Efficient local gene transfection on a tissue scaffold is of crucial importance in facilitating tissue repair and regeneration. In this work, the gelatin-functionalized polycaprolactone (PCL) film surfaces are prepared via surface-initiated atom transfer radical polymerization of glycidyl methacrylate. The resultant covalent attachment of gelatin could enhance the cell-adhesion and local gene transfection properties. The gelatin-functionalized PCL film surfaces exhibit excellent cell-adhesion ability to both adherent and suspension cells. The attached adherent cells demonstrate the characteristic elongated morphologies with good spreading capability, while the attached suspension cells can maintain the original status of the round morphologies without spreading. More importantly, the gelatin coupled on the PCL surface could be used to absorb the cationic vector/plasmid deoxyribonucleic acid (pDNA) complexes via electrostatic interaction. The local gene transfection property on the immobilized cells is dependent on both the density of the immobilized cells and the loading types of pDNA complexes. The transfection efficiency of different assemble methods of pDNA complex was compared. With the pre- and post-loading sandwich-like gene transfection, the gelatin-functionalized PCL film surface can substantially enhance the transfection properties to different cell lines. The present study is very useful to spatially control local gene delivery within PCL-based tissue scaffolds.