In this study, we developed polyethyleneimine-methyl acrylate-polyethyleneimine (PMP) conjugate as a novel, non-viral gene carrier based on low molecular weight polyethyleneimine, and proved PMP derivative exhibit high gene transfection efficiency and low toxicity. We linked methyl acrylate (MA) on polyethyleneimine (PEI) and then grafted PEI onto the PEI-MA. The synthesized PMP was characterized by H-1 NMR and FTIR. When added to DNA solution, PMP condensed with DNA at weight ratios over 1:1 (polymer:DNA) to form 96-108 nm polyplexes with 37 mV of zeta potential. In cytotoxicity assay, PMP turned out less toxic than PEI 25 kDa. In vitro transfection assays showed that the gene transfection of PMP/pDNA complexes was similar or slightly higher than that of the PEI 25 kDa/pDNA complexes in 293, HeLa, and HepG2 cells. Our data suggest that PMP-based gene delivery system is economical, effective, and biocompatible, and it may be a promising gene delivery vector for gene therapies.