This study evaluated the effects of various supplemental lighting conditions on the growth and rhizome production of Wasabia japonica, a high-value crop. Its cultivation poses challenges due to specific environmental requirements, including cool temperatures and high humidity. By tailoring light spectra, we aimed to optimize photosynthetic efficiency and biomass accumulation. Seedlings (cv. ST1) were grown in a controlled plant factory (18 ± 2 °C, 90 ± 5% RH) using a Yamazaki tri-leaf nutrient solution (EC 1.5–2.0 dS/m, pH 5.8–6.8). Lighting was designed to convert blue (450 nm) to red (630–680 nm) wavelengths at 70–40% ratios, maintaining a photosynthetic photon flux density (PPFD) of 50 ± 5 μmol·m⁻²·s⁻¹ during a 12 h photoperiod. In the greenhouse, supplemental lighting (40% blue-to-red conversion) was applied for 4 h daily to complement natural light. After 140 days, the optimized 40% blue-to-red conversion lighting significantly improved plant length (34.4 cm), leaf length (15.3 cm), and photosynthetic rates (2.21 μmol·m⁻²·s⁻¹). When tested in the greenhouse, it increased rhizome fresh weight to 75.6 g compared to 30.0 g under natural light. These results indicate that supplemental lighting with specific spectral ratios and controlled intensities can enhance photosynthesis and rhizome production, providing a sustainable approach to wasabi cultivation.