Fine genetic mapping of <Emphasis Type="Italic">xa24</Emphasis>, a recessive gene for resistance against <Emphasis Type="Italic">Xanthomonas oryzae</Emphasis> pv. <Emphasis Type="Italic">oryzae</Emphasis> in rice

Bacterial blight, caused by Xanthomonas oryzae pv. oryzae (Xoo), is the most devastating plant bacterial disease worldwide. Different bacterial blight resistance (R) genes confer race-specific resistance to different strains of Xoo. We fine mapped a fully recessive gene, xa24, for bacterial blight resistance to a 71-kb DNA fragment in the long arm of rice chromosome 2 using polymerase chain reaction-based molecular markers. The xa24 gene confers disease resistance at the seedling and adult stages. It mediates resistance to at least the Philippine Xoo races 4, 6 and 10 and Chinese Xoo strains Zhe173, JL691 and KS-1-21. Sequence analysis of the DNA fragment harboring the dominant (susceptible) allele of xa24 suggests that this gene should encode a novel protein that is not homologous to any known R proteins. These results will greatly facilitate the isolation and characterization of xa24. The markers will be convenient tools for marker-assisted selection of xa24 in breeding programs. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Identification of novel recessive gene <Emphasis Type="Italic">xa44</Emphasis>(<Emphasis Type="Italic">t</Emphasis>) conferring resistance to bacterial blight races in rice by QTL linkage analysis using an SNP chip

Key message

Using QTL analysis and fine mapping, the novel recessive gene xa44(t) conferring resistance to BB was identified and the expression level of the gene was confirmed through qRT-PCR analysis.

Abstract

Bacterial blight (BB) disease caused by Xanthomonas oryzae pv. oryzae (Xoo) is a major factor causing rice yield loss in most rice-cultivating countries, especially in Asia. The deployment of cultivars with resistance to BB is the most effective method to control the disease. However, the evolution of new Xoo or pathotypes altered by single-gene-dependent mutations often results in breakdown of resistance. Thus, efforts to identify novel R-genes with sustainable BB resistance are urgently needed. In this study, we identified three quantitative trait loci (QTLs) on chromosomes 1, 4, and 11, from an F₂ population of 493 individuals derived from a cross between IR73571-3B-11-3-K3 and Ilpum using a 7K SNP chip. Of these QTLs, one major QTL, qBB_11, on chromosome 11 explained 61.58% of the total phenotypic variance in the population, with an LOD value of 113.59, based on SNPs 11964077 and 11985463. The single major R-gene, with recessive gene action, was designated xa44(t) and was narrowed down to a 120-kb segment flanked within 28.00 Mbp to 28.12 Mbp. Of nine ORFs present in the target region, two ORFs revealed significantly different expression levels of the candidate genes. These candidate genes (Os11g0690066 and Os11g0690466) are described as “serine/threonine protein kinase domain containing protein” and “hypothetical protein,” respectively. The results will be useful to further understand BB resistance mechanisms and provide new sources of resistance, together with DNA markers for MAS breeding to improve BB resistance in rice.

     

Development of SCAR markers linked to <Emphasis Type="Italic">sin</Emphasis>-<Emphasis Type="Italic">2</Emphasis>, the stringless pod trait in pea (<Emphasis Type="Italic">Pisum sativum</Emphasis> L.)

