Intervarietal and interspecific chimera formation by in vitro graft-culture method in Brassica

An efficient chimera formation method by tissue culture combined with grafting was studied in Brassica. Cabbage cultivars "Ruby-ball" and "YR-ranpou" (Brassica oleracea) were used for intervarietal chimera formation. Seven-day-old seedlings leaving one of two cotyledons were approach-grafted and cultured in vitro (AGSC method). Chimeric shoots were obtained by the subculture of directly growing chimeric leaves (DGCL) from grafted part and cross-cut section of the united part after the graft culture. These were rooted and grown to complete chimera. An approach-grafted culture method was also available for interspecific chimera formation between "Komatsuna" (B. campestris) and "Ruby-ball" (B. oleracea), resulting in 20% formation per culture. Interspecific chimeras were identified as complex and peripheral-sectorial type by microscopic observation and the electrophoretic analysis of acid phosphatase isozyme pattern. The AGSC method was more effective than the usual Winkler's graft method and a simple mixed culture of heterogeneous cells or tissues.