ldenti fication of the resistance gene to powdery mildew in Chinese wheat landrace Baiyouyantiao

XU Xiao-dan, FENG Jing FAN Jie-ru LlU Zhi-yong, Ll Qiang, ZHOU Yi-lin MA Zhan-hong

1 State Key Laboratory for Biology of Plant Diseases and Insect Pests/Institute of Plant Protection, Chinese Academy of Agricultural Sciences, Beijing 100193, P.R.China

2 College of Plant Protection, China Agricultural University, Beijing 100193, P.R.China

3 State Key Laboratory of Plant Cell and Chromosome Engineering/Institute of Genetics and Developmental Biology, Chinese Academy of Sciences, Beijing 100101, P.R.China

4 College of Plant Protection, Northwest A&F University/State Key Laboratory of Crop Stress Biology for Arid Areas, Yangling 712100, P.R.China

RESEARCH ARTICLE

ldenti fication of the resistance gene to powdery mildew in Chinese wheat landrace Baiyouyantiao

XU Xiao-dan1,2, FENG Jing1, FAN Jie-ru1, LlU Zhi-yong3, Ll Qiang4, ZHOU Yi-lin1, MA Zhan-hong2

1 State Key Laboratory for Biology of Plant Diseases and Insect Pests/Institute of Plant Protection, Chinese Academy of Agricultural Sciences, Beijing 100193, P.R.China

2 College of Plant Protection, China Agricultural University, Beijing 100193, P.R.China

3 State Key Laboratory of Plant Cell and Chromosome Engineering/Institute of Genetics and Developmental Biology, Chinese Academy of Sciences, Beijing 100101, P.R.China

4 College of Plant Protection, Northwest A&F University/State Key Laboratory of Crop Stress Biology for Arid Areas, Yangling 712100, P.R.China

Powdery mildew, caused by Blumeria graminis f. sp. tritici (Bgt), is one of the most damaging diseases to wheat in the world.The cultivation of resistant varieties of wheat is essential for controlling the powdery mildew epidemic. Wheat landraces are important resources of resistance to many diseases. Mapping powdery mildew resistance genes from wheat landraces will promote the development of new varieties with disease resistance. The Chinese wheat landrace Baiyouyantiao possesses characteristic of disease resistance to powdery mildew. To identify the resistance gene in this landrace, Baiyouyantiao was crossed with the susceptible cultivar Jingshuang 16 and seedlings of parents and F1, BC1, F2, and F2:3were tested with Bgt isolate E09. The genetic results showed that the resistance of Baiyouyantiao to E09 was controlled by a single recessive gene, tentatively designated PmBYYT. An Illumina wheat 90K single-nucleotide polymorphism (SNP) array was applied to screen polymorphisms between F2-resistant and F2-susceptible DNA bulks for identifying the chromosomal location of PmBYYT. A high percentage of polymorphic SNPs between the resistant and susceptible DNA bulks was found on chromosome 7B, indicating that PmBYYT may be located on this chromosome. A genetic linkage map of PmBYYT consisting of two simple sequence repeat markers and eight SNP markers was developed. The two flanking markers were SNP markers W7BL-8 and W7BL-15, with genetic distances of 3 and 2.9 cM, respectively. The results of this study demonstrated the rapid characterization of a wheat disease resistance gene and SNP marker development using the 90K SNP assay. The flanking markers of gene PmBYYT will bene fit marker-assisted selection (MAS) and map-based cloning in breeding wheat cultivars with powdery mildew resistance.

wheat landrace, powdery mildew, genetic mapping, single-nucleotide polymorphism (SNP) array

1. lntroduction

Powdery mildew, caused by Blumeria graminis f. sp. tritici(Bgt), is one of the most devastating diseases of wheat in the world. Growing resistant varieties can effectively control this disease and maintain the wheat yield. However, widely planting wheat varieties with Bgt race-speci fic resistance in a certain area often accelerates selection pressures to pathogens and causes a loss of resistance. Therefore,more resistance genes need to be identi fied for resource utilization. Wheat cropping in China has a long history and is practiced through the country. The vast territory, complicated topography, variable climate, and long period of evolution and arti ficial selection have contributed to the rich diversity of wheat landraces. China has more than 13 000 wheat landraces, which enrich genetic resources and play an important role in promoting wheat breeding (Liu et al. 2000).

