In vitro culture and mtDNA fate of ibex-rabbit nuclear transfer embryos.
Rabbit oocyte can be used as the recipient in interspecies somatic cell nuclear transfer (iSCNT). This work was undertaken in order to study the developmental competence of Capra ibex somatic cells reprogrammed by rabbit oocytes and the fate of mitochondria in iSCNT embryos. Metaphase II (MII) oocytes from superovulated rabbit were used as nuclear recipients. The nuclear donors were Capra ibex somatic cells with different proliferative status: population doubling time (PDL) = 15 +/- 2 (group 1), 35 +/- 2 (group 2), 55 +/- 2 (group 3) and 70 +/- 2 (group 4). Oocytes reconstructed with electrical pulses (2.1kV/cm, 10 micros, 2 times) were activated (1.4kV, 20 micros, 2 times) and then cultured in Medium199 containing 10% fetal bovine serum at 38.5 degrees C, 5% CO2 in air. In groups 1, 2, 3 and 4, the fusion rates were 35.83%, 66.03%, 65.40% and 35.35%, respectively. Similar cleavage rates were observed among the four groups. However, the developmental potential to morula/blastocyst from early nuclear donor embryos (16.42%/10.45%) was significantly higher (p < 0.05) than in terminal donor embryos (9.52%/3.81%). Polymerase chain reaction analysis of the mitochondrial (mt) DNA cytb gene demonstrated that mtDNAs from ibex and rabbit could be detected at various developmental stages before implantation. In conclusion, our results provide some original information about rescuing Capra ibex using the iSCNT technique. These results indicate that: (1) enucleated rabbit oocytes make Capra ibex fibroblast nuclei reprogramme; (2) the proliferative status of donor cells affects the efficiency of iSCNT; and (3) rabbit ooplasm rescues the donor-derived mtDNAs, resulting in mtDNA heteroplasmy before implantation.
Jiang
Yan
Y
State Key Laboratory of Reproductive Biology, Institute of Zoology, Chinese Academy of Sciences, Beijing, China.
Chen
Tao
T
Nan
Chang-Long
CL
Ouyang
Ying-Chun
YC
Sun
Qing-Yuan
QY
Chen
Da-Yuan
DY
D000818:Animals; D001755:Blastocyst / Q000166:cytology; D019976:Cloning, Organism / Q000379:methods*; D004272:DNA, Mitochondrial / Q000032:analysis; D010064:Embryo Implantation; D004624:Embryo Transfer; D004622:Embryo, Mammalian; D047108:Embryonic Development / Q000235:genetics; D005260:Female; D005307:Fertilization in Vitro; D005821:Genetic Techniques; D053652:Nuclear Transfer Techniques; D009865:Oocytes; D011817:Rabbits
D004272:DNA, Mitochondrial