Abstract: Photo-or thermo-senstive genic male sterile gene (PGMS or TGMS) has played an important role in the utilization of heterosis in rice. The identification of PGMS or TGMS mutants and genes could help to better understand the mechanism of PGMS or TGMS in rice and enrich the genetic resources. In this study, a male sterile mutant was identified from the transgenic offspring of Minghui 86, an indica rice material. The mutant showed male sterility in the normal growth season, the anthers were smaller than those of the wild type, pale white, and no pollen was detected by microscopy. The fertility could be recovered after the low-temperature treatment at booting stage, and the mutant was a thermo-sensitive male sterile mutant, which was named mh86s. The mutant mh86s was hybridized with their wild parent Minghui 86 and the indica rice material 93-11, respectively, thus to construct the F₂ genetical population. And the genetic analysis showed that the male-sterile character was controlled by a pair of recessive genic gene. By using the map-based cloning, the gene of thermo-sensitive male sterility was located in the 49.8 kb physical interval between the molecular markers Indel7.38 and SNP3 of the chromosome 2, and the cloned thermo-sensitive male sterility gene tms5 was included in the mapping interval. The sequencing analysis revealed that there was a single-base substitution (G to T) in the second exon of tms5 gene in the mutant mh86s compared to the wild type, which led the presumed amino acid was changed from glycine to valine at the 161th position. F₁ was characterized by thermo-sensitive male sterility by the hybridization of the mutant mh86s and the thermo-sensitive male sterile line HD9802S carrying tms5 gene. Therefore, it was inferred that the gene of thermo-sensitive male sterility in the mutant mh86s was the new allele of tms5, which was named tms5-mh86. The results of this study enriched the tms5 gene resources and contributed to the analysis of tms5 gene structure and function, and the corresponding molecular markers could also be used in the marker-assisted selection of tms5-mh86.