人甲状腺钠/碘转运体基因全长的克隆及其重组表达质粒的构建

Study on full length gene cloning of human sodium/iodide symporter and the construction of its expression plasmid

目的:利用基因工程的方法制备甲状腺钠/碘同向转运体(hNIS)cDNA,为今后的转基因治疗奠定物质基础。方法:采用TRIzol一步法,从甲状腺组织中提取总RNA,再应用RT-PCR扩增出hNIS基因全长,然后采用TOPO克隆技术构建重组表达质粒pcDNA3.1D/FLhNIS,并进行酶切和测序鉴定。结果:成功制备了hNIScDNA并构建了其重组表达质粒。结论:为最终实现131I治疗不摄碘的肿瘤提供了分子生物学基础。

Objective： To provide an objective evidence for transferring gene radioiodine therapy in nonthyroid tumor, build human sodium/iodide symporter（hNIS）cDNA by gene engineering. Methods：Total RNA was isolated from the thyroid tissue sample of Graves disease patient by TRIzol reagent, the hNIS gene was amplified by RT-PCR, the target gene were inserted into vector pcDNA3.1/D-V5-His to construct the recombinant expressive plasmid pcDNA3.1D/FLhNIS by TOPO clone methods, then restrictive enzyme digested and sequenced. Results： Successful building hNIS cDNA and its expressive plasmid. Conclusion： This method provides an objective molecular biologic evidence for radioiodine therapy in nonthyroid tumor.