Remodeling of donor nuclei, DNA-synthesis, and ploidy of bovine cumulus cell nuclear transfer embryos: effect of activation protocol.
The purpose of this study was to investigate the effects of two activation protocols on nuclear remodeling, DNA synthesis during the first cell cycle, chromosome segregation after first mitosis and development to blastocyst of embryos produced by somatic nuclear transfer. Pronuclear formation was significantly higher when activation lasted 5 hr compared to 3 hr for both ethanol-cycloheximide and ionomycin-bohemine treatment. However, the presence of a single nucleus was significantly higher in embryos activated for 3 hr in bohemine. Initiation of DNA synthesis was delayed in ethanol-cycloheximide group, however, after 12 hr labeling 100% of embryos synthesized DNA in both groups. Embryos activated with ethanol-cycloheximide developed to blastocysts at a significantly higher rate than those activated with ionomycin-bohemine. Analysis of 2-cell embryos with DNA probes for chromosome 6, 7, and 15 by fluorescence in situ hybridization showed that at least 50% of NT embryos were of normal ploidy independent of the activation stimulus. The results presented in this study show differences between the protocols compared on the nuclear events during the first cell cycle and on the development to blastocyst. Mol. Reprod. Dev. 59: 371-379, 2001.
Alberio
R
R
Department of Molecular Animal Breeding and Genetics, Ludwig-Maximilian University, Oberschleissheim, Germany.
Brero
A
A
Motlík
J
J
Cremer
T
T
Wolf
E
E
Zakhartchenko
V
V
D000818:Animals; D001755:Blastocyst / Q000502:physiology*; D002417:Cattle; D002453:Cell Cycle / Q000502:physiology; D019070:Cell Lineage; D002467:Cell Nucleus / Q000378:metabolism; D002875:Chromosomes / Q000378:metabolism; D019976:Cloning, Organism; D003513:Cycloheximide / Q000494:pharmacology; D004247:DNA / Q000096:biosynthesis*; D004622:Embryo, Mammalian / Q000166:cytology / Q000187:drug effects / Q000502:physiology*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D000431:Ethanol / Q000494:pharmacology; D017404:In Situ Hybridization, Fluorescence; D053652:Nuclear Transfer Techniques; D009865:Oocytes / Q000166:cytology*; D011003:Ploidies; D011500:Protein Synthesis Inhibitors / Q000494:pharmacology; D011687:Purines / Q000494:pharmacology
D004791:Enzyme Inhibitors; D011500:Protein Synthesis Inhibitors; D011687:Purines; C118034:bohemine; D000431:Ethanol; D004247:DNA; D003513:Cycloheximide