With increasing consumer demand for vegetables, edible-podded peas have become more popular. Stringlessness is one of most important traits for snap peas. A single recessive gene, sin-2, controls this trait. Because pollen carrying the stringless gene is less competitive than pollen carrying the stringy gene, there are fewer than expected stringless plants recovered in segregating generations. Marker-assisted selection (MAS) is a valuable tool to identify plants with the traits of interest at an early stage in the breeding process. The objective of this study was to identify robust, user-friendly molecular markers tightly linked to sin-2. A total of 144 target region amplification polymorphism (TRAP) primer combinations were used to screen four DNA bulks, which were constructed from 32 pea breeding lines based on their phenotypes. Sixty polymorphic TRAP primer combinations were identified between bulks of stringless and stringy pods. Five primer combinations, F6_Trap03_168, F6_SA12_145, F10_ODD8_130, F11_GA5_850, and F12_SA12_190, showed more than 90 % association with the stringless phenotype in 32 pea breeding lines. Two of the TRAP markers, F10_ODD8_130 and F12_SA12_190, were cloned, sequenced, and successfully converted to sequence characterized amplified region (SCAR) markers. These two SCAR markers were validated using 20 F₅ recombinant inbred lines derived from a cross between Bohatyr (a dry pea variety with strings) and S1188 (a stringless snap pea variety) and showed strong marker-trait association. The results will have direct application in MAS of stringless edible-podded peas.     

Molecular tagging and validation of microsatellite markers linked to the low germination stimulant gene (<Emphasis Type="Italic">lgs</Emphasis>) for <Emphasis Type="Italic">Striga</Emphasis> resistance in sorghum [<Emphasis Type="Italic">Sorghum bicolor</Emphasis> (L.) Moench]

Striga is a devastating parasitic weed in Africa and parts of Asia. Low Striga germination stimulant activity, a well-known resistance mechanism in sorghum, is controlled by a single recessive gene (lgs). Molecular markers linked to the lgs gene can accelerate development of Striga-resistant cultivars. Using a high density linkage map constructed with 367 markers (DArT and SSRs) and an in vitro assay for germination stimulant activity towards Striga asiatica in 354 recombinant inbred lines derived from SRN39 (low stimulant) × Shanqui Red (high stimulant), we precisely tagged and mapped the lgs gene on SBI-05 between two tightly linked microsatellite markers SB3344 and SB3352 at a distance of 0.5 and 1.5 cM, respectively. The fine-mapped lgs region was delimited to a 5.8 cM interval with the closest three markers SB3344, SB3346 and SB3343 positioned at 0.5, 0.7 and 0.9 cM, respectively. We validated tightly linked markers in a set of 23 diverse sorghum accessions, most of which were known to be Striga resistant, by genotyping and phenotyping for germination stimulant activity towards both S. asiatica and S. hermonthica. The markers co-segregated with Striga germination stimulant activity in 21 of the 23 tested lines. The lgs locus similarly affected germination stimulant activity for both Striga species. The identified markers would be useful in marker-assisted selection for introgressing this trait into susceptible sorghum cultivars. Examination of the sorghum genome sequence and comparative analysis with the rice genome suggests some candidate genes in the fine-mapped region (400 kb) that may affect strigolactone biosynthesis or exudation. This work should form a foundation for map-based cloning of the lgs gene and aid in elucidation of an exact mechanism for resistance based on low Striga germination stimulant activity.     

Fine mapping of <Emphasis Type="Italic">qSTV11</Emphasis><Superscript><Emphasis Type="Italic">TQ</Emphasis></Superscript>, a major gene conferring resistance to rice stripe disease

The indica rice cultivar, Teqing, shows a high level of resistance to rice stripe virus (RSV). It is believed that this resistance is controlled by the gene, qSTV11 ^TQ . For positional cloning of the resistance gene, a set of chromosome single segment substitution lines (CSSSLs) was constructed, all of which had the genetic background of the susceptible japonica cultivar, Lemont, with different single substituted segments of Teqing on chromosome 11. By identifying the resistance of the CSSSLs-2006 in a field within a heavily diseased area, the resistance gene qSTV11 ^TQ was mapped between the markers Indel7 and RM229. Furthermore, in that region, six new markers were developed and 52 subregion CSSSLs (CSSSLs-2007) were constructed. The natural infection experiment was conducted again at different sites, with two replicates used in each site in order to identify the resistance phenotypes of the CSSSLs-2007 and resistant/susceptible controls in 2007. Through the results of 2007, qSTV11 ^TQ was localized in a region defined by the markers, CAPs1 and Indel4. In order to further confirm the position of qSTV11 ^TQ , another set of subregion CSSSLs (CSSSLs-2009) was constructed. Finally, qSTV11 ^TQ was localized to a 55.7 kb region containing nine annotated genes according to the genome sequence of japonica Nipponbare. The relationship between qSTV11 ^TQ and Stvb-i (Hayano-Saito et al. in Theor Appl Genet 101:59–63, 2000) and the reliability of the markers used on both sides of qSTV11 ^TQ for marker-assisted breeding of resistance to rice stripe disease are discussed.     

