The microwave plasma chemical vapor deposition technology was employed to grow the single crystal diamond substrates with different crystalline qualities by changing the nitrogen content in the gas source. The sample size was controlled at 5 mm×5 mm×0.5 mm by laser cutting and polishing. The surfaces of diamond substrates were hydrogenated and the diamond RF transistors were fabricated on them. The influence of the nitrogen content on the crystalline quality of the diamond and the performance of the device has been systematically studied. With the increase of the nitrogen content, although the growth rate of the single crystal diamond increases, the FWHM of Raman and XRD rocking curves, as well as the corresponding NV defects in the photoluminescence spectra also gradually increases, indicating that the crystalline quality became worse, which not only degrade the carrier mobility of the channels, but also make the diamond RF transistor suffer from the problems of current collapse and performance degradations. By reducing the nitrogen content and improving the crystalline quality of the diamond, the carrier mobility significantly increases, the current gain cutoff frequency fT and power gain cutoff frequency fmax significantly increases from 17 GHz and 22 GHz to 32 GHz and 53 GHz, respectively.