Boron (B) toxicity is an important abiotic stress that drastically damages agricultural production worldwide, mostly in arid regions. Several studies have reported large losses in wheat due to high B in arable land. The identification of different B-toxicity-tolerant wheat germplasm and using them in breeding programs to develop tolerant cultivars is a potential solution to B toxicity. However, tolerance to B toxicity in such germplasm largely relies on the molecular changes in plants under B toxicity at the transcriptome level. Thus, the aim of this study is to determine the transcriptomic response of the roots of a B-toxicity-tolerant Triticum zhukovskyi genotype, PI296968, grown in a highly toxic B environment (10 mM B) in comparison with a control (3.1 μM B) treatment. T. zhukovskyi is a hexaploid wheat species forming a separate lineage from the main wheat lineage and can be a good source of genes for various stresses. However, it has been hardly explored for tolerance to any abiotic stress condition. This study is the first in the literature reporting the B toxicity tolerance of a T. zhukovskyi genotype along with the molecular changes occurring in it under B toxicity as compared to the control treatment. In the present study, 5992 genes were found to be significantly differentially expressed, with 1679 and 4313 up- and down-regulated genes, respectively. A number of transcription factors and pathways were identified to be significantly involved in the B toxicity response of the T. zhukovskyi genotype. A total of 12582 novel transcripts were determined in the study, with 9238 and 3344 coding and noncoding transcripts, respectively. The results not only suggest several candidate genes that can be further studied to improve wheat tolerance to B toxicity in upcoming breeding programs, but also enhance the understanding of the regulatory and molecular processes behind the wheat response to B toxicity. Further experiments are suggested to functionally characterize the identified high-B-responsive genes to confirm their role in providing B toxicity tolerance to the plants.