Coix lacryma-jobi L., an annual or perennial plant belonging to the Poaceae family, has long been cultivated as a food and medicine plant in China. In recent years, coix cultivation for high yields and good quality has become a research hotspot in Southwest China. Soil optimization is essential for improving crop growth. To ensure the robust establishment of coix plants, eight soil substrates, prepared from three typical soils, i.e., red clay soil, peat soil, and sandy soil, were used to cultivate two local coix varieties (Pu coix from Fujian Province, China; Qi coix from Hebei Province, China), and the plant growth and root coixol content of the two coix varieties were investigated. It was found that coix plants could maintain growth when cultivated with peat soil or sandy soil, but red clay soil was unfavorable for coix growth. The mixtures of sandy soils and peat soils resulted in synergistic benefits for coix growth and root coixol levels over the effects of sandy soil or peat soil alone. In conclusion, the mixtures of sandy soils and peat soils with appropriate proportions (sandy soils/peat soils = 2:1) were suggested as an ideal soil substrate for coix cultivation. The results provide valuable guidance for the establishment of coix plants, which could contribute to high yields and good quality in coix cultivation.