【Objective】To accelerate the innovation process of germplasm resources of seed-used pumpkin by constructing a mutant library, which is of great significance for variety selection, improvement and broadening of the genetic base. 【Methods】 Ethyl methanesulfonate (EMS), a 1.8% mutagen, was used to treat ZHL4 seeds of seed-used pumpkin for 15h. Then, phenotypic variation was observed in single plants of the M1 and M2 generation populations, and microstructural observation was made on the mutant strain ZHL4-33 of the M2 population. 【Results】 (1) A total of 242 mutant plants with 45 phenotypic variations were screened in the M2 population, and the types of variation covered all growth periods and plant organs of the mutant plants, with a total mutation frequency of 25.17%. (2) The leaf microstructure showed that the thickness of the fenestrated tissue of the mutant was significantly higher than that of the wild type, the arrangement was compact, and the traces of the vascular layer were obvious; the stem microstructure showed that the vascular bundles of the mutant were more and densely packed, the diameter of the conduit was smaller than that of the wild type, and the pith was well developed, with a small cell gap and an increase in the number of cells. 【Conclusion】 A mutant library of 425 M2 lines has been constructed, which lays a material foundation for the study of the functional genome of seed-used pumpkin and the selection and breeding of new varieties of seed-used pumpkin.