title: Improvement of an electrical activation protocol for porcine oocytes.
abstract: Factors influencing pig oocyte activation by electrical stimulation were evaluated by their effect on the development of parthenogenetic embryos to the blastocyst stage to establish an effective activation protocol for pig nuclear transfer. This evaluation included 1) a comparison of the effect of epidermal growth factor and amino acids in maturation medium, 2) an investigation of interactions among oocyte age, applied voltage field strength, electrical pulse number, and pulse duration, and 3) a karyotype analysis of the parthenogenetic blastocysts yielded by an optimized protocol based on an in vitro system of oocyte maturation and embryo culture. In the first study, addition of amino acids in maturation medium was beneficial for the developmental competence of activated oocytes. In the second study, the developmental response of activated oocytes was dependent on interactions between oocyte age at activation and applied voltage field strength, voltage field strength and pulse number, and pulse number and duration. The formation of parthenogenetic blastocysts was optimal when activation was at 44 h of maturation using three 80-microsec consecutive pulses of 1.0 kV/cm DC. Approximately 84% of parthenogenetic blastocysts yielded by this protocol were diploid, implying a potential for further in vivo development.
authors.affiliation: Department of Gene Expression and Development, Roslin Institute, Roslin, Midlothian EH25 9PS, United Kingdom.
authors.forename: Jie
authors.initials: J
authors.lastname: Zhu
authors.forename: Evelyn E
authors.initials: EE
authors.lastname: Telfer
authors.forename: Judy
authors.initials: J
authors.lastname: Fletcher
authors.forename: Anthea
authors.initials: A
authors.lastname: Springbett
authors.forename: John R
authors.initials: JR
authors.lastname: Dobrinsky
authors.forename: Paul A
authors.initials: PA
authors.lastname: De Sousa
authors.forename: Ian
authors.initials: I
authors.lastname: Wilmut
mesh_terms: D000596:Amino Acids / Q000494:pharmacology; D000818:Animals; D001755:Blastocyst / Q000502:physiology; D003470:Culture Media; D046508:Culture Techniques; D004558:Electric Stimulation; D004815:Epidermal Growth Factor / Q000494:pharmacology; D005260:Female; D007621:Karyotyping; D053652:Nuclear Transfer Techniques; D009865:Oocytes / Q000502:physiology*; D010312:Parthenogenesis; D013552:Swine / Q000502:physiology*; D013997:Time Factors
chemical_list: D000596:Amino Acids; D003470:Culture Media; D004815:Epidermal Growth Factor