Marker-assisted selection (MAS) can accelerate the process of plant breeding, and sequence-tagged site (STS) markers are highly specific for regions of DNA being used for MAS. The objective of this research was to develop STS markers tightly linked with Rf1, the fertility restoring gene for cytoplasmic male sterility (CMS) in cotton (Gossypium hirsutum L.). Bulked segregant analysis was employed to screen for Rf1-linked RAPD markers in a backcross population. Four RAPD markers were identified, three of which co-segregated with Rf1 (UBC147(1400), UBC607(500), and UBC679(700)). Another fragment, UBC169(800), co-segregated with the previously reported UBC169(700) in repulsion phase at a distance of 4.5 cM from Rf1. A marker published by others (UBC659(1500)) mapped to 2.7 cM from Rf1 and 1.8 cM from UBC169(800). Four sets of STS primer pairs were designed based on the RAPD fragment sequences. The primer pairs from the UBC147(1400) and UBC607(500) fragments both amplified a single fragment specific to fertile plants. The UBC679(700) and UBC659(1500) STS primer pairs each amplified one fragment specific to fertile plants and a monomorphic fragment. These four Rf1-linked STS markers were also present in the Rf1 donor species G. harknessii (D2-2). The three primer pairs that produced co-segregating STS markers also amplified fragments from G. trilobum (D8). However, the D8 fragment amplified by the UBC147(1400) STS primers was larger than that from D2-2, and G. trilobum does not restore fertility to CMS-D2-2 lines. These STS markers will be useful in the development of restorer parental lines in cotton CMS breeding efforts.