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Buthidae family is a scorpion family distributed throughout the world, especially in North Africa. The individual and geographical variability of this family members is associated with evolutional genetic development reflected in their venoms even in the same species. Intragenic differences can be a result of genetic and environmental factors. DNA barcoding is a system that, based on the conserved DNA sequences, helps in differentiating the different species and same species in different habitat. This study aimed on using the mitochondrial cytochrome c oxidase 1 (COX1 or CO1 or COI) to obtain molecular identification information about the taxonomic status of a set of scorpion species (Androctonus crassicauda, Androctonus bicolor, Androctonus amoreuxi, Leiurus quinquestriatus, and Buthacus arenicola) collected from Egypt. COI gene is slowly evolving comparing to other protein-coding mitochondrial genes. Therefore, it is widely used for estimating molecular phylogenies. Also, COI gene represents one of the largest sequence data sets generated from any group for phylogenetic study and fulfills the phylogenetic accuracy putative. The sequencing data of the cytochrome oxidase 1 gene were applied to provide information for better understanding of the intraspecific variation, evolution, and genetic distance between these species. COI data from the Egyptian A. australi, and those of L. quinquestriatus from Sudan, previously reported in Egypt, had been included for genetic comparison. The results revealed that the high genetic diversity was found among A. amoreuxi and A. australis. There is no detectable genetic variation between the Egyptian samples of L. quinquestriatus and those isolated from Sudan. This study helped us to understand the current evolution of these six scorpions on the genetic level and gave us better understanding of differences of scorpion envenomation (SE) of the same species in different localities. [ABSTRACT FROM AUTHOR]