关键词: Ara h 1过敏原, 重组蛋白, 致敏性, BALB/c小鼠, RBL细胞

Abstract: An Ara h 1 gene was synthesized from total RNA extracted from peanut using oligo primers by reverse transcription polymerase chain reaction (RT-PCR). The gene was cloned into pET-28a vector and a recombinant expression vector was constructed. Then the recombinant vector was transformed into the bacterial strain Rosetta (DE3) for protein expression, and the target protein was purified by Ni affinity chromatography. Sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) showed that the molecular mass of the expressed protein was about 75 kDa, which matched with the theoretical value. Analysis by mass spectrometry (MS) confirmed that the protein was Ara h 1. The allergenicity of recombinant Ara h 1 protein was evaluated using an animal model of BALB/c mice. The results showed that recombinant Ara h 1 protein had a similar allergenicity to the native one. The contents of specific antibody, Th2 type cytokines and histamine in the serum were increased, and inflammation occurred in the lung and jejunum of mice sensitized to recombinant Ara h 1. Meanwhile, recombinant Ara h 1 could trigger degranulation of RBL cell, resulting in release of β-hexosaminidase. Accordingly, recombinant Ara h 1 could cause Th2 type allergic reaction.

Key words: Ara h 1 allergen, recombinant protein, allergenicity, BALB/c mice, RBL cell