High-level expression of fusion protein GGH in Pichia pastoris GS115 by constructing a double plasmid co-expression system / 生物工程学报
Chinese Journal of Biotechnology
;
(12): 983-989, 2011.
Article
in Chinese
| WPRIM
| ID: wpr-324512
ABSTRACT
In order to make a large-scale preparation of(GLP-1A2G)2-HAS (GGH), the double-plamid pPICZalphaB and pPIC9K co-expression system was introduced into Pichia pastoris GS115. Firstly, the GGH fusion gene was amplified by PCR to create the recombinant expression plasmid pPICZalphaB-ggh, which was transformed into P. pastoris GS 115/F2 that was integrated by another recombinant expression plasmid pPIC9K-ggh. The immunology method combined with high concentration antibiotic was used to screen recombinant strain P. pastoris GS115/F3 capable of high-level expression of GGH protein. The GGH fusion protein expressed by GS115/F3 increased 49.7% compared with the GS 115/F2 in the expression conditions of 3% methanol inducing 80 h at 30 degrees C. At the same time, the quantitative PCR results showed that GGH gene dose in GS115/F3 increased 26.7% with respect to that of GS 115/F2. Furthermore, the Western blotting experiment indicated that the recombinant GGH possess the two antigenicities of GLP-1 and HSA.
Full text:
Available
Index:
WPRIM (Western Pacific)
Main subject:
Pichia
/
Plasmids
/
Recombinant Fusion Proteins
/
Serum Albumin
/
Polymerase Chain Reaction
/
Glucagon-Like Peptide 1
/
Genetic Vectors
/
Genetics
/
Metabolism
/
Methods
Limits:
Humans
Language:
Chinese
Journal:
Chinese Journal of Biotechnology
Year:
2011
Type:
Article
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