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.