Abstract: The sesame (Sesamum indicum L.) DNA was extracted from young leaf and taken as tested materials. In order to establish the optimum SRAP reaction system which could amplify high levels of polymorphism, good repeatability and clear band pattern, the SRAP amplification system on sesame was optimized from many factors, such as DNA template, Mg2+, dNTPs, Taq DNA polymerase and primers. The optimum SRAP system (total volume of 15μl) was as follows: dNTPs (10mmol／L) 0.30μl, Mg2 + (25mmol／L) 1.20μl, Taq polymerase 1.00U, forward primer 50ng, reverse primer 50ng, DNA template 80 ng, 10×Buffer 1.5μl. This work is a foundation for applying SRAP marker to sesame molecular biology studies.