Identification and characterization of a major QTL responsible for erect panicle trait in <Emphasis Type="Italic">japonica</Emphasis> rice (<Emphasis Type="Italic">Oryza sativa</Emphasis> L.)

Panicle erectness (PE) is one of the most important traits for high-yielding japonica cultivars. Although several cultivars with PE trait have been developed and released for commercial production in China, there is little information on the inheritance of PE traits in rice. In the present study, 69 widely cultivated japonica cultivars and a double haploid (DH) population derived from a cross between a PE cultivar (Wuyunjing 8) and a drooping panicle cultivar (Nongken 57) were utilized to elucidate the mechanisms of PE formation and to map PE associated genes. Our data suggested that panicle length (PL) and plant height (PH) significantly affected panicle curvature (PC), with shorter PL and PH resulting in smaller PC and consequently more erect. A putative major gene was identified on chromosome 9 by molecular markers and bulk segregant analysis in DH population. In order to finely map the major gene, all simple sequence repeats (SSR) markers on chromosome 9 as well as 100 newly developed sequence-tagged site (STS) markers were used to construct a linkage group for quantitative trait locus (QTL) mapping. A major QTL, qPE9-1, between STS marker H90 and SSR marker RM5652, was detected, and accounted for 41.72% of PC variation with pleiotropic effect on PH and PL. another QTL, qPE9-2, was also found to be adjacent to qPE9-1. In addition, we found that H90, the nearest marker to qPE9-1, used for genotyping 38 cultivars with extremely erect and drooping panicles, segregated in agreement with PC, suggesting the H90 product was possibly part of the qPE9-1 gene or closely related to it. These data demonstrated that H90 could be used for marker-aided selection for the PE trait in breeding and in the cloning of qPE9-1.     

Genome-wide analysis of defense-responsive genes in bacterial blight resistance of rice mediated by the recessive<Emphasis Type="Italic"> R</Emphasis> gene<Emphasis Type="Italic"> xa13</Emphasis>

Defense responses triggered by dominant and recessive disease resistance ( R) genes are presumed to be regulated by different molecular mechanisms. In order to characterize the genes activated in defense responses against bacterial blight mediated by the recessive R gene xa13, two pathogen-induced subtraction cDNA libraries were constructed using the resistant rice line IRBB13—which carries xa13 —and its susceptible, near-isogenic, parental line IR24. Clustering analysis of expressed sequence tags (ESTs) identified 702 unique expressed sequences as being involved in the defense responses triggered by xa13; 16% of these are new rice ESTs. These sequences define 702 genes, putatively encoding a wide range of products, including defense-responsive genes commonly involved in different host-pathogen interactions, genes that have not previously been reported to be associated with pathogen-induced defense responses, and genes (38%) with no homology to previously described functional genes. In addition, R -like genes putatively encoding nucleotide-binding site/leucine rich repeat (NBS-LRR) and LRR receptor kinase proteins were observed to be induced in the disease resistance activated by xa13. A total of 568 defense-responsive ESTs were mapped to 588 loci on the rice molecular linkage map through bioinformatic analysis. About 48% of the mapped ESTs co-localized with quantitative trait loci (QTLs) for resistance to various rice diseases, including bacterial blight, rice blast, sheath blight and yellow mottle virus. Furthermore, some defense-responsive sequences were conserved at similar locations on different chromosomes. These results reveal the complexity of xa13 -mediated resistance. The information obtained in this study provides a large source of candidate genes for understanding the molecular bases of defense responses activated by recessive R genes and of quantitative disease resistance.Electronic Supplementary Material Supplementary material is available in the online version of this article at The first two authors contributed equally to this workCommunicated by R. Hagemann     