More than 50 of ficially designated and many tentatively designated powdery mildew resistance (Pm) loci have been reported in wheat (Zhang et al. 2016). However, only three formally designated Pm loci, including Pm5 (Huang et al.2003), Pm24 (Huang and Röder 2011), and Pm47 (Xiao et al. 2013), and some tentatively designated Pm loci, such as PmTm4 (Hu et al. 2008), mlxbd (Xue et al. 2009a), and MlHLT (Wang et al. 2015), have been identi fied from wheat landraces. Conventional breeding requires many years to select high-yielding cultivars with sound disease resistance.The rapid progress of molecular biology technology has provides the marker-assisted selection (MAS) tool for wheat breeders, which can signi ficantly shorten the breeding period for wheat cultivars. Therefore, more studies are needed to identify new powdery mildew resistance genes from wheat landraces and develop closely linked molecular markers for MAS in wheat breeding programs.

Many types of molecular markers, such as RFLP (restriction fragment length polymorphism), AFLP (ampli fied fragment length polymorphism), and SSR (simple sequence repeats) markers, have been used for identifying and mapping wheat Pm genes (Hartl et al. 1995; Tao et al.2000; Miranda et al. 2007; Hao et al. 2008; Hua et al. 2009;Petersen et al. 2015). However, it is dif ficult to develop a high-density linkage map for a particular gene due to the huge wheat genome and the unavailability of reference genome sequences. Single-nucleotide polymorphism (SNP)refers to a single-nucleotide mutation in the genome. SNP markers are abundant in quantity and polymorphism. Combined with chip technology, the rapid, large-scale screening of SNPs is feasible. High-throughput wheat SNP genotyping platforms offer opportunities to study genetic diversity (Wang et al. 2014; Yu et al. 2014) and conduct association analysis(Zanke et al. 2014), and gene mapping (Dreisigacker et al.2015; Lu et al. 2015; Liu et al. 2016).

The Chinese wheat landrace Baiyouyantiao is resistant to powdery mildew (Sheng et al. 1992; Xue et al. 2009b).However, the application of Baiyouyantiao in wheat breeding programs is limited due to an unclear inheritance mechanism and unavailability of linked markers. The primary objectives of the present study were to study the genetics of resistance to powdery mildew in Baiyouyantiao and map the chromosomal location of the resistance gene using the 90K wheat SNP array, and develop a genetic linkage map of the resistance gene using SSR and SNP markers. Our results may be useful for further determining the identity of the resistance gene as well as in MAS.

2. Materials and methods

2.1. Plant materials and Bgt isolates

The wheat landrace Baiyouyantiao was collected from Shaanxi Province in China and provided by the Institute of Plant Protection, Chinese Academy of Agricultural Sciences. The 31 cultivars (lines) with known Pm gene(s)were evaluated for Pm gene postulation. Wheat cultivars Jingshuang 16 and Chancellor were used as the susceptible controls. Additionally, Jingshuang 16 was crossed with Baiyouyantiao to generate an F1and segregating BC1, F2and F2:3progenies for genetic analysis and gene mapping.This study located the powdery mildew resistance gene PmBYYT on chromosome arm 7BL; therefore, cultivars Fuzhuang 30, Hongyoumai, Xiaobaidongmai, and Tangmai 4,carrying Pm genes mapped on chromosome arm 7BL,were used to compare the reactions to 22 Bgt isolates with Baiyouyantiao. Because the reactions of Baiyouyantiao and Xiaobaidongmai to 22 Bgt isolates were most similar,an F2population of Baiyouyantiao×Xiaobaidongmai was evaluated for allelism.

The Bgt isolates used in this study, collected from different parts of China, were selected for evaluating resistance to powdery mildew.

