The study presents an optimized Agrobacterium-mediated transformation toolkit for Sorghum bicolor that achieves up to 95.7% editing efficiency using CRISPR/Cas9 targeting the SbPDS gene, and demonstrates comparable performance with a PAM‑broadened SpRY variant. This platform enables multiplex genome editing and is positioned for integration of advanced tools such as prime and base editors to accelerate sorghum breeding.

The study applied single-nucleus RNA sequencing to mature Sorghum bicolor leaves under well‑watered and drought conditions, identifying major leaf cell types and their transcriptional responses. Drought induced transcriptomic changes that surpassed cell‑type differences, indicating a common response across mesophyll, bundle sheath, epidermal, vascular, and stomatal cells, and enabling the identification of candidate drought‑responsive regulators for improving water‑use efficiency.

The study shows that drought triggers ABA accumulation and JA reduction in sorghum roots, accompanied by transcriptional activation of genes linked to mineral homeostasis, hormone signaling, and osmotic regulation, while Fe supplementation enhances ferritin expression and mitigates oxidative stress. Drought also diminishes root bacterial diversity but enriches beneficial taxa such as Burkholderia, whereas fungal diversity remains stable, and functional profiling reveals shifts toward phototrophy, methylotrophy, and nitrate reduction. These findings highlight ferritin’s protective role and suggest specific bacterial inoculants for improving sorghum drought resilience.