Simultaneous modification of three homoeologs of TaEDR1 by genome editing enhances powdery mildew resistance in wheat

Abstract

Wheat (Triticum aestivum L.) incurs significant yield losses from powdery mildew, a major fungal disease caused by Blumeria graminis f. sp. tritici (Bgt). enhanced disease resistance1 (EDR1) plays a negative role in the defense response against powdery mildew in Arabidopsis thaliana; however, the edr1 mutant does not show constitutively activated defense responses. This makes EDR1 an ideal target for approaches using new genome-editing tools to improve resistance to powdery mildew. We cloned TaEDR1 from hexaploid wheat and found high similarity among the three homoeologs of EDR1. Knock-down of TaEDR1 by virus-induced gene silencing or RNA interference enhanced resistance to powdery mildew, indicating that TaEDR1 negatively regulates powdery mildew resistance in wheat. We used CRISPR/Cas9 technology to generate Taedr1 wheat plants by simultaneous modification of the three homoeologs of wheat EDR1. No off-target mutations were detected in the Taedr1 mutant plants. The Taedr1 plants were resistant to powdery mildew and did not show mildew-induced cell death. Our study represents the successful generation of a potentially valuable trait using genome-editing technology in wheat and provides germplasm for disease resistance breeding.

The CRISPR/Cas revolution continues: From efficient gene editing for crop breeding to plant synthetic biology

“…And more recently, Wang et al () rendered grapevine resistant to Botrytis cinerea through the knockout of the transcription factor WRKY52 . The establishment of powdery mildew resistance in wheat has been among the agriculturally most relevant achievements using RNA‐guided Cas endonucleases (Zhang et al ). In this study, all three EDR1 homeoalleles of hexaploid wheat were disrupted.…”

The Enhancement of Plant Disease Resistance Using CRISPR/Cas9 Technology

“…Durability is not a specific aspect of resistance genes obtained by genome editing, and the answers are the same as for introgressed resistance genes discovered in the genetic variability of the species: (i) the stacking of several resistance genes, preferably with different modes of action, (ii) a focus on systems other than NBS-LRR receptor kinases known to break down rapidly, and (iii) good agronomic practices, including, in particular, crop rotation and the concomitant use of biocontrol agents. An example of two independent CRISPR/Cas9-derived resistances against the same disease are the knockouts of TaMLO (Wang et al, 2014) and TaEDR1 (Zhang et al, 2017), both conferring resistance to powdery mildew in wheat. Beyond the creation of novel alleles conferring protection, CRISPR/Cas9 technology can also be helpful in the stacking process itself.…”

“…Continued efforts to improve its efficiency, for example by the use of lig4 (Endo et al, 2016) or polQ mutations (Saito et al, 2017), or a copy number increase of the repair matrix by virus vectors (Čermák et al, 2015), are crucial to increasing the range of tools available to plant pathologists. Base editing, to date permitting C to T and A to G transitions in plants, is more limited in scope but has recently emerged as a readily available alternative for certain editing projects (Zhang et al, 2017; Hua et al, 2018). …”

A roadmap for gene functional characterisation in wheat

“…Therefore, to characterise the function of a gene in wheat it is often necessary to knock out all three homoeologs. This may be achieved by simultaneously targeting all three copies using either RNAi (Uauy et al, 2006) or CRISPR (Zhang et al, 2017b). A large number of transformants need to be screened to identify a null in all three genomes from a CRISPR construct (Zhang et al, 2017b).…”

“…This may be achieved by simultaneously targeting all three copies using either RNAi (Uauy et al, 2006) or CRISPR (Zhang et al, 2017b). A large number of transformants need to be screened to identify a null in all three genomes from a CRISPR construct (Zhang et al, 2017b). If the targets are more divergent it may not even be possible to use one guide RNA to target all three homoeologs, in which case several guide RNAs may be used through multiplexing.…”