Melon (Cucumis melo L.) is a protected crop in China with high economic value. Agrobacterium-mediated genetic transformation is a powerful tool to improve agronomic traits and obtain elite germplasm. However, current transformation protocols in melons are inefficient and highly genotype-dependent. To improve transformation in melon, we tested different infiltration methods for Agrobacterium-mediated transformation. Among these methods, micro-brushing and sonication for 20 s, followed by vacuum infiltration at −1.0 kPa for 90 s, resulted in the strongest green fluorescent protein signal and increased the proportion of infected explants. We transformed melon with developmental regulatory genes AtGRF5, AtPLT5, AtBBM, AtWUS, AtWOX5, and AtWIND1 from Arabidopsis and estimated regeneration frequencies as the number of regenerating shoots/total number of inoculated explants in the selection medium. The overexpression of AtGRF5 and AtPLT5 in melon resulted in transformation efficiencies of 42.3% and 33% in ZHF and 45.6% and 32.9% in Z12, respectively, which were significantly higher than those of the control. AtGRF5 and AtPLT5 expression cassettes were added to CRISPR/Cas9 genome-editing vectors to obtain transgenic phytoene desaturase CmPDS knockout mutants. Using AtGRF5 or AtPLT5, multi-allelic mutations were observed at CmPDS target sites in recalcitrant melon genotypes. This strategy enables genotype-flexible transformation and promotes precise genome modification technologies in melons.

Melon (Cucumis melo L.) represents an agriculturally significant horticultural crop that is widely grown for its flavorful fruits. Downy mildew (DM), a pervasive foliar disease, poses a significant threat to global melon production. Although several quantitative trait loci related to DM resistance have been identified, the comprehensive genetic underpinnings of this resistance remain largely uncharted. In this study, we utilized integrative transcriptomics and metabolomics approaches to identify potential resistance-associated genes and delineate the strategies involved in the defense against DM in two melon cultivars: the resistant ‘PI442177′ (‘K10-1′) and the susceptible ‘Huangdanzi’ (‘K10-9′), post-P. cubensis infection. Even in the absence of the pathogen, there were distinctive differentially expressed genes (DEGs) between ‘K10-1′ and ‘K10-9′. When P. cubensis was infected, certain genes, including flavin-containing monooxygenase (FMO), receptor-like protein kinase FERONIA (FER), and the HD-ZIP transcription factor member, AtHB7, displayed pronounced expression differences between the cultivars. Notably, our data suggest that following P. cubensis infection, both cultivars suppressed flavonoid biosynthesis via the down-regulation of associated genes whilst concurrently promoting lignin production. The complex interplay of transcriptomic and metabolic responses elucidated by this study provides foundational insights into melon’s defense mechanisms against DM. The robust resilience of ‘K10-1′ to DM is attributed to the synergistic interaction of its inherent transcriptomic and metabolic reactions.

The root-knot nematode (Meloidogyne incognita) poses a major threat to global agriculture by impairing root function, reducing nutrient uptake, and ultimately limiting seed development and crop productivity. This study investigated the molecular and metabolic defense responses of Cucumis metuliferus (prickly pear) to M. incognita infection. Gene expression and metabolic pathway reprogramming in M. incognita-infected roots were examined using integrated transcriptomics and metabolomics approaches. The identified genes were involved in stress responses and defense activation. Furthermore, metabolite profiling revealed significant shifts in secondary metabolite production, with an upregulation of defense-related compounds like jasmonic acid, salicylic acid, and prostaglandins. KEGG (Kyoto Encyclopedia of Genes and Genomes) pathway enrichment analysis highlighted critical pathways such as biotin metabolism and nucleotide metabolism, underscoring the adaptive metabolic responses of C. metuliferus plants. GO (Gene Ontology) analysis from the integrated transcriptomics and metabolomics data highlighted significant upregulation of enzymatic pathways, transporter activities, and reorganization of cellular structures. Furthermore, KEGG pathway analysis revealed activation of secondary metabolite biosynthesis, immune-related signaling pathways, and metabolic reprogramming including increased carbon metabolism and nucleotide biosynthesis. This study provides a valuable molecular framework for breeding of M. incognita-resistant cultivars, ultimately supporting more stable seed distribution and agricultural productivity in M. incognita-prone regions.

Plant vacuolar H⁺-ATPases (V-type H⁺-ATPases, VHA) and vacuolar H⁺-pyrophosphatases (V-PPase, VHP) are key proton pumps. VHA and VHP regulate cellular pH homeostasis and ion transport via the proton motive force across the vacuolar membrane, functioning in plant responses to environmental stress. However, they remain poorly characterized in melon, a high-value horticultural crop limited by environmental stresses. Here, a total of 24 V-type proton pump genes were identified from the whole melon genome. Collinearity analysis indicates that the proton pump genes are highly conserved in melons, although species-specific expansions and gene duplication events have been observed in the a and c subunits. Predictions of cis-acting elements in the promoters indicate that these genes are rich in regulatory elements associated with responding to plant hormones and abiotic stress. It was found that most genes were stimulated in response to salt treatment and exhibited consistent expression trends in both Elizabeth and Baishami. However, in Elizabeth, the responses of CmVHA-c″1 and CmVHA-a3 were slower, while the induction of CmVHP2;1 was almost undetectable in response to salt stress. This study provides a foundation for further functional analysis of proton pump genes and new insights into melon stress responses.