[Objectives]  To investigate whether the Bemisia tabaci (Q-biotype) UDP-glycosyltransferase (UGT) gene is involved in resistance to thiamethoxam. [Methods]  PCR primers were designed according to the genome database and the full-length sequence of the UGT354A1 gene was cloned. Expression levels of the UGT354A1 gene in different developmental stages and body parts, and in a thiamethoxam-resistant (THQR) and susceptible strain (THQS) were detected by q-PCR. The function of UGT354A1 gene in the resistance of B. tabaci to thiamethoxam was verified by RNAi. [Results]  Sequence analysis shows that the UGT354A1 gene has a conserved UGTs domain that is typical of insect UGTs. Compared with the susceptible strain, the UGT354A1 gene was overexpressed (2.60-fold) in the thiamethoxam resistant strain. In addition, thiamethoxam could significantly induce the expression of the UGT354A1 gene. The experiments of the expression levels of the UGT354A1 gene in different developmental stages and body parts indicate that it is most highly expressed in nymphs and adults, and in the head and thorax. RNAi silencing of UGT354A1 gene expression significantly reduced the resistance of B. tabaci to thiamethoxam. [Conclusion]  The UGT354A1 gene is involved in the formation of thiamethoxam resistance in B. tabaci. The results improve the understanding of the role of UGTs in the evolution of insecticide resistance, and provide a theoretical basis for the comprehensive management of B. tabaci.