High-resolution genetic mapping at the <Emphasis Type="Italic">Bph15</Emphasis> locus for brown planthopper resistance in rice (<Emphasis Type="Italic">Oryza sativa</Emphasis> L.)

Resistance to the brown planthopper (BPH), Nilaparvata lugens Stål, a devastating sucking insect pest of rice, is an important breeding objective in rice improvement programs. Bph15, one of the 17 major BPH resistance genes so far identified in both cultivated and wild rice, has been identified in an introgression line, B5, and mapped on chromosome 4 flanked by restriction fragment length polymorphism markers C820 and S11182. In order to pave the way for positional cloning of this gene, we have developed a high-resolution genetic map of Bph15 by positioning 21 DNA markers in the target chromosomal region. Mapping was based on a PCR-based screening of 9,472 F₂ individuals derived from a cross between RI93, a selected recombinant inbred line of B5 bearing the resistance gene Bph15, and a susceptible variety, Taichung Native 1, in order to identify recombinant plants within the Bph15 region. Recombinant F₂ individuals with the Bph15 genotype were determined by phenotype evaluation. Analysis of recombination events in the Bph15 region delimited the gene locus to an interval between markers RG1 and RG2 that co-segregated with the M1 marker. A genomic library of B5 was screened using these markers, and bacterial artificial chromosome clones spanning the Bph15 chromosome region were obtained. An assay of the recombinants using the sub-clones of these clones in combination with sequence analysis delimited the Bph15 gene to a genomic segment of approximately 47 kb. This result should serve as the basis for eventual isolation of the Bph15 resistance gene.     

High-resolution mapping and gene prediction of <Emphasis Type="Italic">Xanthomonas Oryzae</Emphasis> pv. <Emphasis Type="Italic">Oryzae</Emphasis> resistance gene <Emphasis Type="Italic">Xa7</Emphasis>

Bacterial blight (BB) caused by Xanthomonas oryzae pv. oryzae (Xoo) is a devastating disease in rice worldwide. The resistance gene Xa7, which provides dominant resistance against the pathogen with avirulence (Avr) gene AvrXa7, has proved to be durably resistant to BB. A set of SSR markers were selected from the “gramene” database based on the Xa7 gene initial mapping region on chromosome 6. These markers were used to construct a high-resolution genetic map of the chromosomal region surrounding the Xa7 gene. An F₂ mapping population with 721 highly susceptible individuals derived from a cross between the near isogenic lines (NILs) IRBB7 and IR24 were constructed to localize the Xa7 gene. In a primary analysis with eleven polymorphic SSR markers, Xa7 was located in approximately the 0.28-cM region. To walk closer to the target gene, recombinant F₂ individuals were tested using newly developed STMS (sequence tagged microsatellite) markers. Finally, the Xa7 gene was mapped to a 0.21-cM interval between the markers GDSSR02 and RM20593. The Xa7-linked markers were landed on the reference sequence of cv. Nipponbare through bioinformatics analysis. A contig map corresponding to the Xa7 gene was constructed. The target gene was assumed to span an interval of approximately 118.5-kb which contained a total of fourteen genes released by the TIGR Genome Annotation Version 5.0. Candidate-gene analysis of Xa7 revealed that the fourteen genes encode novel domains that have no amino acid sequence similar to other cloned Xa(xa) genes. Shen Chen and Zhanghui Huang are contributed equally to this work.     

