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目的 构建融合基因sOVA linker β2m ,探讨通过内源性途径递呈的肽在细胞表面形成MHCⅠ类分子复合物的情况。方法 构建了真核表达载体pcDNA3/sOVA linker β2m及不含有 β2m基因的对照载体 ,以研究 β2m是否能在此过程中起到辅助的作用。结果 RT PCR与原位杂交结果显示在各转导细胞中均有融合基因的正确表达 ,且二者的表达水平差异无显著性 (P >0 .0 5 )。进一步对细胞内、细胞表面H 2Kb OVA复合物的分布进行分析 ,表明各转导细胞中均有正确的蛋白质分子翻译合成 ,并能表达在细胞表面 ,但融合有 β2m基因的EL4 /eb2m细胞的表达量远远高于没有融合 β2m基因的转导细胞EL4 /OVA。结论 β2m基因与特异性抗原肽的融合 ,易于形成稳定的MHCⅠ类分子复合物 ,提高抗原呈递效率。
Objective To construct the fusion gene sOVA linker β2m to study the formation of MHC class I complex on the cell surface by peptides presented by endogenous pathway. Methods The eukaryotic expression vector pcDNA3 / sOVA linker β2m and the control vector without β2m gene were constructed to investigate whether β2m can play an auxiliary role in this process. Results The results of RT-PCR and in situ hybridization showed that the fusion gene was correctly expressed in all transduced cells, and there was no significant difference between the two expression levels (P> 0.05). Further analysis of intracellular and cell surface distribution of H 2Kb OVA complex showed that all transduced cells had the correct translational synthesis of protein molecules and could be expressed on the cell surface but the EL4 / eb2m cells fused with β2m gene The expression level is much higher than the EL4 / OVA transduced cells without β2m gene fusion. Conclusion The fusion of β2m gene and specific antigen peptide is easy to form a stable MHC class Ⅰ molecular complex and improve the efficiency of antigen presentation.