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In this paper, a novel method to directly determine the surface roughness scattering-limited mobilities (μsr) of electrons and holes in Si MOSFETs from the experimental data of MOS interface roughness is proposed and compared with the experimental μsr of Si MOSFETs with and without biaxial tensile strain. This method includes the direct evaluation of the scattering potential from the power spectra of Si/SiO2 interface roughness data, which are taken through high-resolution advanced transmission electron microscopy measurements, without assuming any autocorrelation function form of the interface roughness, like a Gaussian or exponential function. It is found, for the first time, that, by employing the present method, experimental electron and hole μsr (both unstrained and strained Si) could be presented by a same model. As a result, the difference in strain dependence between electron and hole μsr, which has experimentally been observed, is systematically explained by the change of power spectra of the interface roughness due to strain.