This work details the preparation of gold nanoparticles/single-walled carbon nanotubes/polyaniline composite-modified disposable pencil graphite electrode (AuNPs/SWCNTs/PANI/PGEs), and its application for electrochemical DNA sensing. Disposable pencil graphite electrode was modified with single-walled carbon nanotubes/polyaniline composite via one-step electropolymerization of aniline solution containing well-dispersed single-walled carbon nanotubes. AuNPs were then immobilized onto the SWCNTs/PANI/PGE. Electrochemical results exhibited improved sensitivity for DNA detection with AuNPs/SWCNTs/PANI/PGE compared to the SWCNTs/PANI/PGE, PANI/PGE or unmodified PGE due to the strong interaction between AuNPs, SWCNTs, and PANI based on the effective degree of electron delocalization. Cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS) were used to characterize the prepared electrode. Thiol-linked ssDNA was immobilized onto the AuNPs/SWCNTs/PANI/PGE for DNA detection. Experimental parameters such as the concentration of AuNPs and ssDNA were investigated to optimize the experimental conditions.