2.2. Disease evaluation of powdery mildew

Baiyouyantiao, 31 cultivars (lines) with known Pm gene(s)and the susceptible controls Jingshuang 16 and Chancellor were planted in culture trays. Six to eight seeds of each cultivar (line) were grown in each well. Twenty-two sets were planted in total, and each set was covered with a see-through plastic bag for preventing cross-contamination among the Bgt isolates. The seedlings were placed in a greenhouse with temperature of 18°C during the day and 16°C at night. When the first leaves fully expanded,seedlings of all sets were inoculated separately with 22 Bgt isolates.

Baiyouyantiao, Jingshuang 16 and their F1, BC1and F2populations were planted in culture trays with one seed per well for evaluating the genetics of resistance to powdery mildew in the seedling stage ( five seeds were planted for the parental lines Baiyouyantiao and Jingshuang 16). The F2:3lines were tested to identify the phenotypes of corresponding F2plants. The seeds of each F2:3line were planted in two wells with eight plants each. After the first leaves were fully expanded, all the tested seedlings were inoculated with Bgt isolate E09. The inoculated seedlings were kept in an MLR-352H-PC Panasonic Incubator (Panasonic Corporation, China) with a temperature of 18°C under a 12-h light/dark cycle.

The infection types (ITs) were scored 10 d post-inoculation based on the 0, 0;, 1, 2, 3, and 4 scales when the susceptible controls Jingshuang 16 and Chancellor were fully developed with spores on the first leaves (Sheng 1988).Standards were de fined as follows: 0, without disease spot;0;, hypersensitive reactions with dying spot; 1, disease spot less than 1 mm with thin hypha layer and fewer spores; 2,disease spot less than 1 mm with thick hypha layer and more spores; 3, scattered disease spot greater than 1 mm with thick hypha layer and more spores; and 4, continuous disease spot greater than 1 mm with thick hypha layer and more spores. Plants with ITs 0–2 were classi fied as resistant,and those with ITs 3–4 were classi fied as susceptible. The χ2tests for goodness-of- fit were used to test for deviations of the observed and expected segregation ratios of the F2population (the disease responses were checked according to F2:3progenies) and F2:3progenies.

2.3. DNA isolation and wheat 90K SNP array

Genomic DNA was extracted from leaf tissues collected from the parents and F2plants using the CTAB (hexadecyltrimethy ammonium bromide) protocol (Rogers and Bendich 1985).Ten homozygous resistant and 10 homozygous susceptible F2plants were selected for generating the resistant and susceptible bulks, respectively. The resistant and susceptible DNA bulks were screened with the Illumina wheat 90K SNP array. Scan data were analyzed using GenomeStudio software. The polymorphic SNP markers were filtered between resistant and susceptible DNA bulks. The chromosome with the highest number of polymorphic SNPs was assumed to carry the resistance gene.

2.4. SSR and SNP markers for gene mapping

Bulked segregant analysis was used to identify markers associated with the powdery mildew resistance gene. SSR primers on wheat chromosome 7B were selected to first screen both parents and bulks. The primers’ sequences were obtained from the Graingenes 2.0 website (http://wheat.pw.usda.gov/GG2/). Then, the polymorphic SSR markers between the resistant bulk and the susceptible bulk as well as the resistant and susceptible parents were used to genotype the whole F2population. The polymerase chain reaction (PCR) system was performed as follows: 5 µL of 2× Pfu PCR Master Mix (Tiangen Biotech (Beijing) Co.,Ltd.), 2 µL of primers, 1 µL of genomic DNA (70 ng µL–1),and 2 µL of double distilled H2O; the total volume was 10 µL.The PCR ampli fications were carried out with the following steps: denaturation for 5 min at 94°C; denaturation for 1 min at 94°C, annealing for 50 s at 50–60°C (depending on the annealing temperature of each primer), extension for 2 min at 72°C, 35 cycles; and extention for 10 min at 72°C. PCR products were separated by 6% PAGE (polyacryl-amide gel electrophoresis).

