Aspergillus flavus is the second most public fungi that causing disease in human and causes diseases in several important agricultural crops. The aim of this study was to investigate the genetic variety among diverse isolates of Aspergillus flavus by the technique of polymerase chain reaction (PCR)-based random amplified polymorphic DNA (RAPD). Five isolates were obtained from Samarra hospital and outpatient clinic and were diagnosed morphologically and microscopically. Results of RAPD profile revealed a high genetic distance (0.244) between isolates number (1 and 5), while the isolates number (2 and 5) were joined with lower genetic distance (0.090). Unique and absent bands were displayed by certain isolates and can be taken as a positive marker for isolate identification. The dendrogram of banding patterns revealed a great relationship between isolates number (2 and 5) from one side and isolates number (1 and 3) from the other side, while the isolate number 4 was separated into particular subgroup which closer to the isolate number 1 and 3.