Isolation of protoplasts from cardamom (Elettaria cardamomum Maton.) and ginger (zingiber officinale Rosc.)

Abstract

Protoplasts were isolated from leaf mesophyll tissue, collected from in vitro grown plantlets and cell suspension cultures of cardamom (Elettaria cardamomum) and ginger (zingiber officinale). In cardamom, a protoplast yield of 3.5*105/g of leaf tissue was obtained when incubated in an enzyme solution containing 0.5% macerozyme R10, 2% cellulose Onozuka R10 and 9% mannitol for 18-20 h at 25oC in dark. The yield of protoplasts from cell suspension culture was 1.5*105/g tissue, when incubated in 1% macerozyme R10 and 2% cellulose Onozuka R10 for 24 h at 25oC with gentle shaking at 53 rpm in dark. The viability of leaf mesophyll protoplast was 75% and that of cell suspension was 40% on Evan’s blue staining. In ginger, a protoplast yield of 2.5*105/g of leaf tissue was obtained on digestion in an enzyme solution containing 0.5% macerozyme R10, 3% hemicellulase and 5% cellulose Onozuka R10, when incubated for 10 h at 15oC followed by 6 h at 30 oC. The protoplast viability was 55% protoplast yield from cell suspension culture was 1*105/g of callus when digested with an enzyme solution of 1% macerozyme R10, 3% hemicellulase and 6% cellulose Onozuka R10 and incubated for 10h at 15oC and later at 30 oC for 8 h. Seventy twoper cent of the protoplast were viable. The protoplasts from both the species could be cultured and made to develop up to microcalli stage