This study presents the first integrated analysis of genotype–medium interactions and temporal morphogenesis profiling in sunflower regeneration. It aims to characterize genotype-specific responses, identify predictive morphological markers, and develop a scalable framework for breeding and transformation. Eighteen sunflower genotypes were evaluated to assess organogenic performance. The model genotype Ha-26-PR was used for a complementary experiment, testing varying sucrose concentrations to examine their influence on morphogenic outcomes. Hierarchical Cluster Analysis (HCA), guided by the Elbow method, identified four optimal clusters (K = 4). These aligned with three biologically meaningful categories: High Regenerators (Cluster 1), Moderate/Specific Regenerators (Clusters 2 and 3), and Non-Regenerators (Cluster 4). On S1 medium, NO-SU-12 and AS-1-PR showed superior shoot regeneration, while on R4 medium, HA-26-PR-SU and NO-SU-12 performed best. Genotypes such as NO-SU-12 and AS-1-PR consistently excelled across both media, whereas AB-OR-8 and FE-7 remained non-regenerators. Medium R4 supported superior regeneration, primarily through root formation, while S1 failed to induce roots in any genotype, highlighting the importance of hormonal composition. Although sucrose promoted callus induction, it did not trigger organogenesis. Callus was consistently present across media and time points, but its correlations with shoot and root formation were weak and temporally unstable, limiting its predictive value. Root formation at 14 days (Root 14D) emerged as a robust early predictor of organogenic success. This integration of morphological, temporal, and statistical analyses offers a genotype-tailored regeneration framework with direct applications in molecular breeding and CRISPR/Cas-based genome editing.

Iron (Fe) is an essential micronutrient for plant development and productivity. Nevertheless, the role of gibberellins (GAs) in the control of iron homeostasis is less studied compared to other growth regulators. We found that GAs modulate iron homeostasis in maize by inducing deficiency-like responses independent of rhizosphere iron availability. Plant phenotyping demonstrated that exogenous GA₃ application under iron-sufficient conditions phenocopied iron deprivation, while inhibiting GA biosynthesis with mepiquat chloride prevented the development of typical symptoms of Fe deficiency (–Fe). Gibberellins positively control strategy II Fe uptake genes, albeit indirectly, as opposed to the direct negative transcriptional regulation of phytosiderophore biosynthesis. Additionally, gibberellins disrupt iron partitioning by suppressing root-to-shoot Fe translocation, causing iron overaccumulation in roots of GA₃ treated plants. A functional ferrous iron uptake pathway was identified and was found to operate in conjunction with the strategy II uptake pathway via the differentially regulated Zea mays Iron-Regulated Transporter (IRT) paralogs ZmIRT1 and ZmIRT2. Root responses are spatially organized: gene expression in the lateral root sector reflects the shoot iron status, while transcriptional responses in the root apex correlate with local Fe demands. This study demonstrates that maize leverages a hybrid ferric/ferrous iron uptake strategy and establishes novel roles of GAs as pivotal regulators of iron homeostasis.

The observed longer and more frequent periods of drought in recent years have drawn more focus on efficient breeding techniques for drought-tolerant crops. Several systems have been used to simulate drought conditions in order to select drought-tolerant lines. However, little is known about the comparability of different screening methods. We compared four different systems in terms of the information provided on drought tolerance, namely, two in vitro systems (liquid and solid medium applying polyethylene glycol as the drought simulant), and two soil-based pot trials were conducted either in a green house or in a climate chamber. Two sunflower inbred lines differing in drought tolerance (AB-OR-8 and DF-AB-2) were characterized at the seedling stage using physiological and morphological parameters of shoots and roots. Transcriptomic analysis (RNA-seq) and quantitative RT-PCR were performed to compare drought responses and stress levels applied by the different systems. In an assessment of all four screening methods, the expressions of five of six selected genes in the ABA signaling pathway were differentially upregulated during drought, while one of them was downregulated. These genes represent valuable indicators of the stress level applied by a drought-screening method. The advantages and disadvantages of different drought-simulating systems were evaluated and combined with next-generation sequencing data to identify the most efficient selection approach for assessing drought tolerance in sunflower. Overall, all systems enabled effective pre-screening of the breeding material for drought tolerance, where the decision for a system depends on experimental goals and resource availability. If the goal is to maximize the molecular resolution of drought stress, the in vitro liquid system is the most effective.

Sunflower in one of the most important oil-crops worldwide. Due to the effects of climate change, it is becoming increasingly important to discover effective and reliable ways to assist it in combating abiotic stresses. A very precise, cost-efficient and reliable way to achieve sunflower abiotic stress resilience, and specifically drought resilience, is through genetic modification. Due to sunflower’s recalcitrant nature to transformation and regeneration and strong genotype dependency, it is important to establish a high efficiency regeneration protocol applicable to a wide range of genotypes. The aim of this study was to evaluate the root regeneration capacity of 4 different sunflower genotypes when cultured on 5 different rooting media with the purpose of identifying a combination of sunflower genotype and medium with the highest root regeneration rate. The percentage of rooting of different genotypes showed inter-assay variability (from 0% regeneration to 100% regeneration), however, most consistent root formation was repeatedly observed for genotypes PH-BC2-91 and HA-26-PR. Statistically significant higher root regeneration rates were observed when explants of different genotypes were cultured in MS DV medium compared to four other media used (from around 90% for MS DV to around 50% for the other media). These results further verify the intricate relationship between genotype and medium interactions. The study provides a reliable basis for establishing an efficient sunflower root regeneration protocol and paves the way for the application of biotechnological approaches in breeding and enhancing resilience to abiotic stresses.