Deep-seeding tolerant seeds can emerge from deep soil where the moisture is suitable for seed germination. Breeding deep-seeding tolerant cultivars is becoming increasingly important in arid and semi-arid regions. To dissect the quantitative trait loci (QTL) controlling deep-seeding tolerance traits, we selected a tolerant maize inbred line 3681-4 and crossed it with the elite inbred line-X178 to generate an F(2) population and the derivative F(2:3) families. A molecular linkage map composed of 179 molecular markers was constructed, and 25 QTL were detected including 10 QTL for sowing at 10 cm depth and 15 QTL for sowing at 20 cm depth. The QTL analysis results confirmed that deep-seeding tolerance was mainly caused by mesocotyl elongation and also revealed considerable overlap among QTL for different traits. To confirm a major QTL on chromosome 10 for mesocotyl length measured at 20 cm depth, we selected and self-pollinated a BC(3)F(2) plant that was heterozygous at the markers around the target QTL and homozygous at other QTL to generate a BC(3)F(3) population. We found that this QTL explained more phenotypic variance in the BC(3)F(3) population than that in the F(2) population, which laid the foundation for fine mapping and NIL (near-isogenic line) construction.