The aim of this study was to determine the effects and underlying molecular mechanisms of humic acid (HA) on foxtail millet (Setaria italica Beauv.) under drought conditions. The rainless climate of the Shanxi Province (37°42'N, 112°58'E) in China provides a natural simulation of drought conditions. Two foxtail millet cultivars, Jingu21 and Zhangza10, were cultivated in Shanxi for two consecutive years (2017–2018) based on a split-plot design. Plant growth, grain quality, and mineral elements were analyzed in foxtail millet treated with HA (50, 100, 200, 300, and 400 mg L^-1) and those treated with clear water. Transcriptome sequencing followed by bioinformatics analysis was performed on plants in the normal control (CK), drought treatment (D), and drought + HA treatment (DHA) groups. Results were verified using real-time quantitative PCR (RT-qPCR). HA at a concentration of 100–200 mg L^-1 caused a significant increase in the yield of foxtail millet and had a positive effect on dry weight and root-shoot ratio. HA also significantly increased P, Fe, Cu, Zn, and Mg content in grains. Moreover, a total of 1098 and 409 differentially expressed genes (DEGs) were identified in group D vs. CK and D vs. DHA, respectively. A protein-protein interaction network and two modules were constructed based on DEGs (such as SETIT_016654mg) between groups D and DHA. These DEGs were mainly enriched in the metabolic pathway. In conclusion, HA (100 mg L^-1) was found to promote the growth of foxtail millet under drought conditions. Furthermore, SETIT_016654mg may play a role in the effect of HA on foxtail millet via control of the metabolic pathway. This study lays the foundation for research into the molecular mechanisms that underlie the alleviating effects of HA on foxtail millet under drought conditions.

Glutathione S-transferases (GSTs) are a critical superfamily of multifunctional enzymes in plants. As a ligand or binding protein, GSTs regulate plant growth and development and detoxification. Foxtail millet (Setaria italica (L.) P. Beauv) could respond to abiotic stresses through a highly complex multi-gene regulatory network in which the GST family is also involved. However, GST genes have been scarcely studied in foxtail millet. Genome-wide identification and expression characteristics analysis of the foxtail millet GST gene family were conducted by biological information technology. The results showed that 73 GST genes (SiGSTs) were identified in the foxtail millet genome and were divided into seven classes. The chromosome localization results showed uneven distribution of GSTs on the seven chromosomes. There were 30 tandem duplication gene pairs belonging to 11 clusters. Only one pair of SiGSTU1 and SiGSTU23 were identified as fragment duplication genes. A total of ten conserved motifs were identified in the GST family of foxtail millet. The gene structure of SiGSTs is relatively conservative, but the number and length of exons of each gene are still different. The cis-acting elements in the promoter region of 73 SiGST genes showed that 94.5% of SiGST genes possessed defense and stress-responsive elements. The expression profiles of 37 SiGST genes covering 21 tissues suggested that most SiGST genes were expressed in multiple organs and were highly expressed in roots and leaves. By qPCR analysis, we found that 21 SiGST genes were responsive to abiotic stresses and abscisic acid (ABA). Taken together, this study provides a theoretical basis for identifying foxtail millet GST family information and improving their responses to different stresses.

Field experiments were conducted to investigate the effects of K application on the nitrogen and potassium utilization efficiency and yield of foxtail millet (Setaria italica L.). The experiment was performed with a completely randomized design with two millet cultivars (Jingu 21 and Zhangza 10) and five K₂O rates (0, 60, 120, 180, and 240 kg/hm²) in 2020 and 2021. We found that K promoted K and N absorption; significantly increased dry matter (DM), N, and K accumulation in millet organs; caused dry matter accumulation to peak earlier; and increased the DM accumulation rate. In addition, K accumulation preceded that of DM or N. Relative to the crop cycle, most K (61.07%) accumulated at booting, whereas N accumulated mostly (33.86%) during grain filling. N absorption efficiency increased by 31.87%, and the apparent and agronomic utilization rate of K fertilizer remained high, increasing millet yield, which peaked at a K rate of 180 kg/hm² for both cultivars, by 29.91% and 31.51% in Jingu 21 and Zhangza 10, respectively, relative to untreated controls. Stepwise regression and path analysis showed that the leaf and spike K accumulation, stem N accumulation, and stem DM were the main factors affecting yield, with DM having the greatest direct effect, followed by leaf K accumulation. The K concentration (0.77–3.04%) in Zhangza 10 was higher than that in Jingu 21 (0.69–2.91%) in untreated plants. Under the same K application rate, N and K accumulation and the harvest index were higher for Zhangza 10 than those for Jingu 21, and the nutrient utilization ability was greater for Zhangza 10. The results demonstrated that rational K fertilizer application can increase K concentration and accumulation in leaves, promote N metabolism and accumulation, increase N and K utilization efficiency, and improve DM accumulation and millet yield.