The polymorphic SNP markers were identi fied using the resistant and susceptible DNA bulks, which were screened by the Illumina wheat 90K SNP array. The polymorphic SNP markers between the two bulks were blasted to the wheat reference genome (ftp://ftp.ensemblgenomes.org/pub/plants/release-28/fasta/triticum_aestivum/dna/).Polymorphic SNP markers from the possible chromosome were used to genotype each DNA of the F2seedlings.Genotyping was performed using the Sequenom iPLEX Gold Assay (Sequenom, Cambridge, MA). Locus-speci fic PCR primers and allele-speci fic detection primers were designed using MassARRAY Assay Design 3.1 Software.DNAs were ampli fied in a multiplex PCR and labeled using a locus-speci fic single-base extension reaction. The products were desalted and transferred to a 384-element SpectroCHIP array. Allele detection was performed using Matrix-Assisted Laser Desorption/Ionization Time-of-Flight Mass Spectrometry (compact MALDI-TOF MS). Mass spectrograms and clusters were analyzed by the TYPER 3.4 Software package. All DNA samples were deposited on a 384-well plate. Baiyouyantiao, Jingshuang 16, and the resistant and susceptible DNA bulks were repeated five times in 384-well plates as controls.

Linkage analysis was performed using the Software Join-Map 4.0. The centimorgan (cM) distance was estimated by Kosambi map function (Kosambi 1943). The genetic map was drawn with Mapdraw ver. 2.1 (Liu and Meng 2003).

3. Results

3.1. Gene postulation and genetic analysis of resistance to wheat powdery mildew in Baiyouyantiao

At the seedling stage, 22 Bgt isolates were used to inoculate Baiyouyantiao in addition to 31 cultivars (lines) with known Pm gene(s) and the susceptible controls Jingshuang 16 and Chancellor for evaluating the powdery mildew resistance and postulating the resistance gene(s). Jingshuang 16 and Chancellor were susceptible to all tested Bgt isolates with IT of 4 (Table 1). Baiyouyantiao was resistant to 20 Bgt isolates and susceptible to two isolates, E07 and E17,indicating broad-spectrum powdery mildew resistance in Baiyouyantiao. The reactions of Baiyouyantiao to 22 Bgt isolates were different from those of cultivars (lines) with known Pm gene(s) used in this study.

To determine the inheritance of powdery mildew resistance, Baiyouyantiao, Jingshuang 16, and their F1, BC1, F2,and F2-derived F3progenies were evaluated for responses to Bgt isolate E09.

Baiyouyantiao was resistant to Bgt isolate E09 with IT 0;, and Jingshuang 16 was susceptible to E09 with IT 4(Table 2, Fig. 1-A). All 14 F1plants and 17 BC1plants were susceptible to E09 with IT 4, indicating the recessive nature of this resistance. The 364 F2plants of cross Baiyouyantiao×Jingshuang 16 segregated as 103 resistant and 261 susceptible, which fit a 1:3 ratio (χ2=2.11, P=0.15). The F2-derived F3progenies included that were 103 homozygous resistant, 182 segregating, and 79 homozygous susceptible, which fit a 1:2:1 ratio (χ2=3.16, P=0.21). The 206 F2plants of the reciprocal cross Jingshuang 16×Baiyouyantiao segregated as 53 resistant and 153 susceptible, also fitting a 1:3 ratio (χ2=0.06, P=0.81). These results indicated that the resistance of wheat landrace Baiyouyantiao to Bgt isolate E09 was controlled by a single recessive gene, tentatively designated PmBYYT.

