Genetic characteristics of genome donors and selection of the trans-mitochondrial cybrids.

aρ0 MDA-MB-231 cells were used as nuclear DNA donors and mtDNA recipients. As mtDNA donors, we used HeLamt231 and HeLamtFt cybrids sharing the HeLa nuclear genome background to exclude the influence of variations in nuclear-coded cytoplasmic factors on the phenotypes. HeLamt231 cybrids carrying nuclear DNA from HeLa cells and mtDNA from MDA-MB-231 cells were isolated by the fusion of ρ∼ HeLa cells with enucleated MDA-MB-231 cells and subsequent cultivation in selection medium with 6-thioguanine medium without uridine and pyruvate (6-tg+UP−). The 6-tg eliminates unenucleated MDA-MB-231 cells and UP− eliminates unfused ρ0 HeLa cells, which require uridine and pyruvate due to their complete respiration defects caused by mtDNA depletion. HeLamtFt cybrids carrying nuclear DNA from HeLa cells and mtDNA from human fetal skin fibroblasts were obtained previously [17] by the fusion of ρ0 HeLa cells with enucleated human fetal skin fibroblasts and subsequent cultivation in the 6-tg+UP− medium.ben represents enucleated.cThe mtDNA donors sharing the HeLa nuclear genome background and expressing 6-tg resistance cannot survive in the presence of hypoxanthine/aminopterin/thymidine (HAT). Nuclear donor ρ0 MDA-MB-231 cells can grow in the HAT selection medium, but not in UP- selection medium. Thus, only transmitochondrial cybrids can survive in the selection medium.