High frequency axillary shoot proliferation and plant regeneration from cotyledonary nodes of pomegranate (Punica granatum L.)

A complete protocol is presented for in vitro regeneration of pomegranate ( Punica granatum L.), a tropical fruit tree, using cotyledonary nodes derive d from axenic seedlings. Shoot development was induced from cotyledonary no des on Murashige and Skoog (1962) (MS) medium supplemented with 2.3-23.0 mu M benzyladenine (BA) or kinetin (Kn). Both type and concentration of cytok inin significantly influenced shoot proliferation. The maximum number of sh oots (9.8 shoots/explant) was developed on a medium containing 9.0 mu M BA. Shoot culture was established by repeatedly subculturing the original coty ledonary node on a fresh batch of the same medium after each harvest of the newly formed shoots. In vitro raised shoots were cut into nodal segments a nd cultured on a fresh medium for further multiplication. Thus, from a sing le cotyledonary node about 30-35 shoots were obtained in 60 days. Shoots fo rmed in vitro were rooted on half-strength MS supplemented with 0.054-5.4 m u M naphthaleneacetic acid (NAA). However, a medium containing 0.54 mu M NA A resulted in the highest per cent rooting of shoots and significantly high er number of roots than other concentrations. Plantlets were successfully a cclimated and established in soil. (C) 2000 Elsevier Science B.V. All right s reserved.