FERTILE PLANTS REGENERATED FROM MESOPHYLL PROTOPLASTS OF COLD-TOLERANT RICE

A protocol for regeneration of fertile plants from leaf-sheath-derived protoplast of Oryza sativa (L.), which is a pre-requisite for utiliza tion of genetic manipulations at the cellular level without or with ve ry little creation of genetic variation, is reported. Isolated protopl asts from inner leaf sheath of 7-day-old seedlings were cultured in mo dified N6 medium in the presence of feeder cells. The presence of feed er cells and reduction of osmoticum played an important role in obtain ing the sustained divisions of mesophyll protoplasts. Among the three. procedures, i.e. protoplast cultured in liquid medium, in 0.15% agaro se and on top of a cellulose nitrate membrane in the presence of feede rs, the latter procedure exhibited the highest number of protoplast-de rived calli (protocalli), The protocalli obtained from 0.15%-agarose-c ultured protoplasts (without membrane) produced a larger number of pla ntlets.