• Premise of the study: Microsatellite markers were developed using second-generation sequencing in Solanum rostratum as a tool to study the reproductive biology and genetic structure of this invasive species.• Methods and Results: Thirteen microsatellites were successfully discovered and amplified in a single multiplexed PCR. All loci showed genetic variation in S. rostratum. Cross-amplification in five closely related taxa was successful for a subset of loci.• Conclusions: The set of 13 microsatellite markers developed here provides a time-effective and cost-effective genetic tool to study the reproductive biology of S. rostratum. The demonstrated transferability of the PCR multiplex to related taxa also highlights its usefulness for evolutionary studies across Solanum sect. Androceras.