Abstract Background Oryza glumaepatula represents an important resource of genetic diversity that can be used to improve rice production. However, hybrid sterility severely restricts gene flow between Oryza species, and hinders the utilization of distant heterosis in hybrid rice breeding. Results In order to fully exploit the beneficial genes of O. glumaepatula and facilitate the conservation of these gene resources, a set of chromosome single-segment substitution lines (SSSLs) was developed using an indica variety HJX74 as the recurrent parent and an accession of O. glumaepatula as the donor parent. During the process of SSSLs development, S23, a locus conferring hybrid male sterility between O. sativa and O. glumaepatula, was identified and fine mapped to 11.54 kb and 7.08 kb genomic region in O. sativa and O. glumaepatula, respectively, encoding three and two candidate ORFs, respectively. qRT-PCR and sequence analysis excluded one common ORF as the candidate gene. In addition, hybrid male sterility caused by S23 was environment-sensitive, and could be observed only in natural short-day (NSD). Conclusion Identification and candidate genes analysis of S23 in this study provides a valuable example to study the crosstalk between interspecific F1 hybrid male sterility and environment-conditioned male sterility in rice, facilitates reserving and utilizing favorable genes or alleles of wild Oryza species, and allows for a more efficient exploitation of distant heterosis in hybrid rice breeding.
