CAJ | 학술논문

Objective: To clone and construct eukaryotic expressing vectors of sense and antisense human Pin1 (hPin1)genes.Methods: Total RNA was extracted from MG-63 cells,then the hPin1 cDNA was amplified by RT-PCR.The same time the sense and antisense hPin1 genes were formed by binding BamH Ⅰ and Hind Ⅲ in cis and trans-directions.At the end they were cloned into the eukaryotic expressing vector pIRES2-EGFP in cis and trans directions using DNA recombinant technology.The recombinant vectors were further identified by digestion of BamH Ⅰ and Hind Ⅲ.Results: The results of sequencing showed that the orientation of the ligations and the reading frame were correct.After digested by BamH Ⅰ and Hind Ⅲ,two fragments exhibiting 5.3 kb and 0.99 kb were formed in sense and antisense eukaryotic expressing vectors.Electrophoretic results were completely coincident with theoretical calculation.Conclusion: Human Pin1 sense and antisense genes were successfully cloned and eukaryotic expressing vectors were successfully constructed.