Nickel loaded porous carbon nanofibers (NiCNF) was deposited on the surface of glassy carbon electrode by potentiostatic method for sensitive determination of guanine (G) and adenine (A) at oxidation potentials 0.787 V and 1.091V, respectively. The synthesized NiCNF were characterized by scanning electron microscopy, X-ray diffraction, Raman and electrochemical impedance spectroscopy. Simultaneous determination of G and A was successfully conducted in 0.1 M phosphate buffer (pH 4) and the linear relationships of G and A were obtained in the range of 0.05-2 mu M. The proposed electrode performed a good reproducibility, selectivity and sensitivity to both DNA purine bases and it showed satisfactory results in the determination of real fish sperm DNA sample. (C) 2015 Published by Elsevier Ltd.