Map-based cloning of the novel stripe rust resistance gene YrG303 and its use to engineer 1B chromosome with multiple beneficial traits
Authors: Fahima Tzion , Dubcovsky Jorge 
Published: January 2022

Research problem: Bread wheat (Triticumaestivum) provides approximately 20% of the calories and proteins consumed by humankind. As the world population continues to increase, it is necessary to improve wheat yields, increase grain quality, and minimize the losses produced by biotic and abiotic stresses. Stripe rust, caused by Pucciniastriiformisf. sp. tritici(Pst), is one of the most destructive diseases of wheat. The new pathogen races are more virulent and aggressive than previous ones and have produced large economic losses. A rich source for stripe-rust resistance genes (Yr) was found in wild emmer wheat populations from Israel. Original Project goals: Our long term goal is to identify, map, clone, characterize and deploy in breeding, novel wild emmer Yr genes, and combine them with multiple beneficial traits. The current study was aiming to map and clone YrG303 and Yr15, located on chromosome 1BS and combine them with drought resistance and grain quality genes. Positional cloning of YrG303/Yr15: Fine mapping of these genes revealed that YrG303 is actually allelic to Yr15. Fine genetic mapping using large segregating populations resulted in reduction of the genetic interval spanning Yr15 to less than 0.1 cM. Physical mapping of the YrG303/Yr15 locus was based on the complete chromosome 1BS physical map of wheat constructed by our group. Screening of 1BS BAC library with Yr15 markers revealed a long BAC scaffold covering the target region. The screening of T. dicoccoidesaccession-specific BAC library with Yr15 markers resulted in direct landing on the target site. Sequencing of T. dicoccoidesBAC clones that cover the YrG303/Yr15 locus revealed a single candidate gene (CG) with conserved domains that may indicate a role in disease resistance response. Validation of the CG was carried out using EMS mutagenesis (loss-of- function approach). Sequencing of the CG in susceptible yr15/yrG303 plants revealed three independent mutants that harbour non-functional yr15/yrG303 alleles within the CG conserved domains, and therefore validated its function as a Pstresistance gene. Evaluation of marker-assisted-selection (MAS) for Yr15. Introgressions of Yr15 into cultivated wheat are widely used now. Recently, we have shown that DNA markers linked to Yr15 can be used as efficient tools for introgression of Yr15 into cultivated wheat via MAS. The developed markers were consistent and polymorphic in all 34 tested introgressions and are the most recommended markers for the introgression of Yr15. These markers will facilitate simultaneous selection for multiple Yr genes and help to avoid escapees during the selection process. Engineering of improved chromosome 1BS that harbors multiple beneficial traits. We have implemented the knowledge and genetic resources accumulated in this project for the engineering of 1B "super-chromosome" that harbors multiple beneficial traits. We completed the generation of a chromosome including the rye 1RS distal segment associated with improved drought tolerance with the Yr gene, Yr15, and the strong gluten allele 7Bx-over-expressor (7Bxᴼᴱ). We have completed the introgression of this improved chromosome into our recently released variety Patwin-515HP and our rain fed variety Kern, as well as to our top breeding lines UC1767 and UC1745. Elucidating the mechanism of resistance exhibited by Yr36 (WKS1). The WHEAT KINASE START1 (WKS1) resistance gene (Yr36) confers partial resistance to Pst. We have shown that wheat plants transformed with WKS1 transcript are resistant to Pst. WKS1 is targeted to the chloroplast where it phosphorylates the thylakoid-associatedascorbateperoxidase (tAPX) and reduces its ability to detoxify peroxides. Based on these results, we propose that the phosphorylation of tAPX by WKS1 reduces the ability of the cells to detoxify ROS and contributes to cell death. Distribution and diversity of WKS in wild emmer populations. We have shown that WKS1 is present only in the southern distribution range of wild emmer in the Fertile Crescent. Sequence analysis revealed a high level of WKS1 conservation among wild emmer populations, in contrast to the high level of diversity observed in NB-LRR genes. This phenomenon shed some light on the evolution of genes that confer partial resistance to Pst. Three new WKS1 haplotypes displayed a resistance response, suggesting that they can be useful to improve wheat resistance to Pst. In summary, we have improved our understanding of cereals’ resistance mechanisms to rusts and we have used that knowledge to develop improved wheat varieties.