Effect of microinjection of a single IVF-derived blastomere on the development of cloned embryos at the eight-cell stage in bovine.
This study was conducted to determine the effect of microinjection of a single blastomere from in vitro fertilization (IVF)-derived eight-cell embryo into eight-cell cloned embryos harboring the gene encoding recombinant human lactoferrin (rhLF), GFP, and NEO markers in bovine. The reconstructed chimeric embryos were assessed for their development to blastocyst, or to term after transfer, and tissues of offspring were evaluated by polymerase chain reaction (PCR) for the presence of nuclear transfer (NT)-derived transgenic cells, and the cloned embryos without microinjection were used as controls. The chimeric embryos showed slightly higher blastocyst rate than that for controls. The single IVF-derived blastomere appeared to preferential contribute to inner cell mass (ICM) in the chimeric blastocysts. After transfer, the rates of development of chimeric embryos to day 60, to term, and to weaning were significantly higher than those of controls. Sixty-three chimeric blastocysts were transferred and 11 calves were born: 7 calves of them were dead, and the remaining 4 calves are apparently normal and healthy. Most of the tissues collected from dead fetus were transgenic, whereas NT-derived transgenic cells were not detected in some tissues of the living calves. Our results indicated that a single blastomere from IVF-derived eight-cell embryo improves the in vivo developmental potential of transgenic cloned eight-cell embryos in bovine; however, the single IVF-derived blastomere appears to be better able to populate the ICM and many tissues of offspring than NT-derived blastomeres.
Chen
Xiuping
X
State Key Laboratory for AgroBiotechnology, China Agricultural University, Beijing 100193, People's Republic of China.
Wang
Jianwu
J
Li
Rong
R
Ding
Fangrong
F
Li
Song
S
Zhang
Lei
L
Dai
Yunping
Y
Li
Ning
N
D000818:Animals; D030801:Animals, Genetically Modified; D001757:Blastomeres / Q000166:cytology*; D002417:Cattle; D002478:Cells, Cultured; D002678:Chimera; D019976:Cloning, Organism; D004622:Embryo, Mammalian / Q000166:cytology* / Q000502:physiology*; D047108:Embryonic Development / Q000502:physiology*; D005260:Female; D005307:Fertilization in Vitro / Q000379:methods*; D006801:Humans; D008297:Male; D008845:Microinjections; D011247:Pregnancy