In vitro development of cloned bovine embryos produced by handmade cloning using somatic cells from distinct levels of cell culture confluence


Autoria(s): GERGER, R. P. C.; RIBEIRO, E. S.; FORELL, F.; BERTOLINI, L. R.; RODRIGUES, J. L.; AMBROSIO, C. E.; MIGLINO, M. A.; MEZZALIRA, A.; BERTOLINI, M.
Contribuinte(s)

UNIVERSIDADE DE SÃO PAULO

Data(s)

18/04/2012

18/04/2012

2010

Resumo

The relationship between the level of cell confluence near the plateau phase of growth and blastocyst yield following somatic cell cloning is not well understood. We examined the effect of distinct cell culture confluence levels on in vitro development of cloned bovine embryos. In vitro-matured bovine oocytes were manually bisected and selected by DNA staining. One or two enucleated hemi-cytoplasts were paired and fused with an adult skin somatic cell. Cultured skin cells from an adult Nellore cow harvested at three distinct culture confluence levels (70-80, 80-90, and > 95%) were used for construction of embryos and hemi-embryos. After activation, structures were cultured in vitro as one embryo (1 x 100%) or as aggregates of two hemi-embryos (2 x 50%) per microwell. Fusion, cleavage and blastocyst rates were compared using the chi(2) test. The fusion rate for hemi-embryos (51.4%) was lower than for embryos (67.6%), with no influence of degree of cell confluence. However, blastocyst rates improved linearly (7.0, 17.5, and 29.4%) with increases in cell confluence. We conclude that degree of cell culture confluence significantly influences subsequent embryo development; use of a cell population in high confluence (> 90%) for nuclear transfer significantly improved blastocyst yield after cloning.

FAPESP[06/53822-4]

CAPES/Brazil[0192058]

PAP/CAV-UDESC[2007-2008]

Identificador

GENETICS AND MOLECULAR RESEARCH, v.9, n.1, p.295-302, 2010

1676-5680

http://producao.usp.br/handle/BDPI/15557

10.4238/vol9-1gmr690

http://dx.doi.org/10.4238/vol9-1gmr690

Idioma(s)

eng

Publicador

FUNPEC-EDITORA

Relação

Genetics and Molecular Research

Direitos

openAccess

Copyright FUNPEC-EDITORA

Palavras-Chave #Somatic cell nuclear transfer #Cell culture confluence #Cell cycle #Embryo aggregation #Cattle #PORCINE FETAL FIBROBLASTS #NUCLEAR TRANSFER #CYCLE SYNCHRONIZATION #SERUM STARVATION #G0/G1 PHASE #ROSCOVITINE #CATTLE #AGGREGATION #DEPRIVATION #GRANULOSA #Biochemistry & Molecular Biology #Genetics & Heredity
Tipo

article

original article

publishedVersion