Plant production in a plant factory is an innovative and smart idea to grow food anytime, anywhere, regardless of the outer environment. However, potato pre-basic seed tuber (PBST) production in a plant factory is a comparatively new initiative. Therefore, the aim of this study was to optimize the artificial LED light spectrum to produce PBST in a plant factory. Two potato varieties such as Golden king (V48) and Chungang (V41) were grown in soil substrate under different combination of artificial LED light combinations (such as red+blue+far-red, red+blue+white, blue+far-red, blue+white, red+far-red, and red+white) maintaining photosynthetic photon flux density (PPFD) of 100 mol m−2s−1, temperature 23/15 °C (day/night), and relative humidity 70%. The study revealed that, overall, potato plant growth (viz.; plant height, node number, leaf number, leaf length and width, fresh and dry weight) was enhanced by the red+far red light for both potato varieties. The total seed tuber number per plant was higher in red+blue+white light for V48, and red+far-red for V41. The fresh tuber weight was the highest in the red+blue+far-red light for V48 and red+blue+white for V41. The highest accumulated photosynthetic pigment (total Chlorophyll, Chlorophyll a, b and Carotenoid) was observed in red+blue+white light for both varieties. The total carbohydrate content and total sucrose content were higher in red+blue+far red and red +far red light treatment for V48 and V41, respectively. Finally, considering all factors, it is concluded that the red+blue+white light combination is deemed to be appropriate for the potato PBST production in plant factory conditions.