Conventional fertilizer management can destroy the structure of soil. Replacing chemical fertilizers with organic fertilizers can improve soil quality and nitrogen use efficiency. We aimed to study the effects of organic fertilizer substitutions for chemical nitrogen fertilizer on soil fertility and nitrogen use efficiency in order to clarify the effectiveness of the available nutrient management measures in improving soil quality and increasing foxtail millet yield. A field experiment was carried out over two consecutive years, and a total of six treatments were set up: no fertilizer (CK), chemical nitrogen fertilizer alone (N), the substitution of 25% of chemical nitrogen fertilizer with bio-organic fertilizer (N25A1), the substitution of 25% of chemical nitrogen fertilizer with fermented mealworm manure (N25B1), the substitution of 50% of chemical nitrogen fertilizer with bio-organic fertilizer (N50A2), and the substitution of 50% of chemical nitrogen fertilizer with fermented mealworm manure (N50B2). The results of this study show the following: (1) Compared with chemical nitrogen fertilizer, the substitution of organic fertilizer for nitrogen fertilizer reduced the bulk density and solid phase of the soil, and it increased the total porosity, water content, liquid phase, and gas phase of the soil. (2) Compared with nitrogen fertilizer, the use of an organic fertilizer increased the contents of nitrate nitrogen, ammonium nitrogen, and total nitrogen in the soil by 13.59~52.56%, 4.47~18.27%, and 4.40~12.09%, respectively. The content of alkaline nitrogen increased by 1.70~32.37%, and the contents of soil available potassium, available phosphorus, and organic matter also increased. (3) The activities of sucrase, urease, glutaminase, and asparaginase were improved by replacing chemical nitrogen fertilizer with organic fertilizer. The N25 treatments performed better than the N50 treatments, and fermented mealworm manure performed better than biological organic fertilizer. (4) A moderate application of organic fertilizer (N25) can increase the grain yield, ear weight, grain weight, and 1000-grain weight of foxtail millet, whereas excessive application of organic fertilizer (N50) can reduce foxtail millet yield. (5) Replacing chemical nitrogen fertilizer with organic fertilizer can improve the agronomic use efficiency, physiological efficiency, biased productivity, harvest index, and apparent use efficiency of nitrogen fertilizer. In this study, the substitution of 25% of chemical nitrogen fertilizer with fermented mealworm manure was the best combination for restoring crop productivity and soil quality.

Grain development of foxtail millet (Setaria italica L.) is essential for yield and quality. However, its transcriptional dynamics molecular mechanisms and morphological analyses remain scarcely described. Thus, we conducted detailed daily morphological analyses of foxtail millet grain development throughout the 30 d post-fertilization development period. On the basis of the morphological analyses, we used RNA-sequencing (RNA-seq) to examine the transcript dynamics involved in foxtail millet grain development at four stages. These genes included those associated with transcriptional regulation, hormone signaling, sucrose and starch metabolism, zein family members, amino acid metabolism, carotenoid metabolism, flavonoid biosynthesis, and folate synthesis. We have validated the accuracy of the transcriptome data by means of reverse transcription quantitative polymerase chain reaction (RT-qPCR). This study provides precious genetic resources for understanding grain developmental process in the future. These results expand our understanding of the molecular mechanisms of grain development in foxtail millet and contribute to the functional studies of genes related to grain development in the future.