Powdery mildew resistance gene Pm4b, originating from Triticum persicum, is effective against the prevalent Blumeria graminis f. sp. tritici (Bgt) isolates from certain regions of wheat production in China. The lack of tightly linked molecular markers with the target gene prevents the precise identification of Pm4b during the application of molecular marker-assisted selection (MAS). The strategy that combines the RNA-Seq technique and the bulked segregant analysis (BSR-Seq) was applied in an F2:3 mapping population (237 families) derived from a pair of isogenic lines VPM1/7∗Bainong 3217 F4 (carrying Pm4b) and Bainong 3217 to develop more closely linked molecular markers. RNA-Seq analysis of the two phenotypically contrasting RNA bulks prepared from the representative F2:3 families generated 20,745,939 and 25,867,480 high-quality read pairs, and 82.8 and 80.2% of them were uniquely mapped to the wheat whole genome draft assembly for the resistant and susceptible RNA bulks, respectively. Variant calling identified 283,866 raw single nucleotide polymorphisms (SNPs) and InDels between the two bulks. The SNPs that were closely associated with the powdery mildew resistance were concentrated on chromosome 2AL. Among the 84 variants that were potentially associated with the disease resistance trait, 46 variants were enriched in an about 25 Mb region at the distal end of chromosome arm 2AL. Four Pm4b-linked SNP markers were developed from these variants. Based on the sequences of Chinese Spring where these polymorphic SNPs were located, 98 SSR primer pairs were designed to develop distal markers flanking the Pm4b gene. Three SSR markers, Xics13, Xics43, and Xics76, were incorporated in the new genetic linkage map, which located Pm4b in a 3.0 cM genetic interval spanning a 6.7 Mb physical genomic region. This region had a collinear relationship with Brachypodium distachyon chromosome 5, rice chromosome 4, and sorghum chromosome 6. Seven genes associated with disease resistance were predicted in this collinear genomic region, which included C2 domain protein, peroxidase activity protein, protein kinases of PKc_like super family, Mlo family protein, and catalytic domain of the serine/threonine kinases (STKc_IRAK like super family). The markers developed in the present study facilitate identification of Pm4b during its MAS practice.

Wheat powdery mildew is caused by Blumeria graminis f. sp. tritici (Bgt), a biotrophic fungal species. It is very important to mine new powdery mildew (Pm) resistance genes for developing resistant wheat cultivars to reduce the deleterious effects of the disease. This study was carried out to characterize the Pm gene in Qingxinmai, a winter wheat landrace from Xinjiang, China. Qingxinmai is resistant to many Bgt isolates collected from different wheat fields in China. F1, F2, and F2:3 generations of the cross between Qingxinmai and powdery mildew susceptible line 041133 were developed. It was confirmed that a single recessive gene, PmQ, conferred the seedling resistance to a Bgt isolate in Qingxinmai. Bulked segregant analysis-RNA-Seq (BSR-Seq) was performed on the bulked homozygous resistant and susceptible F2:3 families, which detected 57 single nucleotide polymorphism (SNP) variants that were enriched in a 40 Mb genomic interval on chromosome arm 2BL. Based on the flanking sequences of the candidate SNPs extracted from the Chinese Spring reference genome, 485 simple sequence repeat (SSR) markers were designed. Six polymorphic SSR markers, together with nine markers that were anchored on chromosome arm 2BL, were used to construct a genetic linkage map for PmQ. This gene was placed in a 1.4 cM genetic interval between markers Xicsq405 and WGGBH913 corresponding to 4.9 Mb physical region in the Chinese Spring reference genome. PmQ differed from most of the other Pm genes identified on chromosome arm 2BL based on its position and/or origin. However, this gene and Pm63 from an Iranian common wheat landrace were located in a similar genomic region, so they may be allelic.

A single recessive powdery mildew resistance gene Pm61 from wheat landrace Xuxusanyuehuang was mapped within a 0.46-cM genetic interval spanning a 1.3-Mb interval of the genomic region of chromosome arm 4AL. Epidemics of powdery mildew incited by the biotrophic fungus Blumeria graminis f. sp. tritici (Bgt) have caused significant yield reductions in many wheat (Triticum aestivum)-producing regions. Identification of powdery mildew resistance genes is required for sustainable improvement of wheat for disease resistance. Chinese wheat landrace Xuxusanyuehuang was resistant to several Bgt isolates at the seedling stage. Genetic analysis based on the inoculation of Bgt isolate E09 on the F, F, and F populations produced by crossing Xuxusanyuehuang to susceptible cultivar Mingxian 169 revealed that the resistance of Xuxusanyuehuang was controlled by a single recessive gene. Bulked segregant analysis and simple sequence repeat (SSR) mapping placed the gene on chromosome bin 4AL-4-0.80-1.00. Comparative genomics analysis was performed to detect the collinear genomic regions of Brachypodium distachyon, rice, sorghum, Aegilops tauschii, T. urartu, and T. turgidum ssp. dicoccoides. Based on the use of 454 contig sequences and the International Wheat Genome Sequence Consortium survey sequence of Chinese Spring wheat, four EST-SSR and seven SSR markers were linked to the gene. An F recombinant inbred line population derived from Xuxusanyuehuang × Mingxian 169 cross was used to develop the genetic linkage map. The gene was localized in a 0.46-cM genetic interval between Xgwm160 and Xicsx79 corresponding to 1.3-Mb interval of the genomic region in wheat genome. This is a new locus for powdery mildew resistance on chromosome arm 4AL and is designated Pm61.