Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 1 de 1
Filtrar
Adicionar filtros








Intervalo de ano
1.
Chinese Journal of Biotechnology ; (12): 297-304, 2010.
Artigo em Chinês | WPRIM | ID: wpr-336228

RESUMO

It is one of the frequently utilized strategies for positive-negative selection to elevate the gene targeting efficiency in somatic cells by enriching targeted colonies. Knocking out prnp in animals by gene targeting can prevent it from expressing Prion protein (Pathogenic protein of transmissible spongiform encephalopathy), which enables it to resist infection of Prion. We constructed a bovine prnp biallelic targeting vector via the positive-negative selection strategy, and transfected the linearized vector into the bovine fetal fibroblasts through electroporation. Then, we selected cells in cell culture medium with G418 under a concentration of 600 microg/mL followed by Ganciclovir (GCV) under a concentration of 200 nmol/mL. In the end, we successfully obtained 176 cell clones. All these clones were identified by means of sequencing, immunofluorescence and western blotting, respectively, confirming that there existed 9 positive cell clones. The results showed that the bovine prnp gene was successfully knocked out. Conclusively, we provide an effective way to knockout bovine prnp gene, which could serve as the basis for producing prion protein gene knockout transgenic cloned cattle.


Assuntos
Animais , Bovinos , Eletroporação , Encefalopatia Espongiforme Bovina , Genética , Feto , Biologia Celular , Fibroblastos , Biologia Celular , Metabolismo , Técnicas de Inativação de Genes , Métodos , Marcação de Genes , Vetores Genéticos , Genética , Príons , Genética , Transfecção
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA