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Sperm-mediated gene transfer: effect on bovine in vitro embryo production

Published online by Cambridge University Press:  18 July 2012

Renata Simões ,
Alessandra Corallo Nicacio ,
Mario Binelli ,
Fabiola Freitas de Paula-Lopes ,
Marcella Pecora Milazzotto ,
José Antonio Visintin  and
Mayra Elena Ortiz D'Ávila Assumpção
Renata Simões
Affiliation:
Centro de Ciências Naturais e Humanas, Federal University of ABC (UFABC); Santo André-SP; Brazil.
Alessandra Corallo Nicacio
Affiliation:
Embrapa Beef Cattle Research Center, Campo Grande, Mato Grosso do Sul, Brazil.
Mario Binelli
Affiliation:
School of Veterinary Medicine and Animal Science, University of São Paulo, Department of Animal Reproduction, São Paulo, Brazil.
Fabiola Freitas de Paula-Lopes
Affiliation:
Department of Biological Science of Federal University of São Paulo; Diadema-SP; Brazil.
Marcella Pecora Milazzotto
Affiliation:
Centro de Ciências Naturais e Humanas, Federal University of ABC (UFABC); Santo André-SP; Brazil.
José Antonio Visintin
Affiliation:
School of Veterinary Medicine and Animal Science, University of São Paulo, Department of Animal Reproduction, São Paulo, Brazil.
Mayra Elena Ortiz D'Ávila Assumpção*
Affiliation:
Department of Animal Reproduction, School of Veterinary Medicine and Animal Science, University of São Paulo, Av. Prof. Dr. Orlando Marques de Paiva, 87–Cidade Universitária–05508-270–São Paulo, SP, Brazil.
*
All corresponding to: Mayra Elena Ortiz D'Avila Assumpção. Department of Animal Reproduction, School of Veterinary Medicine and Animal Science, University of São Paulo, Av. Prof. Dr. Orlando Marques de Paiva, 87–Cidade Universitária–05508-270–São Paulo, SP, Brazil. Tel: +55 11 3091 7915. Fax: +55 11 3091 7412. e-mail: meoaa@usp.br
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Summary

The technique of sperm-mediated gene transfer (SMGT) can be used to delivery exogenous DNA into the oocyte. However, it has low repeatability and produces inconsistent results. In order to optimize this technique, it is necessary to study the mechanism by which DNA enters the sperm cell and integrates in the sperm genome. Furthermore, studies must focus in the maintenance of sperm cell viability and function. The aim of this study was to evaluate different SMGT protocols of sperm electroporation or capacitation (CaI) aiming to maintain sperm viability in the production of bovine embryos in vitro. Frozen–thawed semen was divided in two experimental groups (electroporation or CaI) and one control group (non-treated cells). For the electroporation method, five different voltages (100, 500, 750, 1000 or 1500 V) with 25 μF capacitance were used. For CaI treatment, combinations of two CaI concentrations (250 nM or 500 nM), two incubation periods of sperm cells with CaI (1 or 5 min) and two incubation periods that mimicked time of sperm cell interaction with exogenous DNA molecules (1 or 2 h) were evaluated. According to our data, electroporation and CaI treatments do not prevent sperm penetration and oocyte fertilization and can be an alternative method to achieve satisfactory DNA delivery in SMGT protocols.

Keywords

BovineCalcium ionophoreElectroporationSMGTIVP embryo
Type
Research Article
Information
Zygote , Volume 21 , Issue 4 , November 2013 , pp. 325 - 329
Copyright
Copyright © Cambridge University Press 2012 

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