Exploration And Cloning Of Wheat Powdery Mildew Resistance Genes

Wheat(Triticum aestivum L.)is one of the most important food crops of the world.Powdery mildew caused by Blumeria graminis f.sp.tritici(Bgt)is a globally devastating foliar disease of wheat.Deployment of host resistance(R)genes is the most effective and environmentally safe approach to control this disease in agricultural production.However,R genes with large effect are often defeated by rapidly evolved pathogen populations.It is essential to constantly discover and utilize new powdery mildew R genes in the breeding program.Moreover,cloning of the R genes is beneficial for understanding disease resistance mechanism and performing molecular breeding.To explore the powdery mildew resistance gene in wild emmer wheat,in this study,we identified and mapped the powdery mildew resistance gene in wild emmer wheat C-21.C-21 was immune to Bgt isolate Bgt19.Genetic analysis revealed a single dominant gene in C-21 confering its resistance to Bgt19.This gene was mapped on chromosome bin 2BS-0.84-1.00 through marker analysis.More markers were developed using wheat EST(expressed sequence tags),Chinese spring genome contig sequences and macro-collinearity of gene region with Brachypodium distachyon chromosome 5(Bd5).Eventually,MIC21 was restricted to a 2.0 cM interval.Mbn2033 and Mlm80 from einkorn wheat TA2033 and M80 respectively were two dominant powdery mildew resistance genes isolated by our lab.Both of them belonged to Pml cluster that located on the long arm of chromosome 7A.Comparative genomics study showed that macro-colinearity exists between Bdl and the Mlm2033-Mlm80-Pmla region,which allowed us to develop markers based on the wheat sequences orthologous to genes contained in the Bdl region.With these and other newly developed and published markers,high-resolution maps were constructed for both Mlm2033 and Mlm80 using large F2 populations.Based on the closely linked common markers,Mlm2033,Mlm80,and MlIW172,another powdery mildew resistance gene in the Pml cluster,were not allelic to one another.Moreover,physical map of Mlm2033 was constructed through chromosome walking with TA2033 bacterial artificial chromosome(BAC)clones.Finally,Mlm2033 was delimited to a 722-kb contig by BAC clones sequencing.Severe recombination suppression was noted in the Mlm2033 candidate region,which contained a gene cluster comprised of 19 nucleotide-binding site and leucine-rich repeats(NBS-LRR).Therein,13 highly similar NBS-LRR located in a 516-kb region with multiple tandem duplications.Comparative genomic analysis revealed that this region had undergone rapid evolution and multiple R genes insertion events,leading to disruption of micro-colinearity in this region.Haplotype analysis indicated high variability of Mlm2033 locus in the einkorn wheat gene pool.Through expression analysis and virus-induced gene silencing experiments,we confirmed that an NBS-LRR type gene,Genel8,corresponds to the powdery mildew resistance conferred by Mlm12033.Haplotype analysis of Gene18 revealed that it is highly conserved in different einkorn wheat backgrounds.The physical map of this gene provides useful information for map-based cloning of the R genes at the Pml cluster and interpretation of their evolution.