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Differential gene expression and developmental competence in in vitro produced bovine embryos

Published online by Cambridge University Press:  15 April 2011

Paula Ripamonte ,
Lígia Garcia Mesquita ,
Sylvia Sanches Cortezzi ,
Júlio César de Carvalho Balieiro ,
Giovana Krempel Fonseca Merighe ,
Yeda Fumie Watanabe ,
Alexandre Rodrigues Caetano  and
Flávio Vieira Meirelles
Paula Ripamonte
Affiliation:
Universidade de São Paulo, Faculdade de Zootecnia e Engenharia de Alimentos, Departamento de Ciências Básicas, Pirassununga, SP, Brazil.
Lígia Garcia Mesquita
Affiliation:
Universidade de São Paulo, Faculdade de Zootecnia e Engenharia de Alimentos, Departamento de Ciências Básicas, Pirassununga, SP, Brazil.
Sylvia Sanches Cortezzi
Affiliation:
Universidade de São Paulo, Faculdade de Zootecnia e Engenharia de Alimentos, Departamento de Ciências Básicas, Pirassununga, SP, Brazil.
Júlio César de Carvalho Balieiro
Affiliation:
Universidade de São Paulo, Faculdade de Zootecnia e Engenharia de Alimentos, Departamento de Ciências Básicas, Pirassununga, SP, Brazil.
Giovana Krempel Fonseca Merighe
Affiliation:
Universidade de São Paulo, Faculdade de Zootecnia e Engenharia de Alimentos, Departamento de Ciências Básicas, Pirassununga, SP, Brazil.
Yeda Fumie Watanabe
Affiliation:
Universidade de São Paulo, Faculdade de Zootecnia e Engenharia de Alimentos, Departamento de Ciências Básicas, Pirassununga, SP, Brazil.
Alexandre Rodrigues Caetano
Affiliation:
EMBRAPA Recursos Genéticos e Biotecnologia, Brasília, DF, Brazil.
Flávio Vieira Meirelles*
Affiliation:
Departamento de Ciências Básicas, Universidade de São Paulo, FZEA, Pirassununga, SP, Brazil., Av. Duque de Caxias Norte 225, Pirassununga, São Paulo, Brasil CEP 13635–900.
*
All correspondence to: F.V. Meirelles. Departamento de Ciências Básicas, Universidade de São Paulo, FZEA, Pirassununga, SP, Brazil., Av. Duque de Caxias Norte 225, Pirassununga, São Paulo, Brasil CEP 13635–900. Tel: +55 19 3565 4112. e-mail: [email protected]
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Summary

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.

Keywords

Bovine embryoDDPCRDevelopmental blockGenome activationReal-time PCR
Type
Research Article
Information
Zygote , Volume 20 , Issue 3 , August 2012 , pp. 281 - 290
Copyright
Copyright © Cambridge University Press 2011

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