目的 构建含早期生长应答因子1 （Egr1）辐射敏感启动子调控的人钠碘同向转运体（hNIS）基因重组杆状病毒，为进一步延长核素在细胞中的滞留时间，提高hNIS介导的恶性肿瘤放射性核素基因靶向治疗疗效提供理论基础。方法 空载体pFB取自质粒pFB-CMV-EGFP，Egr1启动子取自质粒PGL3-Egr1-luc，hNIS取自质粒pcDNA3.1-CMV-hNIS，构建重组杆状病毒载体质粒pFB-Egr1-Hnis；同时构建含CMV启动子的质粒pFB-CMV-hNIS作为阳性对照，并将杆状病毒空载体质粒pFB-0作为阴性对照。随后制备各组杆状病毒，扩增收集重组杆状病毒并进行滴度测定。为进一步鉴定病毒功能，一方面通过体外感染U87脑胶质瘤细胞并采用¹³¹I刺激，通过免疫荧光检测¹³¹I刺激后的hNIS蛋白表达；另一方面通过摄碘实验进一步验证所表达的hNIS蛋白的摄碘功能和特性。结果 成功构建重组杆状病毒载体质粒pFB-Egr1-hNIS和pFB-CMV-hNIS；转座成功并提取了重组Bacmid；重组Bacmid转染Sf 9细胞后扩增收集的重组杆状病毒滴度可以达到1×10¹⁰ pfu/mL。体外感染U87细胞后，免疫荧光检测表明¹³¹I刺激后可增加hNIS的表达，感染细胞表达的hNIS蛋白位于细胞膜上，且具有摄碘的功能。结论 成功构建了重组杆状病毒载体质粒pFB-Egr1-hNIS及pFB-CMV-hNIS，获得高滴度的重组杆状病毒Bac-Egr1-hNIS、Bac-CMV-hNIS和Bac-0并得到验证；建立了基于hNIS基因的辐射正反馈重组杆状病毒体系，为进一步提高核素靶向治疗疗效奠定了重要的理论基础。

Objective To prepare and verify a recombinant baculovirus harboring the human sodium iodide symporter (hNIS) under control of the early growth response-1 (Egr-1) promoter, and to provide the foundation for the research on radionuclides gene targeted treatment of malignant tumors. Methods The vector pFB was cut from plasmid pFB-CMV-EGFP; Egr1 was cut from plasmid PGL3-Egr1-luc; and hNIS was cut from plasmid pcDNA3.1-CMV-hNIS. Then recombinant baculovirus plasmid pFB-Egr1-hNIS was constructed. In addition, a positive control plasmid pFB-CMV-hNIS and a control plasmid pFB-0 were constructed. Baculovirus Bac-Egr1-hNIS and its controls were prepared, verified, amplified, and collected for titer determination. After glioma cell line U87 was infected with Bac-Egr1-hNIS and exposed to ¹³¹I, the presence and enhanced expression of hNIS protein was tested by immunofluorescence using anti-hNIS antibody. To verify the function of hNIS after ¹³¹I stimulation, the uptake of iodide was determined after incubation of the processed cells with ¹²⁵I. Results Recombinant plasmid pFB-Egr1-hNIS, pFB-CMV-hNIS, and pFB-0 were correctly constructed and confirmed by DNA sequencing. Transposition was correct and recombinant Bacmid was extracted. The recombinant baculovirus were packed, verified, and its titer was about 1×10¹⁰ Pfu/mL. After incubation with 3 MBq/mL ¹³¹I, immunofluorescence showed that the hNIS expression level of Bac-Egr1-hNIS infected U87 cells increased dramatically. In another side, ¹³¹I stimulated U87 cells accumulated more ¹²⁵I than did non stimulated cells. Conclusion The recombinant baculovirus Bac-Egr1-hNIS and its controls were constructed and verified, and high titer was obtained. This study provides a foundation of radiosensitive positive feedback system for further study.