3.2. Wheat 90K SNP array of the resistant and susceptible DNA bulks

The resistant and susceptible DNA bulks, derived from F2plants of the cross Baiyouyantiao×Jingshuang 16 inoculated with Bgt isolate E09, were screened with the Illumina wheat 90K SNP array. A total of 341 polymorphic SNP markerswere identi fied between resistant and susceptible DNA bulks and distributed on all chromosomes except chromosome 5D(Fig. 1-B). The distribution of polymorphic SNP markers on each chromosome was analyzed. There were 114 (33.43%)located on chromosome 7B, followed by 41 (12.02%) and 36 (10.56%) polymorphic SNP markers on chromosomes 3B and 2B, respectively. All of the remaining 150 (43.99%)polymorphic SNP markers were located on the other 18 chromosomes. Thus, it was speculated that the resistance gene in Baiyouyantiao to Bgt isolate E09 may be located on chromosome 7B.

Table 1 Infection types of Baiyouyantiao and 31 cultivars (lines) with known Pm genes to 22 Blumeria graminis f. sp. tritici (Bgt)isolates

Table 2 Evaluation of resistance of the parents and progenies derived from the cross Baiyouyantiao×Jingshuang 16 to Blumeria graminis f. sp. tritici (Bgt) isolate E09

Fig. 1 Infection types of Baiyouyantiao and Jingshuang 16 to Blumeria graminis f. sp. tritici (Bgt) isolate E09 (A) and chromosomal distribution of polymorphic single-nucleotide polymorphism (SNP) markers between the resistant and susceptible DNA bulks (B).

3.3. Mapping PmBYYT using SSR and SNP markers

A total of 101 SSR markers on wheat chromosome 7B were selected to screen for polymorphisms between Baiyouyantiao and Jingshuang 16 as well as the resistant and susceptible bulks. Five SSR markers, Xgwm46, Xwmc526,Xwmc364, Xgwm333, and Xbarc32, produced polymorphic bands between the parental lines and the bulks. These markers were used to genotype the DNA of all F2plants derived from the cross Baiyouyantiao×Jingshuang 16.Subsequent linkage analysis indicated that only Xwmc526 and Xbarc32 were linked to the gene PmBYYT with genetic distances of 4.4 and 5.5 cM, respectively (Fig. 2-B).

To obtain more genetic markers linked to PmBYYT, the sequences of the 114 polymorphic SNPs located on 7B were obtained and aligned to wheat reference genome sequences.The 21 polymorphic SNP markers on chromosome arm 7BL were selected for PCR primer design and validation on all F2plants. Finally, eight polymorphic SNP markers were linked to PmBYYT (Table 3). The two closest flanking markers were W7BL-8 and W7BL-15 (Fig. 2-A and B), with genetic distances of 3 and 2.9 cM, respectively (Fig. 2-B).

3.4. Relationships of PmBYYT and the other four Pm genes on 7BL

The disease reactions of Baiyouyantiao, Fuzhuang 30,Hongyoumai, Xiaobaidongmai, and Tangmai 4 to 22 Bgt isolates were evaluated. These cultivars (lines) showed the same resistance reactions to 16 Bgt isolates. However, for the other six isolates, including E01, E05, E06, E17, E23-1,and E68, Baiyouyantiao showed different disease responses from Fuzhuang 30, Hongyoumai, Xiaobaidongmai, and Tangmai 4 (Table 4). Baiyouyantiao was resistant to all tested Bgt isolates except for E17, whereas Fuzhuang 30 was susceptible to E06 and E23-1 and resistant to E17.Also, Hongyoumai was susceptible to E01, E06, and E23-1,and Tangmai 4 was susceptible to E01, E05, and E06.However, Baiyouyantiao was resistant to these Bgt isolates.Because Baiyouyantiao showed different disease responses from Fuzhuang 30, Hongyoumai and Tangmai 4, PmBYYT may be different from Pm5e, Pmhym, and PmTm4.

Fig. 2 The F2 population genotyping results with the two closest linked single-nucleotide polymorphism (SNP) markers W7BL-8 and W7BL-15 (A) and linkage map for the powdery mildew resistance gene PmBYYT on chromosome arm 7BL (B). A, channel intensities are plotted and coloured by genotypes, where heterozygotes are green, resistant homozygotes and susceptible homozygotes are orange and blue. B, genetic distances in cM are shown on the left and locus names on the right.

