中文摘要：

目的：克隆粉尘螨变应原第10组分（Der f 10）基因并建立其原核表达体系。方法：提取粉尘螨总RNA,反转录后经套式PCR扩增出目的基因并克隆至pMD19-T载体测序,将测序正确的目的基因插入表达质粒pET28a,转化E.coliBL21（DE3）用IPTG诱导表达,用SDS-PAGE和Western blot鉴定重组蛋白。结果：RT-PCR扩增获得Der f 10,琼脂糖凝胶电泳见一条约888 bp的条带,与参考序列（GenBank No.EU 106617）同源性高达99.8%。将pET28a（＋）-Der f 10质粒转化宿主菌E.coli BL21,SDS-PAGE电泳时出现特异性条带,Western blot表明重组蛋白可与抗His-Tag Mab抗体结合。生物信息学软件预测获得的Der f 10重组体由295个氨基酸组成的疏水性蛋白,相对分子质量为34 233.1 Da,二级结构包括α-螺旋（91.86%）、延伸主链（1.36%）和无规卷曲（6.78%）,含有5个原肌球蛋白模序分别位于84～101、120～140、145～173、175～198和231～256氨基酸处。结论：获得了Der f 10编码基因,成功建立其原核表达体系,为生产重组变应原用于临床诊断与治疗奠定基础。

英文摘要：

Objective：To clone the gene encoding of group 10 allergen of Dermatophagoides farinae（Der f 10）,and to develop its prokaryotic expression system.Methods：The total RNA of Dermatophagoides farinae was extracted,and the gene encoding Der f 10 was amplified and inserted into pMD19-T simple vector.After sequencing,the target gene was then inserted into the expression plasmid pET28a,transformed into E.coli BL21（DE3）,induced by IPTG,and identified by SDS-PAGE and Western blot.Results：The gene encoding Der f 10 was amplified by RT-PCR,which had 99.8% similarity to reference sequence.The expression plasmid pET28a（＋）-Der f 10 was transformed into E.coli BL21,and a specific band was observed on SDS-PAGE and Western blot with antibody His-Tag Mab.By analysis of bioinformatics software,the recombinant protein rDer f 10 was composed with 295 hydrophobic residues.The secondary structure of Der f 10 appears to comprise alpha helix（91.86%）,extended strand（1.36%）,and random coils（6.78%）.Additionally,five tropomyosin motifs were predicted at 84-101,120-140,145-173,175-198 and 231-256 amino acid position.Conclusion：The gene encoding Der f 10 was obtained,and a prokaryotic expression system was developed,which lay the groundwork for future studies,including large-scale production of recombinant Der f 10 allergen for diagnostic and therapeutic agents.