BackgroundWheat (Triticum aestivumL.) is a major cereal crop that is grown worldwide, and it is highly dependent on sufficient N supply. The molecular mechanisms associated with nitrate uptake and assimilation are still poorly understood in wheat. In plants, NRT2 family proteins play a crucial role in NO3–acquisition and translocation under nitrate limited conditions. However, the biological functions of these genes in wheat are still unclear, especially their roles in NO3–uptake and assimilation.ResultsIn this study, a comprehensive analysis of wheatTaNRT2genes was conducted using bioinformatics and molecular biology methods, and 49TaNRT2genes were identified. A phylogenetic analysis clustered theTaNRT2genes into three clades. The genes that clustered on the same phylogenetic branch had similar gene structures and nitrate assimilation functions. The identified genes were further mapped onto the 13 wheat chromosomes, and the results showed that a large duplication event had occurred on chromosome 6. To explore theTaNRT2gene expression profiles in wheat, we performed transcriptome sequencing after low nitrate treatment for three days. Transcriptome analysis revealed the expression levels of allTaNRT2genes in shoots and roots, and based on the expression profiles, three highly expressed genes (TaNRT2-6A.2,TaNRT2-6A.6, andTaNRT2-6B.4) were selected for qPCR analysis in two different wheat cultivars (‘Mianmai367’ and ‘Nanmai660’) under nitrate-limited and normal conditions. All three genes were upregulated under nitrate-limited conditions and highly expressed in the high nitrogen use efficiency (NUE) wheat ‘Mianmai367’ under low nitrate conditions.ConclusionWe systematically identified 49NRT2genes in wheat and analysed the transcript levels of allTaNRT2sunder nitrate deficient conditions and over the whole growth period. The results suggest that these genes play important roles in nitrate absorption, distribution, and accumulation. This study provides valuable information and key candidate genes for further studies on the function ofTaNRT2s in wheat.Supplementary InformationThe online version contains supplementary material available at 10.1186/s12870-023-04333-5.