Microarray analysis of broad-spectrum resistance derived from an <Emphasis Type="Italic">indica</Emphasis> cultivar Rathu Heenati

Rathu Heenati (RHT) is a Sri Lankan rice cultivar that carries a brown planthopper (BPH) resistance gene, Bph3, and shows broad-spectrum resistance to all four biotypes of BPH. The BPH-resistance loci in RHT has been studied extensively and assigned to four different rice chromosomes (3, 4, 6, and 10) by different research groups, but the gene has not been cloned previously. An Affymetrix rice genome array containing 48,564 japonica and 1,260 indica sequences was used to analyze the potential resistance-related genes on the four chromosomes by comparative analysis of the differentially expressed genes between resistant and susceptible rice cultivars exposed to BPH attack. The microarray results showed that at least 17 genes related to induced resistance and at least 193 genes related to constitutive resistance in RHT. On chromosome 3, the AOC4 was hypothesized to be the most important candidate gene. On chromosome 6, no valuable candidate resistance gene was identified in the Bph3 localization region. In the three Quantitative trait locus regions of chromosomes 3, 4, and 10, the numbers of constitutive and induced resistance-related genes found were 17, 26, and 12, respectively. The major probe on chromosome 10 represents a constitutive expression gene with a very high absolute fold-change of 2,588.82. The microarray analysis indicated that BPH resistance in RHT is probably controlled by a series of resistance-related genes. This study provides valuable information for cloning, functional analysis and marker-assisted breeding of these BPH resistance genes.     

Fine mapping and candidate gene analysis of <Emphasis Type="Italic">ptgms2-1</Emphasis>, the photoperiod-thermo-sensitive genic male sterile gene in rice (<Emphasis Type="Italic">Oryza sativa</Emphasis> L.)

Photoperiod-thermo-sensitive genic male sterile (PTGMS) rice exhibits a number of desirable traits for hybrid rice production. The cloning genes responsible for PTGMS and those elucidating male sterility mechanisms and reversibility to fertility would be of great significance to provide a foundation to develop new male sterile lines. Guangzhan63S, a PTGMS line, is one of the most widely used indica two-line hybrid rice breeding systems in China. In this study, genetic analysis based on F₂ and BC₁F₂ populations derived from a cross between Guangzhan63S and 1587, determined a single recessive gene controls male sterility in Guangzhan63S. Molecular marker techniques combined with bulked-segregant analysis (BSA) were used and located the target gene (named ptgms2-1) between two SSR markers RM12521 and RM12823. Fine mapping of the ptgms2-1 locus was conducted with 45 new Insertion–Deletion (InDel) markers developed between the RM12521 and RM12823 region, using 634 sterile individuals from F₂ and BC₁F₂ populations. Ptgms2-1 was further mapped to a 50.4 kb DNA fragment between two InDel markers, S2-40 and S2-44, with genetic distances of 0.08 and 0.16 cM, respectively, which cosegregated with S2-43 located on the AP004039 BAC clone. Ten genes were identified in this region based on annotation results from the RiceGAAS system. A nuclear ribonuclease Z gene was identified as the candidate for the ptgms2-1 gene. This result will facilitate cloning the ptgms2-1 gene. The tightly linked markers for the ptgms2-1 gene locus will further provide a useful tool for marker-assisted selection of this gene in rice breeding programs.     

Fine genetic mapping and physical delimitation of the lesion mimic gene <Emphasis Type="Italic">spotted leaf 5</Emphasis> (<Emphasis Type="Italic">spl5</Emphasis>) in rice (<Emphasis Type="Italic">Oryza sativa</Emphasis> L.)

