For the establishment of a simple,sensitive and rapid method for the diagnosis,prevention and control of the Koi herpes virus disease,based on the KHV DNA polymerase gene conserved sequence (Sph),a set of single cross amplification primers were designed.The constructed Sph gene plasmid DNA was used as the standard template in the single cross priming amplification (SCPA) and conditions optimization.The detection results showed that the cross circulation amplification could be realized at 63 ℃ within 60 min,and the amplified products showed DNA ladder analyzed by agarose gel electrophoresis.KHV-SCPA could specifically detect KHV with a sensitivity that is about 1 000 times higher than that of the traditional PCR method.In addition,combined with the nucleic acid test strip detection technology,the visual detection of KHV amplified products within 3~5 min was realized.The final concentrations of each component in the optimal reaction condition are as follows:1×ThermoPol^® Reaction Buffer,cross primer 2a1s 1.0 μmol/L,displacement primers 2a,3a 0.5 μmol/L,respectively,peeling primers 4s,5a 0.6 μmol/L,Mg²⁺ 8.0 mmol/L,dNTPs 1.2 mmol/L,Betaine 0.7 mol/L,Bst DNA polymerase 8U,DNA template 1 μL,respectively.The nuclease-free water is added to a total volume of 25 μL.The optimal amplification temperature is 63 ℃,the optimal reaction time is 60 min.KHV-SCPA nucleic acid test strip visual detection method does not require expensive equipment and skilled technicians,which could be used in the on-site diagnosis for effective prevention and control of Koi herpesvirus infection in common carps.