Drought is one of the most critical environmental factors constraining maize production. When it occurs at the flowering stage, serious yield losses are caused, and often, the damage is irretrievable. In this study, anthesis to silk interval (ASI), plant height (PH), and ear biomass at the silking date (EBM) of 279 inbred lines were studied under both water-stress (WS) and well-water (WW) field conditions, for three consecutive years. Averagely, ASI was extended by 25.96%, EBM was decreased by 17.54%, and the PH was reduced by 12.47% under drought stress. Genome-wide association studies were carried out using phenotypic values under WS, WW, and drought-tolerance index (WS-WW or WS/WW) and applying a mixed linear model that controls both population structure and relative kinship. In total, 71, 159, and 21 SNPs, located in 32, 59, and 12 genes, were significantly (P < 10⁻⁵) associated with ASI, EBM, and PH, respectively. Only a few overlapped candidate genes were found to be associated with the same drought-related traits under different environments, for example, ARABIDILLO 1, glycoprotein, Tic22-like, and zinc-finger family protein for ASI; 26S proteasome non-ATPase and pyridoxal phosphate transferase for EBM; 11-ß-hydroxysteroid dehydrogenase, uncharacterised, Leu-rich repeat protein kinase, and SF16 protein for PH. Furthermore, most candidate genes were revealed to be drought-responsive in an association panel. Meanwhile, the favourable alleles/key variations were identified with a haplotype analysis. These candidate genes and their key variations provide insight into the genetic basis of drought tolerance, especially for the female inflorescence, and will facilitate drought-tolerant maize breeding.

Subtilisin-like proteases (subtilases) are found in almost all plant species and are involved in regulating various biotic and abiotic stresses. Although the literature on subtilases in different plant species is vast, the gene function of the serine peptidase S8 family and its maize subfamily is still unknown. Here, a bioinformatics analysis of this gene family was conducted by describing gene structure, conserved motifs, phylogenetic relationships, chromosomal distributions, gene duplications, and promoter cis-elements. In total, we identified 18 ZmSPS8 genes in maize, distributed on 7 chromosomes, and half of them were hydrophilic. Most of these proteins were located at the cell wall and had similar secondary and tertiary structures. Prediction of cis-regulatory elements in promoters illustrated that they were mainly associated with hormones and abiotic stress. Maize inbred lines B73, Zheng58, and Qi319 were used to analyze the spatial-temporal expression patterns of ZmSPS8 genes under drought treatment. Seedling drought results showed that Qi319 had the highest percent survival after 14 d of withholding irrigation, while B73 was the lowest. Leaf relative water content (LRWC) declined more rapidly in B73 and to lower values, and the nitrotetrazolium blue chloride (NBT) contents of leaves were higher in Qi319 than in the other inbreds. The qPCR results indicated that 6 serine peptidase S8 family genes were positively or negatively correlated with plant tolerance to drought stress. Our study provides a detailed analysis of the ZmSPS8s in the maize genome and finds a link between drought tolerance and the family gene expression, which was established by using different maize inbred lines.