Protoplast isolation from leaf mesophyll of sugarbeet Beta vulgaris L. axenic seedlings

Protoplasts were isolated from leaf mesophyll of sugarbeet (Beta vulgaris L.) axenic seedlings. Eight enzyme mixtures were tested for cell wall degrading ability. The efficient enzyme solution was mixture II that consist of 1.5% Cellulase RS, 2% Cellulase R10, 1% Macerozym R10 and 0.1% Pectolyase Y23. This mixture was efficient in releasing protoplasts and gave high yield of a density 7.3 × 104 protoplast / ml. These isolated protoplasts were viable 93%, their sizes ranged from 13 up to 52 μm, vacuolated and unvacuolated. This finding enable workers to focus on somatic hybridization through protoplast fusion to overcome some barriers facing gene transfer to improve plant species such as transfer of N2 fixation ability to non-legumes and producing resistant varieties to biotic stress and other traits.