In a previous study, we used mouse zygotes as recipients of mtDNA with a large-scale deletion mutation (Delta mtDNA) and generated respiration-deficient mice (mito-mice) carrying Delta mtDNA. In this study, we used mouse ES cells as recipients of Delta mtDNA, and generated mito-mice with Delta mtDNA only when the ES cells carried 17% Delta mtDNA. No chimera mice or their F, progenies were obtained from ES cells carrying more than 61% Delta mtDNA. These observations suggest that respiratory defects of ES cells inhibit their normal differentiation into chimera mice and mito-mice, and that ES cells are more effective than zygotes for generation of mito-mice carrying mtDNAs without significant pathogenic mutations. (c) 2005 Elsevier Inc. All rights reserved.