Cloning, expression and purification of allergen parvalbumin from Aristichthys nobilis and its allergic activity
Wang, H.; Li, L.; Liu, Z.; Wu, Y.; Chen, J.
Wei Sheng Yan Jiu 40(5): 555-558
2011
ISSN/ISBN: 1000-8020 PMID: 22043700 Document Number: 650905
To clone, express and identify the parvalbumin gene from Aristichthys nobilis, and investigate its allergenicity. The parvalbumin gene was amplified by RT-PCR and cloned into PMD18-T for sequencing and analysis. Then the target gene was subcloned into pET-28a (+) for expression in E. coli BL21 (DE3) by IPTG induction. The recombinant protein was purified by metal (Ni2+) chelating affinity chromatography. Its allergenicity was examined by Western-blotting assay. The length of gene (Accession No. FJ013047) was 330 bp, coding 109 amino acids. The E. coli strain could express a recombinant protein with a molecular weight of 11 537 Da. The recombinant allergen was identified as its affinity to specific IgE antibodies from the allergic patient sera by Western-blotting. The parvalbumin gene from Aristichthys nobilis is successfully cloned and expressed in this study, and the recombinant protein possesses good IgE-binding capacity.