Interspecific protoplast fusion to rescue a cytoplasmic lincomycin resistance mutation into fertile Nicotiana plumbaginifolia plants

Summary A lincomycin-resistant cell line, LR105, was isolated in a mutagenized (0.1 mM N-ethyl-N-nitrosourea) callus culture initiated from a haploid Nicotiana sylvestris plant. The regenerated plants had an abnormal morphology and did not set viable seeds.Transfer of lincomycin resistance was attempted from the original N. sylvestris nuclear background into Nicotiana plumbaginifolia by protoplast fusion, since it was expected that resistance would be cytoplasmically coded. LR105 protoplasts were irradiated with a lethal dose (120 J kg^-1; ⁶⁰ Co source), fused with sensitive N. plumbaginifolia protoplasts and the colonies grown from the fused population were screened for lincomycin resistance. Expression of resistance was expected only if the cytoplasm of the irradiated cells had mixed with nonirradiated cytoplasm, and was reactivated as a result of cell fusion (Menczel et al. 1982).Plants were regenerated in 44 resistant clones. Plants in 41 clones had a N. plumbaginifolia nuclear genome. In three clones somatic hybrids were obtained. The resistant N. plumbaginifolia cybrid plants were fertile, unlike the original LR105 plants. Lincomycin resistance was inherited maternally in the eight clones in which crosses were made. In these clones the introduction of N. sylvestris chloroplasts into a N. plumbaginifolia nuclear background was confirmed by the SmaI restriction endonuclease pattern of the chloroplast DNA. The involvement of chloroplast DNA in determining lincomycin resistance is therefore implied.