His-PCNA融合蛋白的表达与纯化及其与MYCBP2的相互作用

Objective: To construct and express pET-20b-PCNA recombinant plasmid and to verify the interactin between His-PCNA fusion protein and MYCBP2 protein. Methods: Using PCR and recombinant DNA technology, PCNA full gene sequence was amplified from template pcDNA-FLAG-PCNA and inserted into the prokaryotic expression vector pET-20b(+) with His-tag. The recombinant product was then transformed into the E. coli Rosseta strain to produce the His-PCNA recombinant plasmid, which was purified through a His . Mag column, and verified by SDS-PAGE and western blotting using an anti-His polyclonal antibody. Using His-tag pull-down assay, we confirmed the interaction between His-PCNA and MYCBP2 protein. Results and conclusion: We successfully constructed the pET-20b-PCNA plasmid, produced and purified the His-PCNA fusion protein. Furthermore, we for the first time verified the interaction between His-PCNA and MYCBP2 protein.