Application of DNA testing has been developed into a requisite and regularcomponent of recent forensic casework, based on highly sensitive and stable PCR techniques for the analysis of different types of biological specimens. In the current study, development, optimization and validation of a multiplex system was accomplished comprising on 10 autosomal miniSTRs, SE33, Penta E, Penta D and four Y chromosomal STRs (DYS385a/b, DYS438 and DYS392) along with gender detection marker amelogenin into a single PCR reaction simultaneously.