The Effect of TIBA and NPA on Shoot Regeneration of Cannabis sativa L. Epicotyl Explants

Abstract

Industrial hemp (Cannabis sativa L., family Cannabaceae) is a multi-purpose crop, used in the production of food, nutraceuticals, cosmetics and medicines. Therefore, development of new varieties with specific chemical profiles is necessary. In vitro culture methods could be complementary to conventional breeding and a useful tool for large-scale propagation. Strong apical dominance is considered as one of the factors contributing to the recalcitrance of industrial hemp in shoot proliferation. In this study, we tested the polar transport inhibitors N-1-naphtylphtalamic acid (NPA) and 2,3,5-triiodobenzoic acid (TIBA) to enhance shoot regeneration as the result of suppression of apical dominance and to develop in vitro protocols for Diana, Finola and Fedora 17 cultivars. Shoot tips derived from epicotyls were cultivated on Murashige and Skoog medium (MS) supplemented with meta-topolin (mT) and NPA, and also thidiazuron (TDZ) with a combination of TIBA and NPA. The results showed that the combination of TDZ with NPA (1–5 mg L−1) and TDZ with TIBA (0.5–2.5 mg L−1) increased the response of explants and the multiplication rate, but the effect was genotype-dependent and malformations were observed. To optimize the developed protocol, a two-step procedure with shortened time of exposure to inhibitors and reduced concentrations of them was applied. Shoots were rooted on media containing indole-3-butyric acid (IBA) and then successfully acclimatized. The obtained results will be useful in micropropagation of recalcitrant industrial hemp varieties.