Heterosis plays an important role in enhancing crop yield and quality. Dissecting the genetic basis of heterosis can promote hybrid maize selection, however it is unclear up to now. In this study, a set of chromosome segment substitution lines (CSSLs) population, which was constructed using the inbred line lx9801 as the receptor parent and the inbred line Chang 7-2 as the donor parent, was crossed with the inbred line T7296 to construct the corresponding test population. The test population was used to identify the heterotic loci (HL) for grain yield and ear traits in maize, which showed significant difference in heterosis between the corresponding chromosomal region of the inbred line Chang 7-2 and lx9801 as well as the test inbred line T7296. A total of 64 HL were identified for gain yield and ear traits, and among them 23 HL were identified at the two environments simul- taneously, including 4 HL for ear length, 4 HL for ear width, 4 HL for row number, 7 HL for kernels per row, and 4 HL for grain yield. Additionally, the HL for both grain yield and its components simultaneously were found on many chromosomal regions. This study could offer a basic material for thoroughly dissecting the genetic basis of heterosis for grain yield and its components in maize.