Spotted leaf 5 (spl5), a lesion mimic mutant, was first identified in rice (Oryza sativa L.) japonica cv. Norin8 in 1978. This mutant exhibits spontaneous disease-like lesions in the absence of any pathogens and resistance to rice blast and bacterial blight; however, the target gene has not yet been isolated. In the present study, we employed a map-based cloning strategy to finely map the spl5 gene. In an initial mapping with 100 F₂ individuals (spl5/spl5) derived from a cross between the spl5 mutant and indica cv. 93-11, the spl5 gene was located in a 3.3-cM region on chromosome 7 using six simple sequence repeat (SSR) markers. In a high-resolution genetic mapping, two F₂ populations with 3,149 individuals (spl5/spl5) were derived from two crosses between spl5 mutant and two indica cvs. 93-11 and Zhefu802 and six sequence-tagged site (STS) markers were newly developed. Finally, the spl5 gene was mapped to a region of 0.048 cM between two markers SSR7 and RM7121. One BAC/PAC contig map covering these markers’ loci and the spl5 gene was constructed through Pairwise BLAST analysis. Our bioinformatics analysis shows that the spl5 gene is located in the 80-kb region between two markers SSR7 and RM7121 with a high average ratio of physical to genetic distance (1.67 Mb/cM) and eighteen candidate genes. The analysis of these candidate genes indicates that the spl5 gene represents a novel class of regulators controlling cell death and resistance response in plants.     

<Emphasis Type="Italic">Pm37</Emphasis>, a new broadly effective powdery mildew resistance gene from <Emphasis Type="Italic">Triticum </Emphasis><Emphasis Type="Italic">timopheevii</Emphasis>

Powdery mildew is an important foliar disease in wheat, especially in areas with a cool or maritime climate. A dominant powdery mildew resistance gene transferred to the hexaploid germplasm line NC99BGTAG11 from T. timopheevii subsp. armeniacum was mapped distally on the long arm of chromosome 7A. Differential reactions were observed between the resistance gene in NC99BGTAG11 and the alleles of the Pm1 locus that is also located on chromosome arm 7AL. Observed segregation in F_2:3 lines from the cross NC99BGTAG11 × Axminster (Pm1a) demonstrate that germplasm line NC99BGTAG11 carries a novel powdery mildew resistance gene, which is now designated as Pm37. This new gene is highly effective against all powdery mildew isolates tested so far. Analyses of the population with molecular markers indicate that Pm37 is located 16 cM proximal to the Pm1 complex. Simple sequence repeat (SSR) markers Xgwm332 and Xwmc790 were located 0.5 cM proximal and distal, respectively, to Pm37. In order to identify new markers in the region, wheat expressed sequence tags (ESTs) located in the distal 10% of 7AL that were orthologous to sequences from chromosome 6 of rice were targeted. The two new EST-derived STS markers were located distal to Pm37 and one marker was closely linked to the Pm1a region. These new markers can be used in marker-assisted selection schemes to develop wheat cultivars with pyramids of powdery mildew resistance genes, including combinations of Pm37 in coupling linkage with alleles of the Pm1 locus.     

Fine mapping of the recessive genic male sterility gene (<Emphasis Type="Italic">Bnms3</Emphasis>) in <Emphasis Type="Italic">Brassica napus</Emphasis> L.

The Brassica napus oilseed rape line, 7-7365AB, is a recessive epistatic genic male sterile (RGMS) two-type line system. The sterility is controlled by two pairs of recessive duplicate genes (Bnms3 and Bnms4) and one pair of recessive epistatic inhibitor gene (Bnrf). Homozygosity at the Bnrf locus (Bnrfrf) inhibits the expression of the two recessive male sterility genes in homozygous Bnms3ms3ms4ms4 plants and produces a male fertile phenotype. This line has a good potential for heterosis utilization but it is difficult to breed heterotic hybrids without molecular markers. To develop markers linked to the BnMs3 gene, amplified fragment length polymorphism (AFLP) technology was applied to screen the bulks of sterile and fertile individuals selected randomly from a population of near-isogenic lines (NIL) consisting of 2,000 plants. From a survey of 1,024 primer combinations, we identified 17 AFLP markers linked to the BnMs3 gene. By integrating the previous markers linked to the BnMs3 gene into the genetic map of the NIL population, two markers, EA01MC12 and EA09P06, were located on either side of the BnMs3 gene at a distance of 0.1 and 0.3 cM, respectively. In order to use the markers for male sterile line breeding, five AFLP markers, P05MG05, P03MG04, P11MG02, P05MC11₂₅₀, and EA09P06, were successfully converted into sequence characterized amplified region (SCAR) markers. Two of these, P06MG04 and sR12384, were subsequently mapped on to linkage group N19 using two doubled-haploid mapping populations available at our laboratory derived from the crosses Tapidor × Ningyou7 and Quantum × No2127-17. The markers found in the present study should improve our knowledge of recessive genic male sterility (RGMS), and accelerate the development of male sterile line breeding and map-based cloning.     

