A microRNA(miR，miRNA) electrochemical biosensor was proposed in the present research.Firstly，graphene(GN) was dispersed in the chitosan(Chi) solution to promote the conductivity of chitosan film.Then，prussian blue(PB) was in situ deposited on GN to obtain the PB@GN-Chi nanocomposite.The obtained nanocomposites were modified on the surface of electrode.Compared with the PB@GN-Chi prepared by ultrasonic method，the prepared composite could effectively prevent PB leaking into detection solution and display a prominent stability when modified on the electrode.The capture probe was subsequently modified by amino group and covalently linked to the modified electrode via glutaraldehyde.After blocked by hexanethiol，the miRNA biosensor was successfully prepared.The prepared PB@GN-Chi nanocomposite was observed with transmission electron microscope，and the electrochemical characteristics of the miRNA biosensor were investigated by cyclic voltammetry and differential pulse voltammetry.Under the optimal conditions，the biosensor displayed a linear relationship between current response and the logarithm of concentration of miR-21 in the range of 2.8-2.8×104 pmol/L.The detection limit was calculated to be 0.87 pmol/L.The prepared biosensor owns good stability and repeatability，and could be employed to detect miR-21 in human serum sample.