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
在 Zh
| WPRIM
| ID: wpr-302911
Responsible library:
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.
全文:
1
索引:
WPRIM
主要主题:
Pichia
/
Transduction, Genetic
/
Neuropeptide Y
/
Recombinant Fusion Proteins
/
Polymerase Chain Reaction
/
Cloning, Molecular
/
Gene Fusion
/
Tat Gene Products, Human Immunodeficiency Virus
/
Genetic Vectors
/
Genetics
研究类型:
Prognostic_studies
限制:
Animals
语言:
Zh
期刊:
Chinese Journal of Biotechnology
年:
2008
类型:
Article