Development of an efficient regeneration and rapid clonal multiplication protocol for three different Morus species using dormant buds as explants

Abstract

SummaryAn efficient in vitro protocol using dormant buds as explants was standardised for three Morus (mulberry) species: M. alba, M. indica, and M. laevigata. Explants were collected from field-grown plants and cultured on 1.0_ Murashige and Skoog (MS) medium supplemented with different concentrations and combinations of phytohormones. Thidiazuron (TDZ) at 0.1 mg l-1 or benzylaminopurine (BAP) at 1.0 mg l-1, added separately, gave the best rates of shoot initiation and shoot induction, and required less time for bud sprouting in all three species. Basal MS medium resulted in the lowest rate of shoot induction and the longest duration for shoot initiation. Shoot multiplication was induced by a combination of 0.5 mg l-1 BAP, 0.5 mg l-1 kinetin (Kn), and 0.1 mg l-1 indole-3-acetic acid (IAA). Shoot proliferation and the elongation of adventitious shoots were observed using 1.0 mg l-1 BAP plus 0.2 mg l-1 gibberellic acid (GA3). The rooting percentage was highest (90%) on 0.5_ MS medium supplemented with 0.5 mg l-1 indole-3-butyric acid (IBA), which was more effective than rooting (70%) on 1.0 mg l-1 _-naphthaleneacetic acid (NAA). Well-rooted plantlets exhibited normal growth under greenhouse conditions, with a 98% survival rate. Plants derived from this multiple shoot induction protocol showed no apparent differences in phenotype in the greenhouse. This protocol would be of use for large-scale propagation of mulberry to support ex situ conservation.