Development and validation of KASP markers for the greenbug resistance gene <Emphasis Type="Italic">Gb7</Emphasis> and the Hessian fly resistance gene <Emphasis Type="Italic">H32</Emphasis> in wheat

Key message

Greenbug and Hessian fly are important pests that decrease wheat production worldwide. We developed and validated breeder-friendly KASP markers for marker-assisted breeding to increase selection efficiency.

Abstract

Greenbug (Schizaphis graminum Rondani) and Hessian fly [Mayetiola destructor (Say)] are two major destructive insect pests of wheat (Triticum aestivum L.) throughout wheat production regions in the USA and worldwide. Greenbug and Hessian fly infestation can significantly reduce grain yield and quality. Breeding for resistance to these two pests using marker-assisted selection (MAS) is the most economical strategy to minimize losses. In this study, doubled haploid lines from the Synthetic W7984 × Opata M85 wheat reference population were used to construct linkage maps for the greenbug resistance gene Gb7 and the Hessian fly resistance gene H32 with genotyping-by-sequencing (GBS) and 90K array-based single nucleotide polymorphism (SNP) marker data. Flanking markers were closely linked to Gb7 and H32 and were located on chromosome 7DL and 3DL, respectively. Gb7-linked markers (synopGBS773 and synopGBS1141) and H32-linked markers (synopGBS901 and IWB65911) were converted into Kompetitive Allele Specific PCR (KASP) assays for MAS in wheat breeding. In addition, comparative mapping identified syntenic regions in Brachypodium distachyon, rice (Oryza sativa), and sorghum (Sorghum bicolor) for Gb7 and H32 that can be used for fine mapping and map-based cloning of the genes. The KASP markers developed in this study are the first set of SNPs tightly linked to Gb7 and H32 and will be very useful for MAS in wheat breeding programs and future genetic studies of greenbug and Hessian fly resistance.

     

Pea powdery mildew <Emphasis Type="Italic">er1</Emphasis> resistance is associated to loss-of-function mutations at a <Emphasis Type="Italic">MLO</Emphasis> homologous locus

The powdery mildew disease affects several crop species and is also one of the major threats for pea (Pisum sativum L.) cultivation all over the world. The recessive gene er1, first described over 60 years ago, is well known in pea breeding, as it still maintains its efficiency as a powdery mildew resistance source. Genetic and phytopathological features of er1 resistance are similar to those of barley, Arabidopsis, and tomato mlo powdery mildew resistance, which is caused by the loss of function of specific members of the MLO gene family. Here, we describe the obtainment of a novel er1 resistant line by experimental mutagenesis with the alkylating agent diethyl sulfate. This line was found to carry a single nucleotide polymorphism in the PsMLO1 gene sequence, predicted to result in premature termination of translation and a non-functional protein. A cleaved amplified polymorphic sequence (CAPS) marker was developed on the mutation site and shown to be fully co-segregating with resistance in F₂ individuals. Sequencing of PsMLO1 from three powdery mildew resistant cultivars also revealed the presence of loss-of-function mutations. Taken together, results reported in this study strongly indicate the identity between er1 and mlo resistances and are expected to be of great breeding importance for the development of resistant cultivars via marker-assisted selection.     

