Cloning and Prokaryotic Expression of Human Recombinant Calreticulin / 中国生物工程杂志
China Biotechnology
;
(12)2006.
Article
in Chinese
| WPRIM
| ID: wpr-685826
ABSTRACT
Objective:
Clone, express and purify human recombinant calreticulin (CRT).Methods:
Human CRT cDNA was amplified from total RNA of human lung cancer cell line A549 cells by RT-PCR. Then, PCR product was subcloned into prokaryotic expression vector pET-15b. After sequencing, this recombinant plasmid was transformed into E.coli. Rossetta. Recombinant CRT was expressed in host cells by IPTG induction. Resulted protein was purified by Ni-NTA resin under denature condition and dialyzed to recover its native structure. SDS-PAGE and Western blot method were used to identify the expression and purification of reconbinant CRT.Results:
Human CRT cDNA was cloned from total RNA of A549 cells. CRT prokaryotic expression vector pET-15b-crt was constructed. Reconbinant CRT was induced to express in E.coli and purified by Ni-NTA affinity chromatograph.Conclusion:
A method for prokaryotic expression and purification of human recombinant CRT was successfully established. This method laid a foundation for the subsequent CRT research.
Full text:
Available
Index:
WPRIM (Western Pacific)
Language:
Chinese
Journal:
China Biotechnology
Year:
2006
Type:
Article
Similar
MEDLINE
...
LILACS
LIS