人Polo样激酶1基因的克隆表达及其影响c-Abl表达水平功能的初探

摘要

Objective: To clone and express human Polo-like kinase-1(Plk1) gene in 293 cell lines, and study its effect on the expression of non-receptor tyrosine kinase c-Abl. Methods: Plk1 gene was amplified by PCR, and then inserted into eukaryotic expression vector pcDNA3-Flag and pCMV-Myc respectively. Those recombinant expression plasmids were transiently transfected into 293 cell lines, and the expression of Plk1 was identified by Western blotting. Different tagged Plk1 and c-Abl were cotransfected into 293 cells to identify the role of Plk1 on the expression of c-Abl kinase. Results: The Flag-Plk1 and Myc-Plk1 eukaryotic expression plasmids were con⁃structed, and expressed in 293 cell lines with the molecular weight of 68 kD. The expression of c-Abl reduced significantly when contrandfected with tagged Plk1 or c-Abl plasmids into 293 cells. Conclusion: Flag-Plk1 and Myc-Plk1 have been expressed in 293 cells, and it was found that Plk1 can inhibit expression of c-Abl.%　　目的：克隆并在293细胞中表达人Polo样激酶1（Plk1）基因，探索Plk1对非受体型酪氨酸激酶c-Abl表达水平的影响。方法：利用PCR法扩增Plk1基因，分别定向克隆至pcDNA3-Flag及pCMV-Myc真核表达载体，将上述质粒分别转染293细胞进行瞬时表达，Western印迹检测Plk1蛋白的表达；将上述质粒分别与c-Abl表达质粒共转293细胞，检测带有不同标签的Flag-Plk1或Myc-Plk1对细胞中c-Abl激酶表达的影响。结果：构建了Flag-Plk1和Myc-Plk1真核表达质粒，其在293细胞中均可表达，蛋白的相对分子质量为68×103；与其共转的c-Abl激酶表达水平显著下调。结论：在293细胞中表达了Flag-Plk1和Myc-Plk1蛋白，且初步发现Plk1可以抑制c-Abl的表达。