结肠癌是近年来在中国增长迅速、对社会和人民造成严重负担的新兴癌症，现有治疗手段的局限性如手术切除的易复发性、化疗与靶向药物的靶点易突变性、免疫治疗的冷肿瘤无效性等使得寻找结肠癌治疗的新方式、新靶 点刻不容缓。

      本文以小鼠结肠癌细胞 MC38 作为研究对象，通过对潜在治疗靶点 XX 基因及其胞内外共六个片段 X1~X6 的过表达，探究出 XX 基因及其六个片段 在 MC38 细胞发生发展的具体作用。首先，我们从 C57BL/6 小鼠骨髓细胞提 取总 RNA 并逆转录获得 cDNA 模板，通过设计引物成功扩增出 XX 基因；再 通过双酶切连接将 XX 基因克隆在慢病毒过表达穿梭质粒 pLVX-IRES-Puro， 将穿梭质粒和辅助质粒在人胚肾细胞HEK 293T细胞成功包装出XX基因过表 达慢病毒；最后我们将 XX 基因过表达的慢病毒感染 MC38 细胞后进行体内 荷瘤实验和体外增殖实验，成功探究出 XX 基因过表达可以显著抑制 MC38 细胞的增殖并提高荷瘤小鼠的存活时间。同时我们也将 XX 基因过表达慢病 毒感染了黑色素瘤细胞 B16-F10，发现其并没有抑制 B16-F10 增殖，荷瘤小 鼠的存活时间和对照组也没有差异，说明 XX 基因过表达的抑制作用可能只 对结肠癌细胞起作用。这也揭示针对不同肿瘤间发病机制或转移的特异性时， 我们需要找出特异的靶点并研制出对应的治疗手段。

       我们也探究了 XX 基因起作用的独立结构域片段。我们先设计引物分别扩增出 XX 基因的六个片段 X1~X6，再将 X1~X6 六个片段分别克隆到穿梭质粒 pLVX-IRES-Puro 进行慢病毒包装，最后我们将 X1~X6 分别过表达的慢病 毒感染 MC38 进行体外增殖实验，成功探究出可能起调控作用的片段为胞外 片段 X3。

      本实验通过探究 XX 基因过表达抑制结肠癌细胞 MC38 的增殖，并找出 XX 基因可能起具体调控作用的胞外片段 X3，既为探明 XX 基因调控结肠癌 的具体分子机制奠定坚实的基础，也为结肠癌靶向治疗提供新的方向和思路。

       Colon cancer is an malignant tumor that has grown rapidly in China recently and has caused a serious burden on the society and the people. The limitations of existing treatment methods, such as the susceptibility to recurrence of surgical resection, the mutations of target chemotherapy and targeted drugs, and the ineffectiveness of cold tumors in immunotherapy makes it urgent to find new ways and new targets for colon cancer treatment.

      In this paper, the mouse colon cancer cell line MC38 was taken as the research object, and the potential therapeutic target XX gene and its six fragments X1~X6 were overexpressed to explore the role of XX gene and its six fragments X1~X6 in the proliferation process of MC38 cells. First, we extracted total RNA from C57BL/6 mouse bone marrow cells and reverse-transcribed it to obtain a cDNA template, and we successfully amplified the XX gene by designing primers. Then we cloned the XX gene into the lentiviral overexpression shuttle plasmid pLVX- IRES-Puro and successfully produced XX gene overexpression lentivirus in human embryonic kidney cell HEK 293T cells by using shuttle plasmid and helper plasmid. Finally, we infected MC38 cells with XX gene overexpressed lentivirus, and successfully explored in subsequent in vivo tumor-bearing experiments and in vitro proliferation experiments that XX gene overexpression can significantly inhibit the proliferation of MC38 cells and increase the survival time of tumor-bearing mice. At the same time, we also infected melanoma cell B16-F10 with XX gene overexpression lentivirus, and found that it did not inhibit the proliferation of B16-F10, and the survival time of tumor-bearing mice had no difference with the control group, indicating that the inhibitory effect of XX gene overexpression may only be on colon cancer. This also reveals that when we aim at the specificity of the pathogenesis of different tumors, we need to find out their specific targets and develop corresponding treatments.

      We also explored the domain-independent fragments in which the XX gene functions. We first designed primers to amplify the six fragments X1~X6 of the XX gene, and then cloned the six fragments X1~X6 into the shuttle plasmid pLVX-IRES-Puro for lentiviral packaging. Finally, we infect MC38 with overexpressed lentiviruses of X1~X6 for in vitro proliferation experiments, and successfully explored that the fragment that may play a regulatory role is the extracellular fragment X3.

        In this experiment, we have proved the overexpression of XX gene inhibits the proliferation of colon cancer cell MC38, and find out the fragment X3 of XX gene that may play a specific regulatory role. It not only lays a solid foundation for exploring the specific molecular mechanism of XX gene regulation of colon cancer, but also provides new directions and ideas for targeted therapy of colon cancer.