{"paper_id":"13bcb4eb-4b1e-4d0e-bb2a-8032c03a0258","body_text":"Identification and genetic mapping of a rust resistance gene on chromosome Pv04 in the common bean cultivar CNC using bulked segregant sequencing | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Research Article Identification and genetic mapping of a rust resistance gene on chromosome Pv04 in the common bean cultivar CNC using bulked segregant sequencing Giseli Valentini, Anuj Sharma, Jatinder Singh, Upinder Gill This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-6572527/v1 This work is licensed under a CC BY 4.0 License Status: Posted Version 1 posted You are reading this latest preprint version Abstract Rust, caused by the fungus Uromyces appendiculatus , is one of the most significant diseases of common beans in the United States and worldwide. The identification and characterization of new rust resistance genes are essential for developing cultivars with broad and durable resistance. The black common bean cultivar Compuesto Negro Chimaltenango (CNC) exhibits broad resistance to most known races of the rust pathogen, still its resistance has not been fully characterized. To genetically characterize and map the resistance loci in this cultivar, we crossed it with susceptible cultivar UI114 and evaluated the F 2 population for resistance against races 20-6 and 31-1 of U. appendiculatus . Our results showed that rust resistance in CNC for races 20-6 and 31-1 co-segregates and is conditioned by a single dominant gene. Whole genome sequencing-based bulked segregant analysis identified a genomic region on the proximal end of chromosome 4 associated with rust resistance in CNC. To further validate the identified region, molecular markers spanning this region were mapped on the complete F 2 population. This further delimited the resistance locus in a genomic interval of 653.8 kb. The genetic mapping information and the molecular markers developed in this study will be helpful in developing rust-resistant cultivars of common bean. Phaseolus vulgaris L. Uromyces appendiculatus Compuesto Negro Chimaltenango Bulked segregant analysis resistance mapping Figures Figure 1 Figure 2 Figure 3 Key Message Genetic mapping of the rust resistance gene in the common bean cultivar Compuesto Negro Chimaltenango (CNC) using NGS-BSA. Introduction Common bean ( Phaseolus vulgaris L.) is a highly nutritious food and part of the daily diet of about 400 million people in the tropics (CIAT 2024 ). Especially in Latin America and Africa, beans play an important role as the primary source of protein for direct human consumption (Broughton et al. 2003 ). Diseases are often the most severe constraint in the common bean production, negatively impacting the quality and yield. Rust, caused by the fungus Uromyces appendiculatus , is a disease with worldwide distribution, aligning with the distribution of its hosts. Although the disease occurs in many parts of the world, the yield losses are often more severe when common beans are cultivated under warm and humid conditions (Schwartz and Pastor-Corrales 1989 ; Stavely and Pastor-Corrales 1989). The adoption of cultivars with genetic resistance is the most cost-effective and practical strategy to control rust (Miklas et al. 2006 ). In the US, the introduction of rust resistance genes in the mid-1980s was crucial to reduce chemical applications primarily used to control rust in common bean (MacQueen et al. 2020 ). Indeed, using disease-resistant cultivars lowers production costs while averting yield loss, and reduces the dependence on chemical control and the negative environmental impact caused by their application (Miklas et al. 2006 ). However, the wide and complex virulence diversity of U. appendiculatus , distributed into several races, hinders the development and durability of resistance in common bean cultivars (Stavely et al. 1989 ). U. appendiculatus is an autoecious fungus that can potentially undergo sexual reproduction to produce new virulent strains via recombination (McMillan et al. 2003 ). The complexity of the U. appendiculatus races coupled with the potential for the breakdown of monogenic resistances in bean cultivars necessitates the identification of additional resistance ( R ) genes and their pyramiding in elite germplasm (Pastor-Corrales 2002 ; 2003 ; 2006 ). The deployment of a combination of multiple R genes in a single cultivar is an important strategy for obtaining effective and durable genetic resistance to rust (Stavely et al. 1989 ; Pastor-Corrales 2006 ). Genes conditioning rust resistance are recognized by the Bean Improvement Cooperative (BIC) Committee and identified by the Ur- symbol (Kelly et al. 1996 ). The Ur-3 , Ur-5, Ur-7 , Ur-11 , and Ur-14 are from the Middle American bean gene pool (Augustin et al. 1972 ; Ballantyne 1978 ; Stavely 1984 ; Stavely 1990 ; Souza et al. 2011 ), and Ur-4, Ur-6, Ur-9, Ur-12 , and Ur-13 are from the Andean gene pool (Ballantyne 1978 ; Luannfinke et al. 1986 ; Jung et al. 1998 ; Liebenberg and Pretorius 2004 ). These genes have been mapped on six chromosomes of common bean (Pv01, Pv04, Pv06, Pv07, Pv08, and Pv11). All Ur- genes, but Ur-14 , were mapped in the 12 differential cultivars for U. appendiculatus race characterization. This set was proposed by Steadman et al. ( 2002 ) and comprises six Andean and six Middle American cultivars. Compuesto Negro Chimaltenango (CNC), which is one of 12 differentials and presents broad resistance to multiple rust races, still needs to be genetically characterized in detail. CNC is a black bean cultivar from the Middle American gene pool and originally from Guatemala. This cultivar was included in the first set of differential lines developed by Stavely et al. ( 1983 ), and was retained as part of the updated set of rust differential lines proposed in 2002 (Steadman et al. 2002 ). The resistance in CNC is effective against most races of bean rust (Stavely 1984 ; Stavely et al. 1989 ). Despite its rust resistance, CNC has not been extensively used in