Lipofection of siRNA into bovine 8-16-cell stage embryos using zona removal and the well-of-the-well culture system.
J Reprod Dev
; 64(2): 199-202, 2018 Apr 13.
Article
em En
| MEDLINE
| ID: mdl-29353869
ABSTRACT
Bovine preimplantation embryos exhibit dramatic biological changes between before and after the 8-16-cell stage. Here we report a simple lipofection method to transfect siRNA into bovine 8-16-cell stage embryos using zona removal and the well-of-the-well (WOW) culture system. Bovine one-cell embryos produced in vitro were freed from the zona pellucida and cultured up to the 8-16-cell stage in WOW dishes. The 8-16-cell embryos were lipofected with siRNA and the transfection efficiency was assessed at 48 h of transfection. Lipofection with a red fluorescent non-targeting siRNA revealed the importance of zona removal for transfection of siRNA into embryos. Using this method, we knocked down the methionine adenosyltransferase 2A (MAT2A) gene, achieving a significant reduction in MAT2A expression (P < 0.05) concomitant with the marked inhibition of blastocyst development. Our proposed method, tentatively named 'Octo-lipofection', may be useful to analyze gene functions in bovine preimplantation embryos without expensive equipment and skill-intensive techniques.
Palavras-chave
Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Transfecção
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RNA Interferente Pequeno
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Interferência de RNA
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Ectogênese
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Mórula
Limite:
Animals
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Female
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Humans
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Male
Idioma:
En
Revista:
J Reprod Dev
Assunto da revista:
MEDICINA REPRODUTIVA
Ano de publicação:
2018
Tipo de documento:
Article
País de afiliação:
Japão