Glucosyltransferase plays an important role in plant stress resistance and development regulation. In order to explore the members of the glucosyltransferase BnIRX14 gene family and analyze its biological function in Brassica napus, the study based on BnIRX14, which was cloned from B. napus and adopted the methods of sequence comparison and genetic transformation, we identified BnIRX14 gene family members and conducted functional verification to explore the regulation mechanism of BnIRX14 gene family in B. napus development, and to provide theoretical basis for cross breeding and resistance breeding in B. napus. The results showed that: (1) 11 BnIRX14 family members belonging to 3 different subfamilies of glucosyltransferase were successfully identified in B. napus by genomic database comparison analysis. All of them had the domain characteristics of glucosyltransferase GT43 family members, among which 8 genes were located on 6 different chromosomes. 3 subfamilies showed great specificity in gene structure and conserved elements. (2) Twenty transgenic B. napus lines with BnIRX14 RNA interference were obtained by Agrobacterium-mediated transformation, and 5 positive transformation lines were identified by PCR. (3) Phenotypic identification showed that there was a hole-like cavity from the top to the center of the stigma of the two positive transformation lines, the ovary was significantly enlarged than that of the wild type, and the stigma was not fertile after pollination, indicating female sterility. The other three positive plants had normal flower structure, but the stem and branch of the plants had liquid exudation, which adhered to the surface of the stem and branch in the shape of dew. (4) Real-time quantitative PCR(RT-qPCR) analysis showed that the BnIRX14 gene expression in the flowers, kernels and leaves of B. napus transgenic positive plants was significantly lower than that of the wild type, and the BnIRX14 gene expression in the kernels of the 9# positive sterile plant was lower than that of other transgenic interference lines. The results showed that BnIRX14 gene expression was significantly inhibited in the interference-transformed positive plants. It is speculated that BnIRX14 gene may be involved in pistil development and metabolism of secondary substances in rape.