The study engineered Tobacco rattle virus vectors incorporating distinct RNA secondary structures as mobility factors to improve guide RNA delivery to plant meristems. Using Nicotiana benthamiana plants expressing Cas9, optimal virus constructs were identified that generated both somatic and heritable edits, and these constructs were successfully applied to edit the emerging oilseed crop pennycress (Thlaspi arvense).

The authors introduce the ENABLE(R) Gene Editing in planta toolkit, a streamlined two‑step cloning system for creating CRISPR/Cas9 knockout vectors suitable for transient or stable transformation. Validation was performed in Oryza sativa protoplasts and Arabidopsis thaliana plants, and the toolkit includes low‑cost protocols aimed at facilitating adoption in the Global South.

The authors created a fast‑cycling, isogenic barley line (GP‑rapid) by introgressing the wild‑type Ppd‑H1 allele from Igri into the Golden Promise cultivar and performing two backcrosses to limit the donor genome, achieving a 25% reduction in generation time under speed‑breeding conditions while retaining high transformation efficiency. CRISPR/Cas9‑mediated editing of Ppd‑H1 showed regeneration and transformation rates comparable to the original Golden Promise, establishing GP‑rapid as a rapid platform for transgenic and gene‑edited barley research.

The study applied CRISPR/Cas9 gene editing to Physalis peruviana to modify plant‑architecture genes and create a compact growth ideotype. This compact phenotype is intended to increase per‑plot yield and support future breeding efforts for this nutritionally valuable minor crop.

The study examined how Arabidopsis calcium‑dependent protein kinases AtCPK5 and AtCPK6 modulate immunity triggered by bacterial rhamnolipids, finding that RLs up‑regulate these kinases and that mutants, especially cpk5/6, show altered reactive oxygen species production and defense gene expression. However, these kinases did not influence RL‑induced electrolyte leakage or resistance to Pseudomonas syringae pv. tomato DC3000, indicating additional signaling components are involved.

Using a microfluidic valve rootchip, the study simultaneously tracked ROS and calcium dynamics in compressed roots and found three kinetic phases linking mechanosensitive channel activity, NADPH oxidase‑dependent ROS accumulation, and secondary calcium influx. Pharmacological inhibition revealed that a fast calcium response is mediated by plasma‑membrane mechanosensitive channels, while a slower calcium increase is driven by ROS production.

A comparative phosphoproteomics study using Arabidopsis thaliana ahk5 loss‑of‑function mutants and wild‑type seedlings revealed that the histidine kinase AHK5 mediates a rapid shift from multistep phosphorelay signaling to serine/threonine phosphorylation in response to H2O2. AHK5 controls ROS‑responsive phosphorylation of plasma‑membrane nanodomain proteins and orchestrates distinct ABA‑independent stomatal closure and ABA‑dependent root development pathways by modulating key components such as RBOHD, CAS, HPCA1, and auxin transporters.

The study used CRISPR/Cas9 to edit the downstream region of the Arabidopsis thaliana FLOWERING LOCUS T (FT) gene, identifying a 2.3‑kb segment containing the Block E enhancer as crucial for normal FT expression and flowering. Fine‑scale deletions pinpointed a 63‑bp core module with CCAAT‑ and G‑boxes, and revealed a cryptic CCAAT‑box that becomes active when repositioned, highlighting the importance of local chromatin context and motif arrangement for enhancer function.

The study optimized three wheat transformation methods—immature embryo, callus, and in planta injection—by systematically adjusting Agrobacterium strain, bacterial density, acetosyringone concentration, and incubation conditions, achieving transformation efficiencies up to 66.84%. Using these protocols, CRISPR/Cas9 knockout of the negative regulator TaARE1-D produced mutants with increased grain number, spike length, grain size, and a stay‑green phenotype, demonstrating the platform’s potential to accelerate yield and stress‑tolerance improvements in wheat.

The study demonstrates that calcium-dependent protein kinases NbCDPK4 and NbCDPK5 directly phosphorylate the NADPH oxidase NbRBOHB at Ser‑123, enhancing sustained ROS production during effector-triggered immunity in Nicotiana benthamiana. Constitutively active CDPKs also upregulate NbRBOHB transcription, and phosphorylation of Ser‑123 is amplified by Ca2+ influx triggered by an autoactive helper NLR (NRC4). These results define a NbCDPK‑NbRBOHB signaling module that links NLR activation to prolonged ROS bursts in ETI.