Improving ovary and embryo culture techniques for efficient resynthesis of Brassica napus from reciprocal crosses between yellow-seeded diploids B-rapa and B-oleracea

The primary aim of this study was to optimize in vitro culture protocols to establish an efficient reproducible culture system for different Brassica interspecific crosses, and to synthesize yellow-seeded Brassica napus (AACC) for breeding and genetical studies. Reciprocal crosses were carried out between three B. rapa L. ssp. oleifera varieties (AA) and five accessions of B. oleracea var. acephala (CC). All the parental lines were yellow-seeded except one accession of B. oleracea. Hybrids were obtained through either ovary culture from crosses B. rapa x B. oleracea, or embryo culture from crosses B. oleracea x B. rapa. A higher rate of hybrid production was recorded when ovaries were cultured at 4-7 days after pollination (DAP). Of different culture media, medium E (MS with half strength macronutrients) showed good response for ovaries from all the crosses, the highest rate of hybrid production reaching 45% in B. rapa (1151) x B. oleracea (T-2). In embryo culture, the hybrid rate was significantly enhanced at 16-18 DAP, up to 48.1% in B. oleracea (T-3) x B. rapa (JB(2)). The combinations of optimal DAP for excision and media components increased recovery of hybrids for ovary and embryo culture, and constituted an improved technique for B. rapa x B. oleracea crosses. In addition, yellow seeds were obtained from progenies of two crosses, indicating the feasibility of developing yellow-seeded B. napus through the hybridization between yellow-seeded diploids B. rapa and B. oleracea var. acephala.