Indolicidin (IL) is a cationic antimicrobial peptide and our previous study has demonstrated its potential as a cell penetrating peptide (CPP) to promote gene delivery. However, the cytotoxicity of IL arisen from its membrane perturbation capacity may restrict its clinical application. To promote gene delivery safety and efficiency, an almost mirror-symmetric IL derivative, SAP10 (RRWKFFPWRR-CONH2), was designed in this study. All-atom molecular dynamics (MD) simulations were performed to understand the association between SAP10 and model lipid bilayers. By comparison with IL, SAP10 with high positively charged density resisted its deep insertion into lipid bilayers, which thus reduced its perturbation to lipid bilayers and improved biocompatibility. Consequently, we further mixed SAP10, polyethylenimine (PEI) and DNA to form the ternary nanocomplexes for gene delivery investigation. Both IL and SAP10 weakened the interaction between to DNA and PEI, which may be beneficial to promote the dissociation of internalized DNA from the carrier molecules. In vitro experiments demonstrated that the SAP10-associated ternary nanocomplexes highly promoted the transfection efficiency to various cells with low cytotoxicity. The effect of the SAP10 on promoting gene delivery was mainly contributed by the adsorbed peptides on the nanoparticles rather than the free ones. In particular, the dose of SAP10 could be increased to broaden the administration window, which ensured its safety on transfection. Therefore, our results suggested the argument that the designed SAP10 is a safe and an efficient peptide to promote PEI-mediated gene delivery.