As application scopes of photon transfer curve and conversion gain of a CMOS (Complementary Metal-oxide-Semiconductor)image sensor are limited after irradiated by EMVA 1288 standard testing, an improved testing method of photon transfer curve and conversion gain of CMOS image sensor is presented. By adjusting test conditions, the method limits the dark current and the non-uniform noise of dark current from the CMOS image sensor after irradiation to solve the correct device parameters. By which the device performance changes caused by irradiation are intuitively obtained. An experimental test is performed with the proposed method, and the results show that the switching gain caused by irradiation is reduced by 7.82% . On the basis of the results, the degradation mechanism of photon transfer curve and conversion gain of the CMOS caused by irradiation is analyzed. The results point out that conversion gain degradation comes from the increses of dark current and the non-uniform noise of dark current caused by the proton radiation ionization effect and displacement effect. The paper provides a theoretical basis for mastering the spatial radiation effect of CMOS image sensors.