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目的 从感染HTLV-1的中国患者外周血单核细胞（PBMCs）中扩增编码HTLV-1核心蛋白P24的cDNA,并在大肠杆菌中进行表达。方法 提取感染者PBMCs基因组DNA,应用巢式PCR方法扩增出HTLV-1核心蛋白P24的cDNA并测序,与pGEX-20T载体构建重组质粒,在大肠杆菌BL21中表达,采用ELISA和Westernblot分析重组蛋白活性。结果HTLV-1核心蛋白P24基因序列高度保守,构建重组载体后,在大肠杆菌中表达的重组蛋白,经检测具有较强的抗原活性。结论 成功地表达了HTLV-1型病毒的核心蛋白P24基因,为国产诊断试剂和疫苗的研发打下了基础。
OBJECTIVE: To amplify the cDNA encoding the HTLV-1 core protein P24 from Chinese patients with peripheral blood mononuclear cells (PBMCs) infected with HTLV-1 and express in E. coli. Methods The genomic DNA of infected PBMCs was extracted. The cDNA of HTLV-1 core protein P24 was amplified by nested PCR and sequenced. The recombinant plasmid pGEX-20T was constructed and expressed in E. coli BL21. The recombinant protein was analyzed by ELISA and Western blot active. Results The sequence of P24 gene of HTLV-1 core protein was highly conserved. After constructing the recombinant vector, the recombinant protein expressed in Escherichia coli had strong antigenic activity. Conclusion The P24 gene of core protein of HTLV-1 was successfully expressed, which laid the foundation for the development of domestic diagnostic reagents and vaccines.