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[目的]构建真核表达载体pEGFP-N1/rCNP并转染体外培养的人脐静脉内皮细胞,观察CNP基因在其中的表达。[方法]利用RT-PCR方法从家兔腹主动脉组织扩增获得CNP基因全场全长编码区,克隆到含有增强型绿色荧光蛋白报告基因的真核表达载体pEGFP-N1上,构建重组载体pEGFP-N1/rCNP。利用LipofectaminTM 2000脂质体将重组质粒转染人脐静脉内皮细胞。[结果]重组质粒pEGFP-N1/rCNP构建成功;将其转染人脐静脉内皮细胞,观察到目的基因CNP表达。[结论]成功构建了家兔腹主动脉CNP基因的真核表达载体pEGFP-N1/rCNP,并可转染人脐静脉内皮细胞。
[Objective] To construct eukaryotic expression vector pEGFP-N1 / rCNP and transfect human umbilical vein endothelial cells in vitro to observe the expression of CNP gene in it. [Method] The full-length coding region of CNP gene was amplified from abdominal aorta tissue of rabbits by RT-PCR and cloned into eukaryotic expression vector pEGFP-N1 containing enhanced green fluorescent protein reporter gene to construct a recombinant vector pEGFP-N1 / rCNP. The recombinant plasmids were transfected into human umbilical vein endothelial cells using LipofectaminTM 2000 liposome. [Result] The recombinant plasmid pEGFP-N1 / rCNP was successfully constructed. The recombinant plasmid pEGFP-N1 / rCNP was transfected into human umbilical vein endothelial cells and the CNP expression of the target gene was observed. [Conclusion] The eukaryotic expression vector pEGFP-N1 / rCNP of rabbit abdominal aorta CNP gene was successfully constructed and transfected into human umbilical vein endothelial cells.