Fine Mapping QTL for Drought Resistance Traits in Rice (<Emphasis Type="Italic">Oryza sativa</Emphasis> L.) Using Bulk Segregant Analysis

Drought stress is a major limitation to rice (Oryza sativa L.) yields and its stability, especially in rainfed conditions. Developing rice cultivars with inherent capacity to withstand drought stress would improve rainfed rice production. Mapping quantitative trait loci (QTLs) linked to drought resistance traits will help to develop rice cultivars suitable for water-limited environments through molecular marker-assisted selection (MAS) strategy. However, QTL mapping is usually carried out by genotyping large number of progenies, which is labour-intensive, time-consuming and cost-ineffective. Bulk segregant analysis (BSA) serves as an affordable strategy for mapping large effect QTLs by genotyping only the extreme phenotypes instead of the entire mapping population. We have previously mapped a QTL linked to leaf rolling and leaf drying in recombinant inbred (RI) lines derived from two locally adapted indica rice ecotypes viz., IR20/Nootripathu using BSA. Fine mapping the QTL will facilitate its application in MAS. BSA was done by bulking DNA of 10 drought-resistant and 12 drought-sensitive RI lines. Out of 343 rice microsatellites markers genotyped, RM8085 co-segregated among the RI lines constituting the respective bulks. RM8085 was mapped in the middle of the QTL region on chromosome 1 previously identified in these RI lines thus reducing the QTL interval from 7.9 to 3.8 cM. Further, the study showed that the region, RM212–RM302–RM8085–RM3825 on chromosome 1, harbours large effect QTLs for drought-resistance traits across several genetic backgrounds in rice. Thus, the QTL may be useful for drought resistance improvement in rice through MAS and map-based cloning.     

<Emphasis Type="Italic">STAYGREEN</Emphasis> (<Emphasis Type="Italic">CsSGR</Emphasis>) is a candidate for the anthracnose (<Emphasis Type="Italic">Colletotrichum orbiculare</Emphasis>) resistance locus <Emphasis Type="Italic">cla</Emphasis> in Gy14 cucumber

Key message

Map-based cloning identified a candidate gene for resistance to the anthracnose fungal pathogen Colletotrichum orbiculare in cucumber, which reveals a novel function for the highly conserved STAYGREEN family genes for host disease resistance in plants.

Abstract

Colletotrichum orbiculare is a hemibiotrophic fungal pathogen that causes anthracnose disease in cucumber and other cucurbit crops. No host resistance genes against the anthracnose pathogens have been cloned in crop plants. Here, we reported fine mapping and cloning of a resistance gene to the race 1 anthracnose pathogen in cucumber inbred lines Gy14 and WI 2757. Phenotypic and QTL analysis in multiple populations revealed that a single recessive gene, cla, was underlying anthracnose resistance in both lines, but WI2757 carried an additional minor-effect QTL. Fine mapping using 150 Gy14?×?9930 recombinant inbred lines and 1043 F₂ individuals delimited the cla locus into a 32 kb region in cucumber Chromosome 5 with three predicted genes. Multiple lines of evidence suggested that the cucumber STAYGREEN (CsSGR) gene is a candidate for the anthracnose resistance locus. A single nucleotide mutation in the third exon of CsSGR resulted in the substitution of Glutamine in 9930 to Arginine in Gy14 in CsSGR protein which seems responsible for the differential anthracnose inoculation responses between Gy14 and 9930. Quantitative real-time PCR analysis indicated that CsSGR was significantly upregulated upon anthracnose pathogen inoculation in the susceptible 9930, while its expression was much lower in the resistant Gy14. Investigation of allelic diversities in natural cucumber populations revealed that the resistance allele in almost all improved cultivars or breeding lines of the U.S. origin was derived from PI 197087. This work reveals an unknown function for the highly conserved STAYGREEN (SGR) family genes for host disease resistance in plants.