Abstract: Abstract: In order to establish a genomic DNA extraction method and ISSR-PCR system suitable for purple ornamental peppers, three DNA extraction methods were compared and the ISSR-PCR system was optimized by orthogonal design experiment. The results showed that, the A260/A280 values of the genomic DNA extracted from two pepper varieties by high salt / low pH method were 1.807 and 1.818 respectively, and the DNA bands were clear without tailing. The optimal ISSR-PCR conditions were: 1×PCR buffer, 1.5 mmol/L Mg2+, 0.6 μmol/L primer, 0.2 mmol/LdNTPs, 1.0 U Taq DNA polymerase，60 ng template DNA, 20 μL reaction system. The ISSR results are stable and repeatable, and two pepper varieties can be distinguished from ISSR map only using the DNA templates extracted via high salt / low pH method.