Construction and identification of plasmid vector of CDK7 / 军事医学

ABSTRACT

Objective To construct the eukaryotic expression vector of cyclin-dependent kinase ( CDK) 7 labeled with Myc tag, obtain the expressed product , and identify its interaction with FLAG-P53 at the protein level .Methods Human CDK7 coding gene region amplified from the mammary cDNA library by PCR was inserted into the pXJ -40 vector.The recombinant plasmid Myc-CDK7 transfected into human 293T cell lines was investigated and examined by SDS-PAGE and Western blotting.In addition,assay was applied to determine the interaction between Myc-CDK7 and FLAG-p53.Results The coding region of CDK7 was successfully amplified by PCR and cloned into pXJ-40 vector, which was identified by double enzyme digestion and gene sequencing .Myc-CDK7 was successfully expressed in human 293T cell lines according to SDS-PAGE and Western blotting assay indicated that Myc-CDK7 could interact with P53 protein, which verified its known function .Conclusion The eukaryotic expression vector Myc-CDK7 is successfully obtained , which will contribute to further research on CDK 7-mediated cell cycle regulation .