Recombinant Expression,Purification and Identification of Humulus Japonicus Pollen Allergen Humj1
Through vector construction,prokaryotic expression,and purification of the major allergen Humj1 from Humulus japonicus pollen,we obtained a recombinant protein of the Humulus japonicus allergen Humj1 with immunoreactivity. After codon optimization,the synthesis of the Humj1 gene from Humulus japonicus designed with primers containing restriction sites was achieved. The PCR amplified product was ligated into the pGEM-T vector to construct the cloning vector. The target fragment was then ligated into the pET-28a expression vector,successfully creating the pET-28a-Humj1 prokaryotic expression vector. The fusion protein was efficiently expressed in E.coli BL21 and purified by Ni-NTA affinity chromatog-raphy. Bioinformatics analysis of the expressed protein confirmed its hydrophilic nature,strong antigenicity,and predomi-nantly disordered coil structure. Western blotting proved that the recombinant protein could bind to the serum of Humulus japonicus allergic patients and had immune activity. This holds significant clinical importance for the future development of monoclonal antibodies against Humulus japonicus allergens and the preparation of desensitizing vaccines.
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