Abstract: The L₂₅(5⁶) orthogonal diagram was applied to study the effects of different concentrations of Taq polymerase, Mg²⁺, dNTPs, primer and DNA on litchi (Litchi chinensis Sonn.) RAPD-PCR results and to get a suitable PCR system. The results showed that the orthogonal design could be used to optimize the RAPD-PCR system. A suitable RAPD reaction system for litchi was developed, i.e.1×Taq polymerase reaction buffer, 2.0 mmol/L Mg²⁺, 2.0 U Taq DNA polymerase, 0.15 mmol/L dNTPs, 0.6 μmol / L primer and 25 ng genomic DNA templates with total 25 μL reaction solution.