Factors affecting mango ( Mangifera indica L.) protoplast isolation and culture

Abstract

The major factors influencing protoplast isolation and culture of mango ( Mangifera indica L.) cv. Kensington Pride were investigated. The resultant protocol was used to compare plating efficiency among 4 mango cultivars. Most responses differed between proembryonic masses (PEMs) and leaf sources. Protoplast yields of 15.22 × 10 6 g −1 from PEMs and 8.68 × 10 6 g −1 from greenhouse-derived leaves were obtained in a solution of 0.7 M mannitol CPW plus 1.5% cellulase, 1% hemicellulase and 0.75% macerozyme for PEMs or 0.5 M mannitol CPW plus 1.5% cellulose, 1% hemicellulase and 1.5% macerozyme for leaves. Culture in Ca-alginate beads with initial plating densities (IPD) of 2.5 × 10 4 Pp mL −1 for PEMs and 2.5 × 10 5 for leaves gave the highest plating efficiencies (FPE). For PEMs 1 mg L −1 2,4- d and 3.5 mg L −1 kinetin gave an FPE of 2.85% whereas lower kinetin (2 mg L −1) plus 0.5 mg L −1 6-BAP was most effective for leaves (FPE of 2.12%). Most protoplast mortality occurred during the first week of culture and was more severe in liquid culture. In Ca-alginate beads, protoplast survival at 14 days was higher for PEMs (30%) than leaf (21%) as was the frequency of cell division (17.6% compared to 13.6%). PEMs protoplasts continued development through embryogenesis to in vitro plantlet regeneration whereas leaf protoplasts underwent cell division up to 40-cell colonies but failed to proceed further. For PEMs, polyembryonic cvs. Kensington Pride and Keow Savoey produced higher FPE (1.95%) than monoembryonic cvs. Tommy Atkins and Keitt (1.75%). There was no effect of cultivar for leaf protoplasts.