Drought stress is one of the most important abiotic factors limiting crop productivity. A better understanding of the effects of drought on millet (Setaria italica L.) production, a model crop for studying drought tolerance, and the underlying molecular mechanisms responsible for drought stress responses is vital to improvement of agricultural production. In this study, we exposed the drought resistant F₁ hybrid, M79, and its parental lines E1 and H1 to drought stress. Subsequent physiological analysis demonstrated that M79 showed higher photosynthetic energy conversion efficiency and drought tolerance than its parents. A transcriptomic study using leaves collected six days after drought treatment, when the soil water content was about ∼20%, identified 3066, 1895, and 2148 differentially expressed genes (DEGs) in M79, E1 and H1 compared to the respective untreated controls, respectively. Further analysis revealed 17 Gene Ontology (GO) enrichments and 14 Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways in M79, including photosystem II (PSII) oxygen-evolving complex, peroxidase (POD) activity, plant hormone signal transduction, and chlorophyll biosynthesis. Co-regulation analysis suggested that these DEGs in M79 contributed to the formation of a regulatory network involving multiple biological processes and pathways including photosynthesis, signal transduction, transcriptional regulation, redox regulation, hormonal signaling, and osmotic regulation. RNA-seq analysis also showed that some photosynthesis-related DEGs were highly expressed in M79 compared to its parental lines under drought stress. These results indicate that various molecular pathways, including photosynthesis, respond to drought stress in M79, and provide abundant molecular information for further analysis of the underlying mechanism responding to this stress.

Drought is a major limiting factor affecting grain production. Drought-tolerant crop varieties are required to ensure future grain production. Here, 5597 DEGs were identified using transcriptome data before and after drought stress in foxtail millet (Setaria italica) hybrid Zhangza 19 and its parents. A total of 607 drought-tolerant genes were screened through WGCNA, and 286 heterotic genes were screened according to the expression level. Among them, 18 genes overlapped. One gene, Seita.9G321800, encoded MYBS3 transcription factor and showed upregulated expression after drought stress. It is highly homologous with MYBS3 in maize, rice, and sorghum and was named SiMYBS3. Subcellular localization analysis showed that the SiMYBS3 protein was located in the nucleus and cytoplasm, and transactivation assay showed SiMYBS3 had transcriptional activation activity in yeast cells. Overexpression of SiMYBS3 in Arabidopsis thaliana conferred drought tolerance, insensitivity to ABA, and earlier flowering. Our results demonstrate that SiMYBS3 is a drought-related heterotic gene and it can be used for enhancing drought resistance in agricultural crop breeding.

The water-driven AquaCrop model is used extensively for simulating crop growth and water use. A three-year field experiment (2015–2017) of foxtail millet (Setaria italica) that was grown using plastic film mulching (PM) and no mulching (NM) was conducted in a rain-fed region of China to simulate canopy cover (CC), biomass, soil water content (SWC), yield, evapotranspiration (ETc), and water use efficiency (WUE). The year 2015 was much drier and warmer than the two other years. The model was calibrated using field data from 2016 and validated using the data from 2015 and 2017. Simulations of CC, biomass, and yield achieved favorable performance for both PM and NM in all years, as indicated by the high determination coefficient (R²), model efficiency (EF), small root mean square error (RMSE), normalized root mean square error (NRMSE), and deviations < 10%. Simulations of SWC, ETc, and WUE gave acceptable results for both PM and NM in the normal year (2017). However, low R² and EF, and large NRMSE, RMSE, and deviations were observed in the predictions of PM and NM for SWC, ETc, and WUE in the dry year (2015) with a severe drought stress, indicating that the model performed unsatisfactorily under severe drought stress condition that was caused by the adverse weather. In addition, the simulation performance of NM was more favorable than that of PM for most crop growth and water use indexes under no drought stress condition.

Foxtail millet (Setaria italica L.) is cultivated around the world for human and animal consumption. There is no suitable herbicide available for weed control in foxtail millet fields during the post-emergence stage. In this study, we investigated the effect and safety of the post-emergence herbicide tribenuron-methyl (TBM) on foxtail millet in terms of grain yield and quality using a split-plot field design. Field experiments were conducted using two varieties in 2013 and 2014, i.e., high-yielding hybrid Zhangzagu 10 and high-quality conventional Jingu 21. TBM treatments at 11.25 to 90 g ai ha⁻¹ reduced root and shoot biomass and grain yield to varying degrees. In each of the two years, grain yield declined by 50.2% in Zhangzagu 10 with a herbicide dosage of 45 g ai ha⁻¹ and by 45.2% in Jingu 21 with a herbicide dosage of 22.5 g ai ha⁻¹ (recommended dosage). Yield reduction was due to lower grains per panicle, 1000-grain weight, panicle length, and panicle diameter. Grain yield was positively correlated with grains per panicle and 1000-grain weight, but not with panicles ha⁻¹. With respect to grain protein content at 22.5 g ai ha⁻¹, Zhangzagu 10 was similar to the control, whereas Jingu 21 was markedly lower. An increase in TBM dosage led to a decrease in grain Mn, Cu, Fe, and Zn concentrations. In conclusion, the recommended dosage of TBM was relatively safe for Zhangzagu 10, but not for Jingu 21. Additionally, the hybrid variety Zhangzagu 10 had a greater tolerance to TBM than the conventional variety Jingu 21.

NADP-malic enzyme (NAPD-ME), and pyruvate orthophosphate dikinase (PPDK) are important enzymes that participate in C₄ photosynthesis. However, the evolutionary history and forces driving evolution of these genes in C₄ plants are not completely understood. We identified 162 NADP-ME and 35 PPDK genes in 25 species and constructed respective phylogenetic trees. We classified NADP-ME genes into four branches, A1, A2, B1 and B2, whereas PPDK was classified into two branches in which monocots were in branch I and dicots were in branch II. Analyses of selective pressure on the NAPD-ME and PPDK gene families identified four positively selected sites, including 94H and 196H in the a5 branch of NADP-ME, and 95A and 559E in the e branch of PPDK at posterior probability thresholds of 95%. The positively selected sites were located in the helix and sheet regions. Quantitative RT-PCR (qRT-PCR) analyses revealed that expression levels of 6 NADP-ME and 2 PPDK genes from foxtail millet were up-regulated after exposure to light. This study revealed that positively selected sites of NADP-ME and PPDK evolution in C₄ plants. It provides information on the classification and positive selection of plant NADP-ME and PPDK genes, and the results should be useful in further research on the evolutionary history of C₄ plants.