Plant Cell. Tissue and Organ Culture 33: 339-341, 1993. © 1993 KluwerAcademic Publishers. Printedin the Netherlands. In vitro propagation as an aid for cloning of Morus laevigata Wall R. Islam, A. Zaman, O.I. Joarder & A.C. Barman Department of Botany, University of Rajshahi, Rajshahi-6205, Bangladesh Received 10 December 1991; accepted in revised form 7 December 1992 Key words: mulberry, micropropagation, nodal explants Mulberry (Morus sp.) is the most important crop in the sericulture industry because silkworms (Bombyx mori L.) feed on its leaves. Since cross pollination is the rule, enormous heterozygosity occurs in this plant (Das 1991). Morus laevigata is famous for quality timber and sweet edible fruits about 6-12 cm long (Jain & Kumar 1989). Again leaves of M. laevigata when fed to sil- kworms produce a better cocoon than those fed on M. alba (Mahmood et al. 1987). In spite of all these good qualities, M. laevigata is very difficult to propagate through cuttings because of its poor rooting ability (Rajah & Ravindran 1989). Clonal propagation of plants using tissue cul- ture technique has many applications in ag- ronomy (Murashige 1974), especially for woody plants that are extremely difficult to propagate by conventional means. Considerable work has been done in the last few years on in vitro propagation of M. nigra (Yadav et al. 1990), M. indica (Patel et la. 1983) and M. alba (Hossain et al. 1990). However, there is no report on in vitro culture of M. laevigata. Here we describe micropropagation using axillary buds from ma- ture plants of M. laevigata to provide an alter- native for its rapid clonal propagation. Young shoots each having 4-5 nodes were collected from a 12-year-old tree of M. laevigata in June-July and thoroughly washed in running tap water. Surface disinfestation was carried out with 0.1% (w/v) HgC12 for 10 min after a brief rinse in 70% (v/v) aq. ethanol. The material was then washed with sterilized double-distilled water for 10 rain giving 4-5 changes. Single node explants were then excised and cultured on MS medium (Murashige & Skoog 1962) containing different concentrations (2.5-15 IxM) of cyto- kinin, benzyladenine (BA) or kinetin individual- ly or in combination with 0.5 ~xM of an auxin, 2,4-dichlorophenoxyacetic acid (2,4-D) or. naphthaleneacetic acid (NAA). For adventitious root induction, MS medium with half-strength salts and sucrose with or without 0.5 ixM in- dolebutyric acid (IBA) was used. The pH of the medium was adjusted to 5.8 before autoclaving and solidified with 7 g 1-1 Difco Bacto-agar. The explants were cultured singly in 150 × 25mm culture tubes each containing 15 ml of culture medium and fitted with cotton plugs. The cul- tures were grown at 25-+2°C at a relative humidity of 55-65% (ambient conditions) under a 16-h photoperiod with a light intensity of 50- 60~mol m 2 s-1 provided by warm white fluorescent tubes. Data on days taken to bud proliferation, percentage of proliferating cul- tures, number of shoot per culture and shoot length (cm) were recorded after 5 weeks of culture. For each treatment 20 replicates were used and each experiment was repeated at least twice. Establishment of explants in vitro from adult trees was different due to excessive exudation of a latex-like milky substances. Two to three changes of media during the first week of culture were found to be most beneficial for a satisfac- tory response of explants in all instances. Nodal explants cultured on different treatment combi- nations showed their first response by enlarge- ment and break of axillary buds. Rapid and early proliferation was observed in media with lower concentrations of cytokinins (Table 1). Absence of growth regulators in the media delayed bud