Construction of eukaryotic plasmid of human Plk2 gene and the expression and localization of fusion protein

Objective:To construct the expression plasmid of human Polo-like kinase 2(hPlk2) gene and identify the expression and localization of the fusion protein.Methods:Total mRNA was extracted from Hela cells,and cDNA was synthesized by reverse transcription.The hPlk2 coding sequence was amplified by polymerase chain reaction(PCR) and subcloned into pEGFP-C1 vector.After the target region was identified by restriction enzyme digestion and sequencing,the plasmid was transfected into COS-7 cells.The expression of the recombinant plasmid in COS-7 cells was detected by Western blot assay.The localization of pEGFP-hPlk2 in COS-7 cells was observed with laser scanning confocal microscopy.Results:hPlk2 was constructed into the expressing vector pEGFP-C1 successfully.The length of the fragment identified by restriction enzyme digestion was 2058bp.The expression of pEGFP-hPlk2 fusion protein with a molecular weight of 105kDa was detected by Western blot.The pEGFP-hPlk2 fusion protein was mostly localized in the cytoplasm and perinucleus of COS-7 cells.Conclusion:The recombinant plasmid of hPlk2 gene was successfully cloned into eukaryotic expressing vector,and the pEGFP-hPlk2 fusion protein was mostly localized in the cytoplasm and perinucleus of COS-7 cells.