196 of A. rhizogenes resulted in clones that became unstable during antibiotic treatment
In concordance with the above, our results showed an ITC–nitrile shift as a likely consequence of the changes in the abundance of a myrosinase enzyme (MYR) isoenzyme [68]
The surface-sterilized leaf blades and petioles were inoculated with Agrobacterium rhizogenes A4, 15834 and 8196 strains, using a sterile needle
Summary
Horseradish is used today primarily as a condiment, has been known as a medicinal herb since antiquity [1,2,3]. For both utilizations, the pungent, lacrimatory compounds, the isothiocyanates (ITCs) are responsible. The isothiocyanates are the default hydrolytic breakdown products of the glucosinolates (GLS). Glucosinolates are N-hydroxy-sulfates with a highly variable side chain (R) and a sulfur-linked beta-d-glucopyranose (Figure 1). GLSs are odorless molecules found in vacuoles, while the myrosinase enzyme (MYR), which catalyzes the hydrolytic reaction, is stored in different compartments, typically in myrosin cells [2]
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