Objective:To establish high throughput mitochondrial DNA(mtDNA)SNPs detection system and study the genetic polymorphism of the Guangdong Han population by SNaPshot technology. Methods:In the present study,we selected 21 SNPs from mitochondrial genome,designed primers,performed PCR,co-amplified single base extension(SBE) and also combined with capillary electrophoresis. Results:Multiple detection system were successfully established. The genetic polymorphism of Guangdong Han population was investigated,and 90 haplotypes were detected in the studied population,and the haplotype diversity(HD) was 97.1%. Conclusion:The results show that the 21-plex mtDNA SNPs multiplex detection system has a pretty good application prospect in forensic medicine.