Leaf explant response in in vitro culture of St. John’s wort (Hypericum perforatum L.)

Abstract

For centuries Hypericum perforatum has been used in natural medicine. In the last decades, it has also attracted the attention of pharmaceutical industry due to its promising anti-depressant properties. The important factor in pharmaceutical application of plant material is its stable content of active compounds. Such stability requires standardized conditions of growth, e.g. an in vitro culture. Our aim was to establish a medium allowing for an effective regeneration of shoots from the standardized leaf explants in in vitro conditions. Cultures of the leaf explants carried out in darkness, on Murashige and Skoog agar medium, supplemented with auxins (2,4-dichlorophenoxyacetic acid, 2-metoxy-3,6-dichlorobenzoic acid, α-naphtaleneacetic acid, indole-3-acetic acid) and cytokinins (kinetin, N⁶-(benzyl)adenine, thidiazuron) resulted in callus formation. The callus produced roots on media containing indole-3-acetic acid or α-naphtaleneacetic acid alone. On media supplemented with auxins and cytokinins, indirect shoot organogenesis was also observed. The most efficient shoot formation was observed with 2.85 μM of indole-3-acetic acid and 4.44 μM of benzyladenine. Regenerated shoots were rooted on Murashige and Skoog without plant growth regulators medium or on a medium supplemented with indole-3-acetic acid. From a single leaf explant (one fifth of the leaf) after a month of the culture, 35 regenerated shoots were obtained (allowing for the formation of about 180 vegetative shoots per leaf). Successful multiplication of shoots from a standardized explant makes it possible to obtain a great quantity of uniform plant material for biotechnological purposes.