[Cloning, expression and purification of allergen parvalbumin from Aristichthys nobilis and its allergic activity].

OBJECTIVE:To clone, express and identify the parvalbumin gene from Aristichthys nobilis, and investigate its allergenicity. METHODS: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. RESULTS: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. CONCLUSION:The parvalbumin gene from Aristichthys nobilis is successfully cloned and expressed in this study, and the recombinant protein possesses good IgE-binding capacity.