title: Differential gene expression and developmental competence in in vitro produced bovine embryos.
abstract: The embryonic developmental block occurs at the 8-cell stage in cattle and is characterized by a lengthening of the cell cycle and an increased number of embryos that stop development. The maternal-embryonic transition arises at the same stage resulting in the transcription of many genes. Gene expression studies during this stage may contribute to the understanding of the physiological mechanisms involved in the maternal-embryonic transition. Herein we identified genes differentially expressed between embryos with high or low developmental competence to reach the blastocyst stage using differential display PCR. Embryos were analysed according to developmental kinetics: fast cleavage embryos showing 8 cells at 48 h post insemination (hpi) with high potential of development (F8), and embryos with slow cleavage presenting 4 cells at 48 hpi (S4) and 8 cells at 90 hpi (S8), both with reduced rates of development to blastocyst. The fluorescence DDPCR method was applied and allowed the recovery of 176 differentially expressed bands with similar proportion between high and low development potential groups (52% to F8 and 48% in S4 and S8 groups). A total of 27 isolated fragments were cloned and sequenced, confirming the expected primer sequences and allowing the identification of 27 gene transcripts. PI3KCA and ITM2B were chosen for relative quantification of mRNA using real-time PCR and showed a kinetic and a time-related pattern of expression respectively. The observed results suggest the existence of two different embryonic genome activation mechanisms: fast-developing embryos activate genes related to embryonic development, and slow-developing embryos activate genes related to cellular survival and/or death.
authors.affiliation: Universidade de São Paulo, Faculdade de Zootecnia e Engenharia de Alimentos, Departamento de Ciências Básicas, Pirassununga, SP, Brazil.
authors.forename: Paula
authors.initials: P
authors.lastname: Ripamonte
authors.forename: Lígia Garcia
authors.initials: LG
authors.lastname: Mesquita
authors.forename: Sylvia Sanches
authors.initials: SS
authors.lastname: Cortezzi
authors.forename: Júlio César
authors.initials: JC
authors.lastname: de Carvalho Balieiro
authors.forename: Giovana Krempel
authors.initials: GK
authors.lastname: Fonseca Merighe
authors.forename: Yeda Fumie
authors.initials: YF
authors.lastname: Watanabe
authors.forename: Alexandre Rodrigues
authors.initials: AR
authors.lastname: Caetano
authors.forename: Flávio Vieira
authors.initials: FV
authors.lastname: Meirelles
mesh_terms: D000818:Animals; D002417:Cattle; D002453:Cell Cycle; D004622:Embryo, Mammalian / Q000378:metabolism*; D005260:Female; D005307:Fertilization in Vitro; D020869:Gene Expression Profiling; D018507:Gene Expression Regulation, Developmental; D008562:Membrane Glycoproteins / Q000235:genetics / Q000378:metabolism; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism
chemical_list: D008562:Membrane Glycoproteins; D009687:Nuclear Proteins; D012333:RNA, Messenger