Cactaceae species are an important component of the Brazilian Chaco landscape. Sixteen species are reported to this region, including 13 genera representing three Cactaceae subfamilies. All these species are native and have been locally threatened by the advance of the deforestation, which can negatively impact their genetic diversity. In order to test genetic markers that can potentially be used to screen the population diversity of these species, we checked the cross-amplification performance of 27 nuclear and 23 plastid microsatellite loci in all 16 cacti species from Brazilian Chaco. We tested the cross-amplification of the 50 microsatellite (SSR) loci in one specimen of each cacti species and considered it successful when at least one band of the expected size was generated. Thirteen species (81%) had at least 18 nuclear microsatellite loci amplified, while seven species (43%) had at least 11 chloroplast microsatellite loci amplified. We also reviewed current knowledge of SSR studies with Cactaceae in 50 studies available in the Web of Science®, and found that only five cacti species that occur in the Brazilian Chaco have representatives of the same genus with described SSR loci. The high cross-amplification rates indicated that these microsatellites markers can be helpful for future population genetic studies with cacti species from the Brazilian Chaco. Since their diversity levels and gene flow patterns are still poorly known, analyses with universal and transferable markers provide important tools to guide conservation efforts on this highly neglected region.