Self-assembled monolayers (SAMs) are ultra-thin organic films capable of precisely tailoring the physical and chemical properties of substrate surfaces. In this study, we demonstrate that introducing anisotropic morphology into the substrate effectively guides the lateral diffusion of SAM molecules along predefined directions during the self-assembly process, thereby reducing random molecular orientation at the macroscale. Notably, amplified optical signals obtained through liquid crystal (LC) texture analysis were used to visualize defects arising from molecular alignment within the SAM, while a multi-level analysis of surface energy, molecular packing density, and alignment quality further enhanced the precision of LC-based texture interpretation. These findings demonstrate that the proposed bottom-up strategy is broadly applicable across a wide range of alkyl chain lengths and may serve as a versatile platform for advanced electronic and optoelectronic substrate applications. ⓒ 2025 Elsevier B.V.