Prokaryotic expression and purification of fusion protein PTD-Cyclin D1

AIM: To construct the recombinant expression vector and to obtain large amount of mouse Cyclin D1 protein fused to PTD. METHODS: A 888 bp of mouse Cyclin D1 gene fragment was amplified by PCR method and cloned into pET16b vector immediately downstream of the PTD fragment, an E.coli expression vector, to construct a recombinant plasmid pET16b-PTD-CCND1. The plasmid was transformed into E.coli BL-21 (DE3) and induced to express fusion protein PTD-Cyclin D1 with IPTG. The expression of PTD-Cyclin D1 was detected by SDS-PAGE electrophoresis and Western blot. RESULTS: A novel protein with expected molecular mass was expressed upon induction with IPTG. The expressed product showed good reactivity to anti-His tag antibody, and was mostly in the form of inclusion bodies. CONCLUSION: Our successful cloning and expression of Cyclin D1 gene and purification of Cyclin D1 protein lay a basis for further study on the application of this protein to accelerate cell proliferation in vitro and in vivo.