In the present work, an electrochemical nucleic acid biosensor has been designed for the purpose of detection of miR-122 in real samples, i.e cell lysates. The fabrication of the biosensor has been done first by immobilization of complemantary anti-miR-122 to the surfaces of the graphene (GRP) modified pencil graphite (PGEs) electrodes then solid phase hybridization with either synthetic miR-122 or miR-122 included in total RNA isolated from HUH-7 cell line. The characterization of GRP modification onto the PGE surface has been proved via Raman spectroscopy analysis, Electrochemical Impedance Spectroscopy (EIS) and Differential Pulse Voltammetry (DPV). The intrinsic guanine oxidation signal measured via DPV and the charge transfer resistance, R-ct values recorded via electrochemical circle fit option of EIS have been used for hybridization detection. The proposed biosensor with the limit of detection (LOD) 1 pmol is applicable for analysis of certain miRNAs as well as miR-122 from total RNAs isolated from cell lysates. (C) 2016 The Electrochemical Society. All rights reserved.