Flesh flies comprise a large fly family distributed worldwide that has great importance for forensic entomology. A robust and updated checklist of Sarcophaginae flies recorded in Southern Brazil is presented, based on material collected in the Rio Grande do Sul state and on a literature review. The forensic importance (high, moderate, or none) of the recorded flesh flies for estimating the postmortem interval (PMI) or inferring cases of neglect was determined based on their ecological habits. A total of 2,831 specimens representing 38 taxa were collected from three studies. Oxysarcodexia was the most abundant and species-rich genus in all three studies. Dexosarcophaga carvalhoi (Lopes) was registered for the first time in Southern Brazil. The checklist for Southern Brazil comprises 91 species distributed in 19 genera, with Oxysarcodexia, Peckia, and Lepidodexia representing the richest genera. Considering the importance of species for PMI estimation, 10, 42, and 39 species were classified as having high, moderate, or no forensic relevance, respectively. Moreover, five myiasis-causing sarcophagine flies recorded in Southern Brazil were considered to have the potential to reveal cases of neglect or mistreatment. This study revealed that Microcerella halli (Engel) and Peckia (Euboettcheria) australis (Townsend) are the most important species for PMI estimation in Southern Brazil, as they are often found breeding in corpses in this region.