Drought is one of the major environmental stresses limiting crop growth and production. MYB family transcription factors play crucial roles in response to abiotic stresses. Previous studies found that TaMYB31 is transcriptionally induced by drought stress. However, the biological functions of TaMYB31 in drought stress responses remained unknown. In this study, three TaMYB31 homoeologous genes from hexaploid wheat, designated TaMYB31-A, TaMYB31-B, and TaMYB31-D, were cloned and characterized. Expression analysis showed that TaMYB31 genes have different tissue expression patterns, and TaMYB31-B has relatively high expression levels in most tested tissues. All the three homoeologs were up-regulated by polyethylene glycol (PEG) 6000 and abscisic acid (ABA) treatments. Subcellular localization analyses revealed that TaMYB31 is localized to the nucleus. Ectopic expression of the TaMYB31-B gene in Arabidopsis affected plants growth and enhanced drought tolerance. In addition, seed germination and seedling root growth of TaMYB31-B transgenic plants were more sensitive to exogenous ABA treatment compared to wild type control. RNA-seq analysis indicated that TaMYB31 functions through up-regulation of wax biosynthesis genes and drought-responsive genes. These results provide evidence that TaMYB31 acts as a positive regulator of drought resistance, and justify its potential application in genetic modification of crop drought tolerance.

In plants, the HAK (high-affinity K⁺)/KUP (K⁺ uptake)/KT (K⁺ transporter) family represents a large group of potassium transporters that play important roles in plant growth and environmental adaptation. Although HAK/KUP/KT genes have been extensively investigated in many plant species, they remain uncharacterized in wheat, especially those involved in the response to environmental stresses. In this study, 56 wheat HAK/KUP/KT (hereafter called TaHAKs) genes were identified by a genome-wide search using recently released wheat genomic data. Phylogenetic analysis grouped these genes into four clusters (Ι, II, III, IV), containing 22, 19, 7 and 8 genes, respectively. Chromosomal distribution, gene structure, and conserved motif analyses of the 56 TaHAK genes were subsequently performed. In silico RNA-seq data analysis revealed that TaHAKs from clusters II and III are constitutively expressed in various wheat tissues, while most genes from clusters I and IV have very low expression levels in the examined tissues at different developmental stages. qRT-PCR analysis showed that expression levels of TaHAK genes in wheat seedlings were significantly up- or downregulated when seedlings were exposed to K⁺ deficiency, high salinity, or dehydration. Furthermore, we functionally characterized TaHAK1b-2BL and showed that it facilitates K⁺ transport in yeast. Collectively, these results provide valuable information for further functional studies of TaHAKs, and contribute to a better understanding of the molecular basis of wheat development and stress tolerance.

The relatively low efficiency of biolistic transformation and subsequent integration of multiple copies of the introduced gene/s significantly complicate the genetic modification of wheat (Triticum aestivum) and other plant species. One of the key factors contributing to the reproducibility of this method is the uniformity of the DNA/gold suspension, which is dependent on the coating procedure employed. It was also shown recently that the relative frequency of single copy transgene inserts could be increased through the use of nanogram quantities of the DNA during coating. A simplified DNA/gold coating method was developed to produce fertile transgenic plants, via microprojectile bombardment of callus cultures induced from immature embryos. In this method, polyethyleneglycol (PEG) and magnesium salt solutions were utilized in place of the spermidine and calcium chloride of the standard coating method, to precipitate the DNA onto gold microparticles. The prepared microparticles were used to generate transgenics from callus cultures of commercial bread wheat cv. Gladius resulting in an average transformation frequency of 9.9%. To increase the occurrence of low transgene copy number events, nanogram amounts of the minimal expression cassettes containing the gene of interest and the hpt gene were used for co-transformation. A total of 1538 transgenic wheat events were generated from 15,496 embryos across 19 independent experiments. The variation of single copy insert frequencies ranged from 16.1 to 73.5% in the transgenic wheat plants, which compares favourably to published results. The DNA/gold coating procedure presented here allows efficient, large scale transformation of wheat. The use of nanogram amounts of vector DNA improves the frequency of single copy transgene inserts in transgenic wheat plants.