甜荞麦16 kDa过敏原基因的克隆及其原核表达载体的构建

Cloning and prokaryotic expression vector construction of common buckwheat 16 kDa major allergen gene

中文关键词:  甜荞麦  克隆  16 kDa过敏原  原核表达载体

英文关键词:common buckwheat  clone  16 kDa allergen  prokaryotic expression vector

中文摘要:

      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16 kDa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16 kDa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16 kDa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450 bp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16 kDa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。

英文摘要:

      Objective To study the molecular biology of allergens from common buckwheat. Methods In this paper, the RT-PCR was applied to clone the full-length allergen genes from common buckwheat and the sequences were analyzed, and then the specific primers were designed. At last, the ORF of 16 kDa allergen gene of common buckwheat was subcloned into the prokaryotic expression vector pET 28a. Results The cDNA sequence coding for this allergenic protein contained 450 bp and encoded 149 aa. Sequence analysis showed that this clone shared high identities with 16 kDa allergen gene from common buckwheat. The deduced protein was therefore regarded as the major allergen of 16 kDa of common buckwheat (GenBank database entry No. EU883600). Conclusion This research lay a foundation for the recombinant expression of allergen protein and the diagnosis of allergic diseases.