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目的获得粉尘螨变应原第3组分(Der f 3)的编码基因并了解其序列多态性。方法根据GenBank已公布的Der f 3核酸序列设计引物,用RT-PCR扩增获得其编码基因,插入pMD19-T载体,进行序列测定和多态性分析。结果获得粉尘螨变应原Der f 3编码基因为780bp。5株Der f 3基因cDNA克隆的核苷酸序列与参考序列(GenBankNo.AY 283291,GenBank No.D63858,GenBank No.EU312162,GenBank No.EU312163)相比,有19处cDNA序列发生突变,但有7处突变至少在3个cDNA克隆中序列一致;推导出氨基酸序列后进行比对,有11处氨基酸序列发生变化,有4处与上述7个位点相一致。结论获得了粉尘螨变应原Der f 3编码基因且证明其序列具有多态性,为进一步开展相关研究奠定了基础。
Objective To obtain the coding gene of der f 3 allergen and to understand its sequence polymorphism. Methods Based on the published Der f 3 nucleic acid sequence in GenBank, primers were designed and their coding genes were amplified by RT-PCR and inserted into pMD19-T vector for sequence analysis and polymorphism analysis. Results Der f 3 gene of Dermatophagoides farinae allergen was 780bp. The nucleotide sequence of the 5 cDNA clones of the Der f 3 gene was mutated in 19 cDNA sequences compared to the reference sequence (GenBank No. A 283291, GenBank No. D63858, GenBank No. EU312162, GenBank No. EU312163) Seven of the seven mutations were consistent in at least three cDNA clones. After amino acid sequence deduced, 11 amino acid sequences were changed, and four of them were consistent with the above seven loci. Conclusion Der f 3 gene encoding Dermatophagoides pteronyssinus was obtained and its sequence was proved to be polymorphic, which laid the foundation for further research.