Fine Mapping Of Wheat Stripe Rust Resistance Gene Yr26

Wheat stripe rust, caused by the fungal pathogen Puccinia striiformis Westend. f. sp.tritici Erikss.（Pst）, is an epidemic and destructive disease to wheat production all over theworld. Growing resistant cultivars is the most effective method to control the disease. TheYr26gene, conferring resistance to all currently important races of Puccinia striiformis f. sp.tritici （Pst） in China, was still effective in controlling the wheat stripe rust. Thus, Cloning ofYr26gene is important for both understanding the molecular mechanisms of resistance andhelpful for marker assisted selection （MAS） in wheat breeding.The Yr26gene was previously mapped to wheat chromosome deletion bin C-1BL-6-0.32with low-density markers. In this study, genomes of Brachypodium distachyon and rice wereused to develop markers to saturate the chromosomal region containing the Yr26locus, whichprovide a sound basis for further fine mapping and map-based cloning.In this study, F2and F_2:3progenies derived from Avocet S×92R137were used for finemapping of Yr26. Wheat expressed sequence tags （ESTs） located in deletion binC-1BL-6-0.32were used to develop sequence tagged site （STS） markers. The EST-STSmarkers flanking Yr26were used to identify collinear regions of the rice and B. distachyongenomes. Wheat ESTs with significant similarities in the two collinear regions were selectedto develop conserved markers for fine mapping of Yr26. The details are as follows:1. To determine inheritance of the Yr26gene in wheat variety92R137, a total of6,700F2plants （36lines） and551F_2:3progenies were inoculated with Chinese PST race CYR32inthe greenhouse. Among them,2,341plants （15lines） were initially used for fine mapping ofYr26gene. The F2population segregated in1,747resistant and594susceptible, fitting a_3:1ratio (χ²_3:1=0.17, P=0.68), indicating that Yr26in the AvS x92R137population behaved asa single dominant gene. The segregation of551F_2:3families these families conformed to a1:2:1ratio (χ²_1:2:1=1.36, P=0.51) as expected for a single gene. Simultaneously, the other4,359F2plants provide a sufficient basis for further selection of recombinants and markerdevelopment.2. Previous selected SSR markers linked with Yr26were firstly used to test forpolymorphism in present population. Only five SSR markers （Xgwm11, Xgwm18, Xgwm413,Xbarc181and Xwmc419） showed clear polymorphisms between the parents and bulks. Of the62SSR markers physically mapped on chromosome1B, four markers （Xgwm33, Xgwm273, Xbarc61and Xwmc230） were validated to be linked with Yr26with a F2population of196plants. The four SSR markers were linked with Yr26gene with genetic distances of11.8cM、9.7cM、6.3cM and3.1cM, respectively, and the two closet flanking loci were Xbarc61andXwmc230. Yr26gene was previously assigned to wheat chromosome deletion binC-1BL-6-0.32with six EST-STS markers （WE201, WE202, WE210, WE171, WE173andWE177）, these markers were firstly used to test for polymorphisms between the presentparents and bulks. In addition to those markers,163new pairs of EST-STS primers weredesigned from wheat ESTs mapped in the Yr26region and used in the bulked segregantanalysis. The results showed that two previous markers （WE201and WE173） and eight newlydeveloped markers （STS-BQ5, STS-BQ6, STS-CD28, STS-BQ33, STS-BE46, STS-BE68,STS-BQ74and STS-CD77） produced stable polymorphic bands in the bulk segregant analysis.All ten EST-STS markers were used to genotype the entire F2population of2,341plants. Theten EST-STS markers were mapped within a genetic interval of4.02cM region carrying Yr26.Two EST-STS markers, STS-CD28and STS-BQ74, flanked the Yr26locus at genetic distancesof0.48and0.44cM.3. To accurately characterize the collinearity between the Yr26region and the genomicregions of B. distachyon and rice, ten sequences corresponding to the mapped wheat ESTswere used as queries to perform a BLAST search against the rice and B. distachyon genomesequences. Comparative genomic analysis established the collinearity of the Yr26genomicregion with a4.48Mb region （Bradi3g28070-Bradi3g31630） in B. distachyon chromosome3and a3.33Mb region （Os10g0462900-Os10g0524500） in rice chromosome10. To developmore markers for saturation of Yr26region, a comparative genomics approach were used.Genes located in the collinear regions of rice and B. distachyon were firstly analyzed to searchfor resistance gene analogs （RGAs）. cDNA sequences of genes matching RGAs were selectedto develop RGA-based markers; then genes in collinear regions of Brachypodium and ricewere also used to search against all wheat ESTs and develop conserved markers. All thenewly developed markers were used to determine polymorphisms between the parents andbulks and21conserved markers were found to be polymorphic. Six markers （CON-6, CON-7,CON-8, CON-9, CON-10and CON-11） among them, cosegregated with Yr26. The newlydeveloped markers CON-4and CON-12narrowed the genomic region carrying Yr26to a1.92Mb （Bradi3g28410-Bradi3g29600） on B. distachyon chromosome3and1.17Mb（Os10g0470700-Os10g0489800） on rice chromosome10. There are135and68genes in thenarrowed collinear regions of B. distachyon and rice, respectively. No typical NBS-LRRresistance gene analog was found in the collinear regions of rice （Os10g0470700-Os10g0489800） and B. distachyon （Bradi3g28410-Bradi3g29600）. However, Bradi3g28590 was annotated as “leucine-rich repeat （LRR） protein kinase”, and Bradi3g29120wasannotated as “protein kinase”.4. The31markers, including25closely-linked markers and6cosegregated markers,were used to test wheat cultivars/lines and to assess their potential in marker-assisted selectionfor Yr26. The results indicated that11markers （STS-BQ33, STS-CD77, STS-BQ74, WE173,CON-1, CON-3, CON-4, CON-5, CON-6, CON-10and CON-19） could be useful in selectionof Yr26（Yr24, YrCH42） in breeding programs.