Cloning of PTD-NPY fusion gene and its secretory expression in Pichia pastoris / 生物工程学报
Chinese Journal of Biotechnology
; (12): 1620-1624, 2008.
Article
em Zh
| WPRIM
| ID: wpr-302911
Biblioteca responsável:
WPRO
ABSTRACT
The PTD-NPY fusion gene derived from HIV-1 TAT protein transduction domain and rat neuropeptide Y was amplified by overlap extension PCR, digested and subcloned into yeast expression vector pPICZ alpha A to construct recombinant expression plasmid pPICZ alpha-PTD-NPY. The cloned PTD-NPY fusion gene was identified by PCR and restriction enzyme digestion and sequenced. The exact recombinant plasmid was linearized by Sac I and integrated by electrotransformation into the genome of Pichia pastoris GS115 cells. Then, these positive recombinant yeast cells were induced by 10 mL/L methanol to express soluble PTD-NPY fusion protein. After 120 h of methanol induction, the SDS-PAGE electrophoresis result indicated PTD-NPY fusion protein was efficiently secreted into the medium. Western blotting analysis proved that the expressed fusion protein had specific NPY binding activity. The successful expression of PTD-NPY fusion protein in Pichia pastoris provided basis for its further application study.
Texto completo:
1
Índice:
WPRIM
Assunto principal:
Pichia
/
Transdução Genética
/
Neuropeptídeo Y
/
Proteínas Recombinantes de Fusão
/
Reação em Cadeia da Polimerase
/
Clonagem Molecular
/
Fusão Gênica
/
Produtos do Gene tat do Vírus da Imunodeficiência Humana
/
Vetores Genéticos
/
Genética
Tipo de estudo:
Prognostic_studies
Limite:
Animals
Idioma:
Zh
Revista:
Chinese Journal of Biotechnology
Ano de publicação:
2008
Tipo de documento:
Article