Abstract: Gufeng B is a broad-spectrum, high resistance maintainer line to rice blast. Its blast resistance genes, Pi-d2, Pi-d3 and Pigm, were identified using the molecular marker technology. To study and verify the role of these genes, their multiple sites editing vectors were constructed by the high-efficiency and designated-editing CRISPR/Cas9 system. The vectors were subsequently transformed to Gufeng B by means of the agrobacterium-mediated method. Through the mutant analysis targeting mutation types, a variety of combinations in the T₀ plants was found. The indoor inoculation of the T₁ generation of 5 homozygous mutants indicated that Pi-d2, Pi-d3 and Pigm were significantly resistant to 86/501-3, KJ201/CHE86061 and 86/CHE86061/501-3. The resistance of the knockout lines of Pi-d2+Pi-d3, Pi-d2+Pigm and Pi-d2+Pi-d3+Pigm showed a simple superposition that was not completely equivalent to a single gene. The mutant of Pi-d2+Pi-d3+Pigm had a completely different sensitivity to CHL768 from the other mutants. This study unveiled genetic information on the blast resistance of Gufeng B of value for rice breeding.