In addition, Xiaobaidongmai was resistant to E17 and susceptible to E68, whereas Baiyouyantiao was susceptible to E17 and resistant to E68. To further determine the relationship between PmBYYT and mlxbd, 858 F2plants derived from the cross Baiyouyantiao×Xiaobaidongmai were inoculated with Bgt isolate E09. Baiyouyantiao and Xiaobaidongmai were resistant to E09 with IT 0;, all of the 858 F2plants were resistant to E09 with IT 0;, indicating that the resistance genes, PmBYYT and mlxbd, may be allelic or very closely linked.

4. Discussion

The Chinese wheat landrace Baiyouyantiao showed resistance to 20 out of 22 tested Bgt isolates, indicating broad-spectrum resistance against B. graminis f. sp. tritici.Therefore, Baiyouyantiao can be combined with other effective resistance genes to develop wheat cultivars with high-level resistance. Comparing resistance reactions of Baiyouyantiao and 31 cultivars (lines) with known Pm gene(s) revealed that the resistance reactions of Baiyouyantiao were different from those of comparative cultivars(lines), indicating that the powdery mildew resistance gene in Baiyouyantiao may be different from these known Pm genes. The genetic studies indicated that a single recessive gene, PmBYYT, in Baiyouyantiao confers resistance to Bgt isolate E09. It is very interesting that the recessive mode of action is a common feature of all known alleles at the Pm5 locus or genes nearby this locus, including Pm5a–Pm5e(Law and Wolfe 1966; Hsam et al. 2001; Huang et al. 2003),Pmhym (Wang et al. 2009), mlxbd (Xue et al. 2009a), and PmTm4 (Hu et al. 2008). The mechanisms by which all known alleles at the Pm5 locus or genes nearby this locus are recessive genes are unknown, and further research on this topic is necessary.

The combination of bulked segregant analysis and SNP arrays provides a rapid approach for mapping wheat genes on a particular chromosome. In this study, the powdery mildew resistance gene of Baiyouyantiao was found to be located on chromosome 7B using the Illumina wheat 90K SNP array. And yet, a few polymorphic SNP markers have been detected on other chromosomes due to the possibility of a marker enrichment section of the genome (Liao et al.2009). The large size of the wheat genome and the difference in the genetic background of parental cultivars also increase the rate of incorrect detection.

Polymorphic SNPs provide useful information to develop PCR-based markers and construct a high-density genetic linkage map of the target gene using the Sequenom iPLEX Gold Assay platform. However, a low ef ficiency of polymorphic SNP detection was observed when DNA bulks were screened with the 90K SNP assay. Only 114 out of 2 347 array probes on chromosome 7B detected polymorphisms between the resistant and susceptible DNA bulks.Finally, only eight SNP markers were found to be linked to PmBYYT.Signi ficant progresses have been made in sequencing wheat and the genomes of relatives with the development of high-throughput sequencing technologies (Brenchley et al.2012; Ling et al. 2013;Jia et al. 2013; IWGSC et al. 2014). Complying with these resources,another two higher density wheat SNP assays,including the 660K SNP array and 820K SNP array, were respectively developed by the Chinese Academy of Agricultural Sciences(CAAS) and Biotechnology and Biological Sciences Research Council (BBSRC)-funded Wheat Improvement Strategic Programme(WISP) in the UK. In the future, these new arrays may be able to improve the ef ficiency of polymorphic marker development.

