Abstract:

Ara h 2 is one of paramount allergens in peanut. The goal of the present study was to biosynthesize recombinant peanut allergen Ara h 2. Peanut cDNA was synthesized from total RNA using Oligo primers by RT-PCR in order to provide a template for the PCR amplification of Ara h 2 gene. The purified amplification products were cloned into the pMD19-T simple vector to construct a recombinant vector carrying Ara h 2 gene, named pMD19-T-Ara h 2. The recombinant plasmid was digested by Nco I and Hind III. The purified digestion products were then ligated to the pGEX-4T-1 expression vector and transformed into the BL21-codonPlus(DE3)-RIPL for 24 h expression under IPTG induction. The target product, GST-Ara h 2 fusion protein, was purified with Glutathione Sepharose 4B. The results of SDS-PAGE and western-blotting showed that GST-Ara h 2 fusion protein was 46 kD in size, with 90% purity and could specifically react with anti-Ara h 2 sera from rabbits, indicating that the recombinant protein has high specificity of immune reaction.

Key words: food allergy, peanut allergen Ara h 2, biosynthesis