中文摘要：

目的：构建卡波氏肉瘤病毒（Kaposi′s sarcoma-associated herpesvirus，KSHV）K9基因的重组慢病毒载体，并探究 K9基因编码的病毒干扰素调节因子1（viral IFN regulatory factor 1，vIRF1）对人脐静脉血管内皮细胞（HUVECs）增殖的影响。方法：根据 K9基因的核酸序列设计聚合酶链式反应（PCR）的上下游引物。以实验室已有的真核表达载体 pCI-K9-Flag 为模板，PCR 扩增 K9基因序列。PCR 产物经限制性内切酶双酶切后插入慢病毒载体 pHAGE-CMV-MCS-Izs-Green（简称 pHAGE）中，构建重组慢病毒质粒 pHAGE-K9。将 pHAGE-K9质粒与包膜质粒 pMD2．G、包装质粒 psPAX2共同转染人胚肾上皮细胞（293 T 细胞），包装 K9基因的重组慢病毒。采用病毒梯度稀释法测定病毒滴度。慢病毒感染 HUVECs 后，观察绿色荧光蛋白（GFP）的表达，蛋白质印迹法检测 HUVECs 中 K9编码的 vIRF1蛋白的表达。采用流式分选技术获得稳定表达 vIRF1蛋白的 HUVECs。运用细胞增殖实验检测 vIRF1对 HUVECs 增殖的影响。结果：核酸序列测定结果表明，重组慢病毒质粒 pHAGE-K9构建成功。蛋白质印迹结果显示，K9基因重组慢病毒感染 HUVECs 后其编码蛋白成功表达。细胞增殖实验结果表明，稳定表达 vIRF1的 HUVECs 增殖能力显著强于对照组。结论：KSHV K9基因编码的 vIRF1能够促进 HUVECs 的增殖。

英文摘要：

Objective：To construct the recombinant plasmid carrying K9 gene of Kaposi′s sarcoma-as-sociated herpesvirus（KSHV）,and to explore the effect of K9 encoded protein vIRF1 on the proliferation of human umbilical vein endothelial cells（HUVECs）.Methods：According to the sequence of K9,forward and reverse PCR primers with XhoⅠ and BamHⅠrestriction endonucleases sites were designed.Then the expression plasmid pCI-K9-Flag was used as template to amplify K9.Subsequently,PCR product digested by restriction endonucleases was inserted into pHAGE-CMV-MCS-Izs-Green vector （abbreviated as pHAGE）to construct recombinant plasmid pHAGE-K9.Then pHAGE-K9,package plasmid psPAX2 and envelope plasmid pMD2.G were co-transfected into the 293 T cells to produce lentivirus.The viral titer was determined by gradient dilution.Next,pHAGE-K9 lentivirus infected HUVECs was observed and the expression of vIRF1 protein was detected by Western blotting.Cells expressing vIRF1 were sorted by flow cytometry.Finally,the effect of vIRF1 on the proliferation of HUVECs was measured by CCK-8 assay. Results：Based on sequencing of recombinant plasmid corresponded with K9 gene,it was validated that recombinant plasmid was successful constructed.Western blotting showed lentivirus-infected HUVECs＆nbsp;expressed vIRF1 successfully.The result of CCK-8 showed that compared with control group,proliferation of HUVECs stablely expressing vIRF1 was significantly increased （P 〈0.01,n =5）.Conclusion：KSHV K9 encoded protein,vIRF1,enhanced the proliferation of HUVECs.