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目的:构建白血病相关基因LRP16真核表达质粒,并检测其在人子宫内膜癌HEC-1-B细胞中的表达。方法:从人子宫内膜癌HEC-1-B细胞中提取总RNA,应用PCR技术,扩增获得LRP16基因编码序列片段,克隆入真核表达载体EX-Y2069-M29,对重组质粒进行酶切和测序鉴定后,以脂质体介导法转染至HEC-1-B细胞,采用Western blot法检测LRP16蛋白的表达。结果:酶切和测序结果证明LRP16基因真核表达质粒EX-Y2069-M29的DNA序列完全正确,将其转染HEC-1-B细胞后,LRP16蛋白表达明显增加。结论:LRP16基因重组真核表达质粒EX-Y2069-M29构建成功,并能在HEC-1-B细胞中表达,为进一步研究LRP16基因奠定了基础。
Objective: To construct a leukemia-associated gene LRP16 eukaryotic expression plasmid and detect its expression in human endometrial carcinoma HEC-1-B cells. Methods: Total RNA was extracted from human endometrial carcinoma cell line HEC-1-B and amplified by PCR. The fragment encoding LRP16 gene was cloned into the eukaryotic expression vector EX-Y2069-M29 and the recombinant plasmid was digested After sequencing, the cells were transfected into HEC-1-B cells by lipofectamine. The expression of LRP16 protein was detected by Western blot. Results: The results of enzyme digestion and sequencing showed that the sequence of LRP16 gene was exactly correct. The expression of LRP16 protein was significantly increased after transfected into HEC-1-B cells. CONCLUSION: The recombinant eukaryotic expression plasmid EX-Y2069-M29 of LRP16 gene was successfully constructed and expressed in HEC-1-B cells, which laid the foundation for further study of LRP16 gene.