According to two SSR markers and eight SNP markers, the gene PmBYYT was located on chromosome 7BL,flanked by SNP markers W7BL-8 and W7BL-15 with genetic distances of 3.0 and 2.9 cM, respectively. These markers will be bene ficial to the MAS-based breeding for powdery mildew tolerance in wheat. To date, 10 powdery mildew resistance genes have been located on chromosome 7B. Among them, Pm40 from wheat-Elytrigia intermedium introgression line GYR19 (Luo et al. 2009) and Pm47 from Chinese landrace Hongyanglazi (Xiao et al.2013) were located on chromosome arm 7BS. However,PmBYYT was located on chromosome arm 7BL. Therefore,PmBYYT should be different from Pm40 and Pm47. Among the other eight genes (including Pm5a–Pm5e, Pmhym,mlxbd, and PmTm4), Pm5e, Pmhym, mlxbd, and PmTm4 were derived from Chinese wheat landraces Fuzhuang 30,Hongyoumai, Xiaobaidongmai, and Tangmai 4, respectively.One SSR marker, Xwmc526, linked to PmBYYT with a genetic distance of 5.5 cM, was also linked to Pmhym with a genetic distance of 57.2 cM (Wang et al. 2009), suggesting that PmBYYT and Pmhym may be present at different loci.All the markers linked to PmBYYT were different from the markers linked to Pm5e, mlxbd, and PmTm4. Twenty-two Bgt isolates were used to classify the disease response pattern of Baiyouyantiao and Fuzhuang 30 (Pm5e),Hongyoumai (Pmhym), Xiaobaidongmai (mlxbd), and Tangmai 4 (PmTm4). The disease reactions of Fuzhuang 30,Hongyoumai, and Tangmai 4 were different from those of Baiyouyantiao on more than two Bgt isolates, which suggested that PmBYYT may be different from Pm5e, Pmhym, and PmTm4. In addition, an allelism test between Baiyouyantiao and Xiaobaidongmai was conducted because their disease responses were very similar. Additionally, no susceptible plants were detected in the F2population derived from Baiyouyantiao×Xiaobaidongmai, which indicated that the gene PmBYYT and mlxbd may be allelic or very closely linked.

According to previous reports (Huang et al. 2003; Hu et al. 2008; Xue et al. 2009b), Pm5e, PmTm4, and mlxbd were located on or nearby the Pm5 locus. Moreover, these genes were all derived from Chinese wheat cultivars or landraces. Using allelism tests, Huang et al. (2000) reported that Pm5e and mlxbd were all closely linked to Pm5a but not the allelic genes of Pm5, whereas mlxbd and Pm5e may be allelic. According to the allelism test of this study, PmBYYT and mlxbd may be allelic or linked gene; therefore, PmBYYT and Pm5e may also be allelic or linked. Additional studies are needed to further determine the allelic relationships among PmBYYT, PmTm4, and other Pm5 locus genes.

5. Conclusion

In the present study, wheat landrace Baiyouyantiao was highly resistant to powdery mildew, and the resistance to Bgt isolate E09 was controlled by a single recessive gene,PmBYYT. PmBYYT was mapped to chromosome arm 7BL and closely flanked by two SNP markers, W7BL-11 and W7BL-14. A comparison of the disease reactions of Pm genes on chromosome 7B and an allelic analysis of genes PmBYYT and mlxbd showed that PmBYYT may be allelicor linked with mlxbd and Pm5e. The gene PmBYYT and closely linked SNP markers will be of bene fit for breeders to combine this locus with valuable genes and develop new cultivars (lines) resistant to powdery mildew.

Table 4 Different reactions of Baiyouyantiao, Fuzhuang 30, Hongyoumai, Xiaobaidongmai, Tangmai 4 and the susceptible controls Jingshuang 16 and Chancellor to 22 Blumeria graminis f. sp. tritici (Bgt) isolates

Acknowledgements

The study was funded by the National Key Research and Development Program of China (2017YFD0201701), the Special Fund for Agro-scienti fic Research in the Public Interest, China (201303016) and the Science and Technology Project for Xingjiang Uygur Autonomous Region, China(2013911092).

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28 December, 2016 Accepted 1 March, 2017

XU Xiao-dan, Mobile: +86-13141384399, E-mail: xxdalice@163.com; Correspondence ZHOU Yi-lin, Tel: +86-10-62815946,E-mail: yilinzhou6@aliyun.com; MA Zhan-hong, E-mail: mazh@cau.edu.cn

© 2018 CAAS. Publishing services by Elsevier B.V. All rights reserved.

10.1016/S2095-3119(16)61610-6

Section editor ZHANG Xue-yong

Managing editor WANG Ning