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目的对甜荞麦(common buckwheat)过敏原的分子生物学开展研究。方法通过RT-PCR克隆甜荞麦16kDa过敏原蛋白的全长基因,并根据序列设计带有酶切位点的特异性引物,扩增甜荞麦16kDa过敏原基因的完整开放阅读框,与pET-28a载体连接,构建原核表达载体。结果本研究成功克隆了甜荞麦16kDa过敏原蛋白的基因,且构建了其原核表达载体。该基因含有长度为450bp的开放阅读框,编码149个氨基酸,在GenBank数据库中的登录号为EU883600,同源性分析发现其与数据库中已知的荞麦过敏原基因有高度的同源性。结论本研究为甜荞麦16kDa过敏原蛋白的重组表达和临床过敏性疾病的诊断奠定了基础。
Objective To study the molecular biology of common buckwheat allergens. Methods The full-length gene of buckwheat 16kDa allergen was cloned by RT-PCR and the complete open reading frame (ORF) of the 16kDa allergen was amplified according to the designed sequence. Vector to construct prokaryotic expression vector. Results The 16kDa allergen protein gene was successfully cloned and the prokaryotic expression vector was constructed. The gene contains an open reading frame of 450 bp and encodes a protein of 149 amino acids. Its accession number in the GenBank database is EU883600. Homology analysis revealed a high degree of homology with the known buckwheat allergen gene in the database. Conclusion The present study laid the foundation for the recombinant expression of 16 kDa allergen and the diagnosis of clinical allergic diseases in sweet buckwheat.