Dihydroflavonol 4-reductase (DFR) significantly influences the modification of flower color. To explore the role of DFR in the synthesis of strawberry anthocyanins, in this study, we downloaded the CDS sequences of the DFR gene family from the Arabidopsis genome database TAIR; the DFR family of forest strawberry was compared; then, a functional domain screen was performed using NCBI; the selected strawberry DFR genes were analyzed; and the expression characteristics of the family members were studied by qRT-PCR. The results showed that there are 57 members of the DFR gene family in strawberry, which are mainly expressed in the cytoplasm and chloroplast; most of them are hydrophilic proteins; and the secondary structure of the protein is mainly composed of α-helices and random coils. The analysis revealed that FvDFR genes mostly contain light, hormone, abiotic stress, and meristem response elements. From the results of the qRT-PCR analysis, the relative expression of each member of the FvDFR gene was significantly different, which was expressed throughout the process of fruit coloring. Most genes had the highest expression levels in the full coloring stage (S4). The expression of FvDFR30, FvDFR54, and FvDFR56 during the S4 period was 8, 2.4, and 2.4 times higher than during the S1 period, indicating that the DFR gene plays a key role in regulating the fruit coloration of strawberry. In the strawberry genome, 57 members of the strawberry DFR gene family were identified. The higher the DFR gene expression, the higher the anthocyanin content, and the DFR gene may be the key gene in anthocyanin synthesis. Collectively, the DFR gene is closely related to fruit coloring, which lays a foundation for further exploring the function of the DFR gene family.

Sucrose phosphate synthase (SPS) is an important link in the process of sugar metabolism. In addition, it is also involved in abiotic stresses in plants. In order to study the SPS gene family and its role in abiotic stress, we identified the MdSPS gene family members by bioinformatics methods such as correlation analysis, the HMM method, and the Clustering method, and analyzed the transient expression of MdSPS genes by quantitative real-time fluorescence analysis (qRT-PCR). The MdSPS gene family consists of a total of 19 members divided into three subfamilies distributed on 14 chromosomes in apples. The MdSPS gene family has 12 collinearity gene pairs, indicating significant duplication. Most members of this family contain a large number of plant hormone response elements, light-inducible elements, and abiotic stress response elements 2kb upstream of the promoter. Codon bias analysis shows that there are 28 high-frequency codons and no codons with strong preference in this family. Gene chip results showed that only MdSPS2, MdSPS3, MdSPS11, and MdSPS17 were up-regulated in roots, and they were all members of subfamily C. The qRT-PCR analysis showed that all members of this family responded significantly to drought stress, salt stress, and low temperature stress. Interestingly, the relative expression of MdSPS12 was significantly down-regulated under salt stress and low temperature stress. In addition, the expression of MdSPS3, MdSPS8, MdSPS11, and MdSPS17 was more than 20 fold higher than that of the control under drought stress, salt stress, and low temperature stress. These four genes could be candidates for molecular breeding in the MdSPS family.

Flesh mealiness, a textural disorder in apples, reduces storage quality and consumer acceptance. The ‘Delicious’ and ‘Fuji’, prominent apple cultivars in China, exhibit contrasting susceptibility to mealiness, though the underlying mechanisms remain unclear. This study compared cytological, physiological and cell wall metabolic changes between mealy ‘Oregon Spur II Delicious’ and non-mealy ‘Miyazaki Spur Fuji’ during ambient storage. Toluidine blue staining and scanning electron microscopy revealed that ‘Delicious’ exhibited larger intercellular spaces and cell separation in contrast to ‘Fuji’. This observation aligns with the earlier onset of mealiness in ‘Delicious’: its mealiness degree increased from 3.06% at harvest to 19.62% after 28 d of storage (a 6.4-fold rise), whereas that of ‘Fuji’ only increased from 2.13% to 3.90% (1.8-fold). This pronounced increase in ‘Delicious’ was accompanied by a significant increase in air space volume and a reduction in expressible juice. Furthermore, the occurrence of mealiness in ‘Delicious’ involved a sharp increase in respiration rate and ethylene production, alongside rapid declines in firmness and starch content. Notably, there was a substantial accumulation of water-soluble pectin (WSP) and chelator-soluble pectin (CSP) in ‘Delicious’, whereas the content of Na₂CO₃-soluble pectin (NSP) remained consistently lower. Monosaccharide composition analysis confirmed significantly reduced arabinose and galactose levels across pectin fractions (WSP, CSP, and NSP) in ‘Delicious’. Correspondingly, immunofluorescence labeling showed a pronounced degradation of arabinan and galactan within the side chains of rhamnogalacturonan-I (RG-I). In addition, the activities of pectin methylesterase, α-L-Arabinofuranosidase, and β-D-Galactosidase remained significantly elevated in ‘Delicious’. Collectively, these findings demonstrate that cultivar differences in flesh mealiness are attributable to divergent physiological senescence and cell wall disassembly processes.

