Abstract: The major pathogen of corn sheaths rot is Fusarium proliferatum.In order to clarify the optional response system of ISSR-PCR of F.proliferatum.DNA was used as a template to screen the concentration range of each factor,and orthogonal test was used to optimize 5 factors to determine the best response system.The primers with clear amplified bands and good polymorphism were selected from 13 primers and selected optional annealing temperature.The results showed that ISSR-PCR optional reaction system:template 40 ng·μg-1,magnesiumion concentration 2.5 mmol·L^-1,primer concentration 0.4μmol·L^-1,dNTPs concentration0.2 mmol·L^-1,Taqenzyme concentration 2.5 U·μL^-1,the optional primer was 808 and optional annealing temperature was 58℃,a total of 14 bands were amplified with the screening out of primers 808.And polymorphic bands were 13 among them.The percentage of polymorphic bands was 92.9%.