The cotyledon protoplasts culture and plant regeneration of Brassica napus L.

Protoplasts were prepared from the cotyledon of Brassica napus cultivar Zhongshuang 6,cultured with solid-liquid combined method.The optimized conditions about the isolation,culture and regeneration of protoplast were tested and ultimately the regeneration plantlets were obtained.The results indicated that high recovery of protoplast could be achieved with enzyme mixture(0.5% cellulase,0.1% pectolyase + 0.2mol/L mannitol + 80mol/L CaCl2) diluted with SCM solution as 3∶7 ratio.It is fine to culture protoplast on solid-liquid combined media B and C.The optimum differentiation medium was E+1.0g/L 6-BA+0.1g/L NAA+0.02g/L with GA3+30μmol/L AgNO3.The best rooting media was MS without hormones.This study provided new formula and data for establishing sophisticated protoplast regeneration system of Brassica napus L..