MAC-T cells as a tool to evaluate lentiviral vector construction targeting recombinant protein expression in milk.
Anim Biotechnol
; 26(2): 136-42, 2015.
Article
em En
| MEDLINE
| ID: mdl-25380466
Prior to generating transgenic animals for bioreactors, it is important to evaluate the vector constructed to avoid poor protein expression. Mammary epithelial cells cultured in vitro have been proposed as a model to reproduce the biology of the mammary gland. In the present work, three lentiviral vectors were constructed for the human growth hormone (GH), interleukin 2 (IL2), and granulocyte colony-stimulating factor 3 (CSF3) genes driven by the bovine ß-casein promoter. The lentiviruses were used to transduce mammary epithelial cells (MAC-T), and the transformed cells were cultured on polystyrene in culture medium with and without prolactin. The gene expression of transgenes was evaluated by PCR using cDNA, and recombinant protein expression was evaluated by Western-blotting using concentrated medium and cellular extracts. The gene expression, of the three introduced genes, was detected in both induced and non induced MAC-T cells. The human GH protein was detected in the concentrated medium, whereas CSF3 was detected in the cellular extract. Apparently, the cellular extract is more appropriate than the concentrated medium to detect recombinant protein, principally because concentrated medium has a high concentration of bovine serum albumin. The results suggest that MAC-T cells may be a good system to evaluate vector construction targeting recombinant protein expression in milk.
Palavras-chave
Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Proteínas Recombinantes
/
Lentivirus
/
Leite
/
Vetores Genéticos
Tipo de estudo:
Prognostic_studies
Limite:
Animals
Idioma:
En
Revista:
Anim Biotechnol
Assunto da revista:
BIOTECNOLOGIA
Ano de publicação:
2015
Tipo de documento:
Article
País de afiliação:
Brasil
País de publicação:
Reino Unido