Establishing japonica rice suspensions retaining a high regeneration potential after 14 months of culture

We report here a method of ‘two cycles of selection’ for rapidly establishing rice embryogenic cell suspensions which had high regeneration potential even after long-term culture (up to 14 months). Embryogenic calli were induced from immature embryos of 17 genotypes of cultivated japonica rice (Oryza sativa L.) and tested for their regeneration potential. Five of the 17 genotypes showed relatively high regeneration frequencies (27%–45%) and were selected. This process was the first cycle selection, namely, the selection for responding genotypes. The second cycle of selection, or the selection for responding individual callus, was then conducted. Calli from different immature embryos of the same genotype were found to differ in regeneration potential, therefore calli with high regeneration potential were selected again in each of the five genotypes. Five finely-dispersed and rapidly-growing embryogenic cell suspensions, one for each of the genotypes, were established from the calli chosen by the two cycles of selection. The regeneration potential of these cell suspensions was tested every two months over a period of 14 months. All of the five genotypes retained high regeneration frequency (above 62%) over the whole period tested.