Factors influencing plant regeneration from protoplasts isolated from long-term cell suspension culture of recalcitrant Indica rice cultivar IR36

Summary Factors influencing successful establishment of embryogenic cell-suspension cultures and plant regeneration from longterm cell suspension-derived protoplasts of the recalcitrant Indica rice cultivar IR36 were studied. The factors included cell and protoplast culture medium, protoplast culture procedure, the source of nurse cells, and the regeneration procedure. Embryogenic cell suspension cultures could only be established from mature seed-derived callus of IR36 in AA-based medium (Müller and Grafe, 1978). Protoplast-derived colonies could be obtained only using the filter-membrane nurse-culture procedure when Lolium multiflorum suspension cells served as nurse, rather than wild rice (Oryza ridleyi) and Japonica rice (Oryza sativa cv. Taipei 309) cells. The utilization of a two-step regeneration procedure led to regeneration of fertile plants from protoplasts isolated from 2-yr-old cell suspensions of IR36, one of the most important but recalcitrant rice cultivars.