The mirror carp breeding of generations targeted at Koi herpes virus (CyHV-3) has been carried out to F₃ generation. F₂ and F₃ survival rates are similar and significantly higher than F₁. In this study, theCyHV-3 viral loads and the gene expression of IL-1β, TRAF6, MyD88a, I-IFN and TLR7a are compared by real-time fluorescence quantitative PCR technique. Immune gene expression levels and viral loads in combination with disease resistance survival rates are used to evaluate the disease resistance of different generations. The results showed that: (1) CyHV-3 virus load decreased sequentially in F₁, F₂ and F₃, and F₃ was significantly lower than F₁ and F₂ (P<0.05); (2) Summerlings group wasn't infected CyHV-3, only the expression of TRAF6 gene in F₃ was significantly higher than that in F₁ and F₂ (P<0.05). (3) In the infected group, the levels of IL-1β and MyD88a in F₃ spleen were significantly higher than those in F₁ and F₂ (P<0.05), and TRAF6 was significantly higher than that of F₁ (P<0.05). The expression of IL-1β and TRAF6 in F₃ kidney was significantly higher than that in F₁ and F₂ (P<0.05). There was no significant difference between F₁ and F₂ (P>0.05). The above results show that F₃ disease resistance is significantly higher than F₁, F₂, only the TRAF6 gene was breeding. Based on the fact that F₂, F₃ survival rate was similar and significantly higher than F₁, it can be concluded that F₂ disease resistance is not stable and F₃ resistance has stabilized, and that using the immune gene expression or viral load method to evaluate the disease resistance is feasible.