携带鼠白细胞介素-12基因重组腺病毒载体的构建及其对致瘤性的影响

目的 :构建携带鼠白细胞介素 - 1 2基因腺病毒载体 ( Ad CMVIL- 1 2 )并观察其影响 B1 6黑色素瘤致瘤性的效果。方法 :将鼠 IL- 1 2基因构建进含有 CMV启动子的双顺反子重组腺病毒载体内 ,分离纯化后 ,转染 2 93细胞 ,48h后提取细胞总 RNA,进行 RT- PCR、琼脂糖凝胶电泳 ,选取特异性片段 TA克隆、测序 ;将 B1 6黑色素瘤细胞与 Ad CMVIL- 1 2在 37℃、5 %CO2 培养箱中孵育2 h后注射到 C57/ BL6小鼠右侧腹股沟皮下 ( MOI=1 0 0 ) ,观察 Ad CMVIL- 1 2对 B1 6黑色素瘤致瘤性的抑制作用。结果 :测序结果证明 Ad CMVIL- 1 2构建成功 ,实验组未见肿瘤生长 ,抑瘤率达1 0 0 %。结论 :成功地构建 Ad CMVIL- 1 2 ,并具有明显的抑制肿瘤生长的作用。

Construction of Recombinant Adenovirus Vector Carrying Murine Interleukin-12 Gene and the Experimental Study of Its Inhibitory Effect on B16 Melanoma

Objective: To construct AdCMVIL 12 and observe its inhibit effect on B16 melanoma. Methods: Murine IL 12 gene was inserted into a bicistronic recombinant adenovirus vector contains CMV promoter, then the virus was separated and purified. After AdCMVIL 12 transferred 293 cells, the total RNA of cell was extracted to make RT PCR after 48 hours, did the agarose gel electrophoresis, the specific fragment was selected for sequencing. B16 melanoma cells were incubated with AdCMVIL 12 (MOI=100) at 37℃ 5% CO 2 for 2 hours, then the cells were injected subcutaneous at right Groin of C 57 /BL 6 mice, observes the inhibit effect of AdCMVIL 12 on B16 melanoma. Results: The result of sequencing verified AdCMVIL 12 was constructed successfully, the experimental group had no tumor growth, inhibit percent of tumor was 100%. Conclusion: The construction of AdCMVIL 12 was successful, and the inhibit effect on tumor growth is obvious.