PRODUCTION OF NEW CMS BRASSICA-OLERACEA BY TRANSFER OF ANAND CYTOPLASM FROM B-RAPA THROUGH PROTOPLAST FUSION

New types of cytoplasmic male sterility (CMS) in Brassica oleracea wou ld be useful for F-1 hybrid seed production. The 'Anand' cytoplasm der ives from the wild species B. tournefortii. Rapid cycling stocks of B. rapa and B. oleracea were used in cybridization experiments as donor and recipient of 'Anand' (= 'tour') CMS, respectively. Prior to fusion with PEG, donor protoplasts were inactivated with 30 krad gamma-rays and recipient ones with 3 mM iodoacetate, respectively. No calli were obtained from the pre-treated protoplasts. The frequency of shoot rege neration was 21-43% in untreated B. oleracea controls, but only 0-0.5% in 'Anand' B. rapa. Putative cybrids were regenerated from about 3% o f the calli from fused protoplasts. Regenerated plants were analyzed f or nuclear DNA content, plant and flower morphology, pollen production , female fertility, cold tolerance, and organelle composition. Eighty- one percent of the regenerated controls and 63% of fusion-derived plan ts were diploid. The rest showed DNA contents corresponding to 2x-4x, 4x, or higher ploidy levels, presumably due to somatic doubling in vit ro and/or fusions in which the donor nucleus was not completely elimin ated. Sixty-four percent of the cybrids had stamens and petals varying in size and shape and were male-sterile, with indehiscent anthers. Th eir phenotype was otherwise similar to that of B. oleracea. The remain ing plants had normal flowers and were male-fertile. Data from crosses with fertile pollinators indicated good female fertility in some of t he sterile lines, both after hand and insect pollinations in cages. Mi tochondrial (mt) segregation in the cybrids was slightly biased toward s 'Anand' mitochondria, and the presence of 'Anand' mtDNA fragments wa s strongly associated with male sterility. Evidence of mtDNA rearrange ments was obtained in some cybrids. Segregation of chloroplasts was sl ightly biased towards B. oleracea. The presence of 'Anand' chloroplast s with a B. oleracea nucleus did not result in cold temperature chloro sis, as seen in 'Ogura' CMS plants.