title: Transmission of mitochondrial DNA in pigs and progeny derived from nuclear transfer of Meishan pig fibroblast cells.
abstract: In embryos derived by nuclear transfer (NT), fusion, or injection of donor cells with recipient oocytes caused mitochondrial heteroplasmy. Previous studies have reported varying patterns of mitochondrial DNA (mtDNA) transmission in cloned calves. Here, we examined the transmission of mtDNA from NT pigs to their progeny. NT pigs were created by microinjection of Meishan pig fetal fibroblast nuclei into enucleated oocytes (maternal Landrace background). Transmission of donor cell (Meishan) mtDNA was analyzed using 4 NT pigs and 25 of their progeny by PCR-mediated single-strand conformation polymorphism (PCR-SSCP) analysis, PCR-RFLP, and a specific PCR to detect Meishan mtDNA single nucleotide polymorphisms (SNP-PCR). In the blood and hair root of NT pigs, donor mtDNAs were not detected by PCR-SSCP and PCR-RFLP, but detected by SNP-PCR. These results indicated that donor mtDNAs comprised between 0.1% and 1% of total mtDNA. Only one of the progeny exhibited heteroplasmy with donor cell mtDNA populations, ranging from 0% to 44% in selected tissues. Additionally, other progeny of the same heteroplasmic founder pig were analyzed, and 89% (16/18) harbored donor cell mtDNA populations. The proportion of donor mtDNA was significantly higher in liver (12.9 +/- 8.3%) than in spleen (5.0 +/- 3.9%), ear (6.7 +/- 5.3%), and blood (5.8 +/- 3.7%) (P < 0.01). These results demonstrated that donor mtDNAs in NT pigs could be transmitted to progeny. Moreover, once heteroplasmy was transmitted to progeny of NT-derived pigs, it appears that the introduced mitochondrial populations become fixed and maternally-derived heteroplasmy was more readily maintained in subsequent generations.
authors.affiliation: Department of Animal Breeding and Reproduction, National Institute of Livestock and Grassland Science, National Agricultural Research Organization, Tsukuba, Japan. kumiko@affrc.go.jp
authors.forename: Kumiko
authors.initials: K
authors.lastname: Takeda
authors.forename: Mariko
authors.initials: M
authors.lastname: Tasai
authors.forename: Masaki
authors.initials: M
authors.lastname: Iwamoto
authors.forename: Tomiji
authors.initials: T
authors.lastname: Akita
authors.forename: Takahiro
authors.initials: T
authors.lastname: Tagami
authors.forename: Keijiro
authors.initials: K
authors.lastname: Nirasawa
authors.forename: Hirofumi
authors.initials: H
authors.lastname: Hanada
authors.forename: Akira
authors.initials: A
authors.lastname: Onishi
mesh_terms: D000818:Animals; D019976:Cloning, Organism / Q000379:methods*; D004272:DNA, Mitochondrial / Q000235:genetics*; D005111:Extrachromosomal Inheritance / Q000235:genetics*; D005347:Fibroblasts / Q000166:cytology*; D008845:Microinjections; D053652:Nuclear Transfer Techniques; D009865:Oocytes / Q000166:cytology / Q000502:physiology; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013552:Swine
chemical_list: D004272:DNA, Mitochondrial