Modulation of brassinosteroid biosynthesis by genome editing: advancing sustainability and resilience in durum wheat production
Author(s)
Date Issued
2024
Type
conferenceObject
Abstract
Durum wheat (Triticum turgidum spp durum) is a staple cereal wide cultivated in countries surrounding the Mediterranean Basin. In recent years, this region has been particularly affected by extreme climatic conditions impairing crop yields. The aim of this work is to increase the resilience of durum wheat to stress by modulating the accumulation of brassinosteroid (BR) phytohormones, which are crucial for regulating plant development and stress responses. Using a genome editing approach in durum wheat, two transcription factors involved in brassinosteroid biosynthesis pathway, IBH1 and GATA7, were knocked out.
Fourteen days post-anthesis embryos from the Svevo variety were transformed using Agrobacterium tumefaciens engineered to deliver four different constructs: two for IBH1 knock-out and two for GATA7 knock-out, each with a pair of guides. The constructs were cloned using the Golden Gate (MoClo) method, which offers efficiency and flexibility. The final constructs included five essential components for transformation and regeneration: a hygromycin resistance gene, Cas9 nuclease under the control of rice ubiquitin promoter, small guide RNAs under the ubiquitin-6 promoter of T. aestivum, and a growth regulation factor.
Transformation efficiency was tested through copy number PCR screening, using primer designed for the Hygromycin resistance gene (hpt), revealing 43 T0 plants positive for IBH1 construct insertion and 44 plants for the GATA7 construct insertion. Respectively, 18 and 15 plants contained a single insertion copy.
Sequencing by NGS and Sanger methods is ongoing for IBH1 and GATA7, respectively, to investigate deletion events. The edited durum wheat lines are expected to exhibit improved agronomic traits, such as increased yield and enhanced resilience to abiotic stresses like heat and drought, challenges that are intensifying due to climate change.
Biochemical assays and phenotyping will be performed on the edited lines to evaluate productivity trends and resilience against abiotic stress. Additionally, molecular analyses will be conducted to deeply characterize the edited plants and to deepen the role of IBH1 and GATA7 in durum wheat.
Conference(s)
Expanding frontiers in crop genetics - siga 2024