Molecular modulation of the brassinosteroids pathway in durum wheat to enhance abiotic stress resilience
Author(s)
Date Issued
2025
Type
conferenceObject
Abstract
Durum wheat (Triticum turgidum spp. durum) is a staple crop in the Mediterranean region, which is currently suffering yield losses due to extreme events related to climate change. This study aims to enhance resilience in durum wheat cv. Svevo by modulating brassinosteroid (BR) phytohormone signaling, which is crucial for plant development under stress conditions.
Using a genome editing approach, the transcription factor IBH1, involved in BR biosynthesis, was knocked out.
Immature embryos were transformed via Agrobacterium tumefaciens carrying CRISPR/Cas9 constructs targeting IBH1. Two independent transformation experiments were performed using constructs with different guide RNA pairs: sgRNA-1 and sgRNA-3 for the first set, and sgRNA-2 and sgRNA-4 for the second. Both constructs were assembled using the Golden Gate (MoClo) method for efficiency and modularity.
Transgenesis was confirmed, and transgene copy number was analysed by TaqMan qPCR, identifying 43 T₀ transgenic plants, of which 18 carried a single-copy transgene insertion. Next-generation sequencing revealed five lines with editing events in both homeologs. sgRNA-3 showed the highest editing efficiency, as predicted in silico. All events displayed a single-base deletion at the expected cut site, inducing a frameshift likely causing loss of function.
These edited lines were taken to the T1 and T2 generations. In the T2 progeny, gene expression and phenotypic analyses are ongoing to evaluate the effects of IBH1 knock-out on BR-related transcription and stress responses. Oxidative stress assays will be performed to evaluate the tolerance of edited genotypes to environmental stresses.