Fate of donor mitochondrial DNA in cloned bovine embryos produced by microinjection of cumulus cells.
This study examined the fate of donor mitochondrial DNA during preimplantation development after nuclear transfer (NT) in cattle. Frozen-thawed cumulus cells were used as donor cells in the nuclear transfer. Mitochondrial DNA heteroplasmy in the nuclear transfer embryos was analyzed by allele-specific PCR (AS-PCR), direct DNA sequencing, and DNA chromatography. AS-PCR analysis for the detection of donor mitochondrial DNA was performed at the 1-, 2-, 4-, 8-, 16-cell, morula, and blastocyst stages of the embryos. The mitochondrial DNA from donor cells was detected at all developmental stages of the nuclear transfer embryos. However, mitochondrial DNA heteroplasmy was not observed in direct DNA sequencing of displacement-loop sequence from nuclear-transfer-derived blastocyst embryos. To confirm the mtDNA heteroplasmy in cloned embryos, the AS-PCR product from NT-derived blastocysts was analyzed by DNA sequencing and DNA chromatography. The nucleotides of NT-derived blastocysts were in accordance with the nucleotides from donor cells. These results indicate that the foreign cytoplasmic genome from donor cells was not destroyed by cytoplasmic events during preimplantation development that followed nuclear transfer.
Do
Jeong Tae
JT
Animal Resources Research Center, Department of Animal Science, Konkuk University, Kwangjin-gu 93-1, Seoul 143-702, Korea.
Lee
Jeong Woong
JW
Lee
Bo Yon
BY
Kim
Seung Bo
SB
Ryoo
Zae Young
ZY
Lee
Hoon Taek
HT
Chung
Kil Saeng
KS
D000483:Alleles; D000818:Animals; D001483:Base Sequence; D001755:Blastocyst; D002417:Cattle; D002467:Cell Nucleus / Q000235:genetics*; D019976:Cloning, Organism; D004272:DNA, Mitochondrial / Q000378:metabolism*; D005260:Female; D008845:Microinjections; D008969:Molecular Sequence Data; D009841:Oligonucleotides / Q000737:chemistry; D009865:Oocytes / Q000502:physiology; D010053:Ovary / Q000166:cytology*; D011110:Polymorphism, Genetic / Q000235:genetics; D011247:Pregnancy; D020133:Reverse Transcriptase Polymerase Chain Reaction
D004272:DNA, Mitochondrial; D009841:Oligonucleotides