CYBRID PRODUCTION BASED ON MUTAGENIC INACTIVATION OF PROTOPLASTS AND RESCUING OF MUTANT PLASTIDS IN FUSION PRODUCTS - POTATO WITH A PLASTOME FROM S-BULBOCASTANUM AND S-PINNATISECTUM

A procedure for cybrid production, based on double treatment of donor protoplasts by physical and afterwards chemical mutagens at superletha l doses (gamma-irradiation at a dose of 1000 Gy was applied for the in activation of nuclei; 3-5 mM N-nitroso-N-methylurea was used for the e fficient induction of plastome mutation) and the rescuing of mutant pl astids after fusion with untreated recipient protoplasts, was develope d. For identification of mutant donor-type plastids in fusion products a selection for streptomycin was performed. In two sets of experiment s, in which S. tuberosum served as the recipient of foreign cytoplasm with the wild tuber-bearing species S. bulbocastanum and S. pinnatisec tum as donors, a total of about 40 streptomycin-resistant colonies was isolated. Eight regenerants from the S. tuberosum + S. bulbocastanum fusion combination and four from S. tuberosum + S. pinnatisectum were further investigated using chromosome counting, analysis of esterase i soenzymes, restriction analysis of organelle DNA, and blot hybridizati on. All but one plant from both combinations were characterised as pot ato cybrids possessing exclusively foreign plastids and retaining a mo rphology typical of the recipient. Only in one line was rearranged mtD NA detected. The availability of potato cybrids facilitates the analys is of plastome-encoded breeding traits and the identification of the m ost valuable source of cytoplasm among the wild potato species. The de scribed system for producing cybrids without genetic selectable marker s in the parental material offers the possibility for the rescue of cy toplasmic mutations which are impossible to isolate by conventional ap proaches.