In order to clarify the genetic diversity and population genetic structure of Botrytis cinerea on solanaceous vegetables in Qinghai Province, 213 strains of Botrytis cinerea isolated from solanaceous vegetables were analyzed by ISSR molecular marker technology. A total of 243 polymorphic loci was amplified by 12 primers. Cluster analysis based on ISSR data showed that 213 strains could be divided into 9 groups at the level of 0.65 similarity coefficient. The Nei's gene diversity index (H) was 0.437 0, the percentage of polymorphic loci (P) was 100%, the effective number of alleles (Ne) was 1.807 3, the Shannon's information index (I) was 0.625 6, and the observed number of alleles was 2.000 0. The genetic differentiation coefficient (Gst) was 0.203 6, and the gene flow (Nm) was 1.956 4. The genetic variation of B. cinerea mainly occurred within the population (85%). Based on the similarity coefficient of 0.89, the five geographical populations could be divided into two main groups and three subgroups. The PCoA analysis reveals that although there is some degree of overlap among the populations, they generally exhibit a geographically clustery tendency. The genetic variation among the three host populations is only 2%. Compared with the geographical origin population, the population genetic distance is close and the genetic similarity is high. In the PCoA analysis, most of the individual strains formed a close cross, and a small number of strains were discrete.