The calcium ion-transporting ATPase (Ca²⁺-ATPase) gene family maintains plant intracellular Ca²⁺ homeostasis and regulates growth, development and stress immunity; however, its functions remain poorly characterized in Malus domestica. Here, we performed a genome-wide identification of apple Ca²⁺-ATPase genes and obtained 45 members, which were classified into MdACA (39) and MdECA (6) subfamilies and unevenly distributed on 14 chromosomes. Phylogenetic analysis of Ca²⁺-ATPase genes from Malus domestica, Arabidopsis thaliana, and Oryza sativa classified these proteins into five subgroups. The ACA and ECA subfamilies were highly conserved across species, whereas Group D was apple-specific. Collinearity and Ka/Ks analyses indicated that segmental duplication and purifying selection dominated the evolution of apple Ca²⁺-ATPase genes. Promoter cis-element prediction uncovered numerous regulatory elements related to phytohormone signaling, growth, development and stress defense. Codon usage bias analysis indicated that AUG (methionine) was the dominant codon. Tissue expression profiles showed differential expression of apple Ca²⁺-ATPase genes in various organs. Quantitative real-time PCR (qRT-PCR) assays demonstrated widespread responses of Ca²⁺-ATPase genes to Alternaria alternata infection, exogenous CaCl₂, salicylic acid (SA) and methyl jasmonate (MeJA), among which MdACA39 was strongly induced under all treatments. Subcellular localization verified that MdACA39 resides on the plasma membrane. Moreover, transient overexpression of MdACA39 significantly enhanced apple resistance to A. alternata, likely due to the activation of SA, MeJA and Ca²⁺ signaling-mediated immune pathways, the induction of disease resistance-related genes, and elevated antioxidant enzyme activity. Collectively, this study systematically characterizes the apple Ca²⁺-ATPase family and identifies MdACA39 as a key regulator of fungal resistance, providing valuable gene resources for dissecting Ca²⁺ signaling-mediated disease resistance in apple.

Long-term clean tillage in apple orchards on the Loess Plateau has degraded soil properties and diminished fruit quality, highlighting the urgent need for sustainable cover cropping strategies. In a two-year field experiment (two consecutive growing seasons), eight green manure species (annual ryegrass, perennial ryegrass, white clover, smooth vetch, common vetch, rapeseed, hairy vetch, and cocksfoot) were compared with a clean-tillage control. Each treatment was replicated three times, and each plot was 100 m². Soil physicochemical properties, microbial communities (16S rRNA and ITS amplicon sequencing), and fruit quality parameters were systematically assessed. All grass treatments significantly lowered soil pH and increased soil organic matter and total nitrogen, phosphorus, and potassium contents. Hairy vetch (T7) produced the highest soil organic matter (25.86 g kg⁻¹) and total nitrogen (1.43 g kg⁻¹), while rapeseed (T6) reduced pH most markedly (4.28%). White clover (T3) enhanced bacterial and fungal alpha-diversity and enriched beneficial genera (Sphingomonas, Lysobacter, Mortierella, Chaetomium). However, functional predictions based on 16S/ITS data using FAPROTAX and Tax4Fun (PICRUSt-type predictive tools) suggested a potential increase in chemoorganoheterotrophy and saprotrophy; these predicted functions should be interpreted as hypothesis-generating rather than as direct evidence of microbial function. Common vetch (T5) increased soluble sugar content by 28.1%, and cocksfoot (T8) decreased titratable acidity by 30.8%. The comprehensive analysis indicates that hairy vetch and white clover as the most effective species for improving soil fertility, microbial diversity, and fruit quality. We therefore recommend these two legumes as optimal cover crops for sustainable orchard floor management in rain-fed semi-arid regions of the Loess Plateau.