Protoplast isolation in Daucus carota and Momordica charantia: regeneration and cold storage studies in carrot

Carrot and bitter gourd are two important vegetable crops of the Indian subcontinent with immense nutritional and medicinal value. Both of these crops experience inconsistent yields when propagated vegetatively. Plant regeneration through protoplasts is known to develop plantlets in a shorter time and higher quantity at a larger scale than other tissue culture methods. In the present study, optimization of the enzymatic methodology for protoplast isolation of leaves of carrot (Daucus carota) and bitter gourd (Momordica charantia) was executed. The Champion F1 carrot variety resulted in a higher protoplast yield (4.9 ± 0.20 × 106 protoplasts per gram) using Cellulase Onozuka R-10 and Pectolyase Y-23 enzyme combination compared to the Naigara one. For both varieties, protoplast viability was found to be ≥ 70%. Embryogenesis and non-cryogenic cold storage studies were conducted at 2–8 °C for the Champion F1 carrot variety. The Champion F1 protoplasts in carrot petiole protoplast media stored at 2–8 °C for five months were successfully regenerated. 216 out of 253 plants survived, giving 85% efficiency of cold-stored protoplasts and yielded healthy orange carrots with acclimatization after three months of the growth cycle. The present study also reports the protoplast isolation of bitter gourd with a yield of 1.18 ± 0.07 × 106 protoplasts per gram using the optimum combination of Cellulase Onozuka R-10 and Macerozyme R-10 enzymes. To the author’s knowledge, this is the first reported study of protoplast isolation from bitter gourd.