Development of parthenogenetic and cloned ovine embryos: effect of activation protocols

Preliminary experiments carried out on ovine oocytes were designed to establish correlations between activation protocols and subsequent rates of embryonic development. The best activation protocols were thereafter used in studies on ovine parthenogenesis and cloning. The first study established tha...

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Bibliographic Details
Published in:Biology of reproduction 1998-05, Vol.58 (5), p.1177-1187
Main Authors: Loi, P, Ledda, S, Fulka, J. Jr, Cappai, P, Moor, R.M
Format: Article
Language:English
Subjects:
6-DIMETHYLAMINOPURINE
Adenine - analogs & derivatives
Adenine - pharmacology
Animals
Biological and medical sciences
Blastocyst - drug effects
Blastocyst - metabolism
Blastocyst - ultrastructure
Blastomeres - drug effects
Blastomeres - ultrastructure
Cell Nucleus - drug effects
Cell Nucleus - physiology
CLONACION
CLONAGE
Clone Cells - drug effects
Clone Cells - physiology
CLONING
CULTIVO DE EMBRIONES
CULTURE D'EMBRYON
DESARROLLO EMBRIONARIO
DEVELOPMENTAL COMPETENCE
DEVELOPPEMENT EMBRYONNAIRE
Diploidy
DNA REPLICATION
EMBRYO CLONING
EMBRYO CULTURE
EMBRYO TRANSFER
Embryo, Mammalian - cytology
Embryo, Mammalian - drug effects
Embryo, Mammalian - physiology
Embryology: invertebrates and vertebrates. Teratology
Embryonic and Fetal Development - drug effects
Embryonic and Fetal Development - physiology
EMBRYONIC DEVELOPMENT
ENZYME INHIBITORS
Enzyme Inhibitors - pharmacology
Female
FETAL DEVELOPMENT
Fundamental and applied biological sciences. Psychology
Germ Cells - drug effects
Germ Cells - physiology
Germ Cells - ultrastructure
INHIBIDORES DE ENZIMAS
INHIBITEUR D'ENZYME
Molecular embryology
NOYAU CELLULAIRE
NUCLEI
NUCLEO
NUCLEUS
OOCYTE ACTIVATION
OOCYTES
Oocytes - drug effects
Oocytes - physiology
Oocytes - ultrastructure
OVA
OVIN
OVINOS
OVULE
OVULO
PARTENOGENESIS
PARTHENOGENESE
PARTHENOGENESIS
Parthenogenesis - physiology
PARTHENOGENETIC EMBRYOS
Pregnancy
Protein Kinase Inhibitors
REPLICACION
REPLICATION
SHEEP
Stimulation, Chemical
TRANSFERENCIA DE EMBRIONES
TRANSFERT EMBRYONNAIRE
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Summary:Preliminary experiments carried out on ovine oocytes were designed to establish correlations between activation protocols and subsequent rates of embryonic development. The best activation protocols were thereafter used in studies on ovine parthenogenesis and cloning. The first study established that chemical activators induce pronuclear development at a slightly higher rate than physical activation (ionomycin, 96%; ethanol, 95%; electro activation, 80%). Inhibition of second polar body extrusion and one single pronucleus were observed in the majority of the oocytes (approximately 90%) treated for 3 h with 6-dimethylaminopurine (6-DMAP) following either ionomycin or ethanol activation. While over 80% of these oocytes cleaved after transfer to the oviducts of recipients, progression to the blastocyst stage was higher after ionomycin as compared with ethanol activation (58% vs. 19%). The ionomycin plus 6-DMAP activation protocol was used to produce parthenogenetic blastocysts whose subsequent development was monitored both by ultrasonography and by direct fetal examination. Over 70% of parthenogenotes were viable on Day 21 of pregnancy but dead by Day 25. The effects of 6-DMAP on nuclear remodeling and fetal development of cloned embryos was then investigated. Control cloned embryos underwent nuclear envelope breakdown (NEBD), premature chromatin condensation (PCC), and inhibition of DNA synthesis. By contrast, reconstructed embryos treated with 6-DMAP exhibited intact nuclear membranes, interphase chromatin, and no interference on DNA synthesis. Moreover, cloned embryos developed to blastocyst stage in higher percentage after 6-DMAP treatment (83% vs. 25%). We conclude that ionomycin followed by 6-DMAP incubation yields high percentages of diploid parthenogenetic embryos that develop to Day 25 before dying. Cloned embryos activated by the ionomycin-6-DMAP protocol develop readily to term.
ISSN:0006-3363
1529-7268
DOI:10.1095/biolreprod58.5.1177