Sperm genome cloning used in biparental bovine embryo reconstruction

The generation of androgenetic haploid embryos enables several haploid blastomeres to be obtained as identical copies of a single spermatozoon genome. In the present study, we compared the developmental ability of bovine androgenetic haploid embryos constructed by different methods, namely IVF and i...

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Detalles Bibliográficos
Otros Autores: Vichera, Gabriel Damián, Olivera, Ramiro, Sipowicz, Pablo, Radrizzani Helguera, Martín, Salamone, Daniel Felipe
Formato: Artículo
Lenguaje:Inglés
Materias:
SEXED HAPLOID NT
SPERM CLONING
TRANSGENESIS
ENHANCED GREEN FLUORESCENT PROTEIN
ANIMAL CELL
BLASTOCYST
BLASTOMERE
COW
EMBRYO
ENUCLEATION
GENE EXPRESSION
GENOME
HAPLOIDY
INTRACYTOPLASMIC SPERM INJECTION
MITOSIS
MOLECULAR CLONING
NONHUMAN
OOCYTE
SPERM
ANIMALS
BLASTOMERES
BREEDING
CATTLE
CLONING, MOLECULAR
EMBRYO CULTURE TECHNIQUES
EMBRYO, MAMMALIAN
FEMALE
FERTILIZATION IN VITRO
GREEN FLUORESCENT PROTEINS
IN SITU HYBRIDIZATION, FLUORESCENCE
MALE
NUCLEAR TRANSFER TECHNIQUES
POLYMERASE CHAIN REACTION
PREGNANCY
SPERM INJECTIONS, INTRACYTOPLASMIC
SPERMATOZOA
BOVINAE
Acceso en línea:http://ri.agro.uba.ar/files/intranet/articulo/2011Vichera2.pdf
LINK AL EDITOR
Aporte de:Registro referencial: Solicitar el recurso aquí
Biblioteca Central (FAUBA) de Universidad de Buenos Aires
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245 1 0 |a Sperm genome cloning used in biparental bovine embryo reconstruction 
520 |a The generation of androgenetic haploid embryos enables several haploid blastomeres to be obtained as identical copies of a single spermatozoon genome. In the present study, we compared the developmental ability of bovine androgenetic haploid embryos constructed by different methods, namely IVF and intracytoplasmic sperm injection [ICSI] before and after oocyte enucleation. Once obtained, the blastomeres of these androgenetic haploid embryos were used as male genome donors to reconstruct biparental embryos by fusion with matured oocytes. To verify the cytoplasmic contribution of androgenetic haploid blastomeres, we used spermatozoa incubated previously with exogenous DNA that coded for a green fluorescent protein gene [pCX-EGFP] and the enhanced green fluorescent protein [EGFP]-positive androgenetic haploid blastomeres generated were fused with mature oocytes. Of the reconstructed embryos reaching the cleavage and blastocyst stages, 85.1 percent and 9.0 percent, respectively, expressed EGFP [P greater than 0.05]. EGFP expression was observed in 100 percent of reconstructed embryos, with 91.2 percent exhibiting homogenic expression. To confirm sperm genome incorporation, androgenetic haploid blastomeres generated by ICSI prior to enucleation and using Y chromosome sexed spermatozoa were used for biparental embryo reconstruction. Incorporation of the Y chromosome was confirmed by polymerase chain reaction and fluorescence in situ hybridisation analysis. In conclusion, the results of the present study prove that it is possible to use sperm genome replicates to reconstruct biparental bovine embryos and that it is a highly efficient technique to generate homogeneous transgene-expressing embryos. 
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700 1 |a Radrizzani Helguera, Martín   |9 58157 
700 1 |9 61021  |a Salamone, Daniel Felipe 
773 |t Reproduction, Fertility and Development  |g Vol.23, no.6 (2011), p.769-779 
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