The direct bonding process of a diamond-on-insulator (DOI) substrate enables monolithic integration of diamond photonic structures for quantum computing by improving photon collection efficiency and entanglement generation rate between emitters. It also addresses key fabrication challenges, such as robustness, bonding strength, and scalability. This study investigates strain effects in DOI substrates following direct bonding. Strain generation is expected near the diamond–SiO₂/Si interface due to the thermal expansion coefficient mismatch between the bonded materials. Strain-induced lattice distortions are characterized using nitrogen-vacancy (NV) centers in diamond via optically detected magnetic resonance (ODMR) and photoluminescence (PL) mapping. PL mapping reveals interference fringes in unbonded regions, indicating bonding irregularities. Depth-resolved ODMR measurements show a volumetric strain component increase of ≈0.45 MHz and a shear component increase of ≈0.71 MHz between the top surface and the DOI interface. However, ODMR signal contrast and peak linewidth remain largely unaffected, suggesting no visible deterioration in the optical properties of the emitters. By combining ODMR and PL mapping, this work establishes a robust methodology for assessing bonding quality and strain impact on NV centers, an essential step toward advancing scalable quantum technologies and integrated photonic circuits.