cultivar development, and the genetic basis of its resistance remains poorly understood. Previous reports indicate CNC possesses a single dominant gene that conditions rust resistance against race 49 of U. appendiculatus (Rasmussen et al. 2002 ). Although other races were not tested on CNC, the presence of multiple resistance genes in the line has been suggested (Rasmussen et al. 2002 ). Given the importance of highly effective rust resistance in CNC, we present its genetic characterization, mapping, and the development of molecular markers to facilitate its potential use in future bean cultivars. Material and Methods Plant material The F 1 and F 2 populations were developed from the cross between UI114 and CNC at North Dakota State University, Fargo, ND. CNC is resistant to most of the known races of U. appendiculatus , while UI114, a pinto bean cultivar, is highly susceptible. The dominant violet flower trait, inherited from the male parent CNC, was observed in the F 1 plants, confirming that they were hybrids. A total of 275 F 2 plants were obtained by F 1 self-pollination and used for the inheritance test. Rust phenotyping The races used in this study were obtained from the collection maintained at North Dakota State University. Six F 1 plants from the cross UI114 × CNC were inoculated with races 13 − 2, 20 − 6, 31 − 1, and 20 − 3 of U. appendiculatus . Later, races 20 − 6 and 31 − 1 of U. appendiculatus were selected to study the genetic inheritance on the F 2 population. Seeds of the F 1 and F 2 populations were planted in 12.7-cm diameter pots containing Pro-mix BX general purpose growing mix (Premier Tech) and maintained under greenhouse conditions at 20 ± 5°C. Inoculations of U. appendiculatus were performed nine days after planting when the primary leaves were about 2/3 expanded. Parental lines and the cultivars Aurora ( Ur-3 ), Early Gallatin ( Ur-4 ), Mexico 309 ( Ur-5 ), Golden Gate Wax ( Ur-6 ), and ND Falcon ( Ur-11 ) were used as internal controls. The inoculum solution was prepared by suspending urediniospores of U. appendiculatus in distilled water containing Tween 20 (0.01%) to obtain a concentration of 2 × 10 4 uredospores mL − 1 . The F 1 and F 2 populations were simultaneously inoculated with four and two races of U. appendiculatus , respectively, on the abaxial side of the primary leaves using a cotton swab containing the spore solution, avoiding contamination between races. After inoculation, plants were transferred to a mist chamber (20 ± 1°C, relative humidity > 95%) for 18 hours, under darkness. After this period, plants were relocated to the greenhouse and kept for approximately 10–12 days, until disease symptoms appeared. Disease evaluation were performed using a 1–6 scale proposed by Stavely et al. ( 1983 ), where 1 = without any visible rust reactions; 2 = necrotic spots without sporulation (hypersensitive reaction or HR); 2 + = necrotic spots 300–1000 µm (1 mm) in diameter; 2++ = necrotic spots 1–3 mm in diameter; 2+++ = necrotic spots larger than 3 mm in diameter; 3 = tiny sporulating uredinia (rust pustules) less than 300 µm in diameter; 4 = sporulating uredinia 300–500 µm in diameter; 5 = sporulating uredinia 500–800 µm in diameter; and 6 = sporulating uredinia larger than 800 µm in diameter. Plants with grades 1, 2 and 3 were classified as resistant, whereas plants with grades 4, 5, and 6 were considered susceptible. Evaluations including more than one grade were recorded and separated by “,” with the most predominant type of reactions listed first. The chi-square (χ 2 ) test was performed to study the genetic segregation (resistant/susceptible) ratios. Next generation sequencing based bulked segregant analysis (NGS-BSA) Based on the disease phenotype of the F 2 population, plants were selected for NGS-BSA. Two bulks were developed: a resistant bulk containing 18 F 2 plants resistant to races 20 − 6 and 31 − 1 and a susceptible bulk containing 18 F 2 plants susceptible to races 20 − 6 and 31 − 1. Young leaves were collected from each plant and lyophilized for further DNA extraction using the OmniPrep DNA extraction kit (G-Biosciences, St. Louis, MO) following the manufacture’s protocol with a few modifications. DNA purity and concentration were verified using a NanoDrop (Thermo Fisher Scientific, Waltham, MA). For the two bulks, DNA of plants were bulked separately in equimolar concentrations based on the NanoDrop readings. The paired-end DNA sequencing data (2×150 bp) from bulks were generated using Illumina NovaSeq X Plus by outsourcing to Novogene ( www.novogene.com ). For data analysis viz. , quality check, adapter trimming, read mapping and variant calling, Bulk2SNPs pipeline ( https://github.com/NDSUrustlab/Bulk2SNPs ) was used. This pipeline uses a set of bioinformatics software, such as fastqc (Andrews 2010 ), fastp (Chen 2023 ), bwa-mem2 (Vasimuddin et al. 2019), and GATK (McKenna et al. 2010 ) for analyzing raw sequencing data from two bulks and output a Single Nucleotide Polymorphism (SNP) table file for downstream QTL analysis. The reference genome P. vulgaris UI111 v1.1 ( https://phytozome-next.jgi.doe.gov/ ) was used for mapping the sequence reads for each bulk. For QTL analysis, we used an R package, QTLseqr (Mansfeld and Grumet 2018 ) available at https://github.com/bmansfeld/QTLseqr . QTLseqr implements the bulked segregant analysis via G’ and Δ (SNP-index), described by Magwene et al. ( 2011 ) and Takagi et al. ( 2013 ), respectively. Low-confidence SNPs were filtered out based on the reference allele frequency (refAlleleFreq = 0.05), and minimum and maximum read depth (minTotalDepth = 40 and maxTotalDepth = 400). Identification of candidate QTL regions was performed using a 2 Mb sliding window, whereby the confidence intervals for the ΔSNP-indices was determined using 10,000 simulations. Marker development and genetic mapping Individual F 2 plants were genotyped to confirm marker-trait association. The targeted genomic region containing the gene of interest identified by NGS-BSA was examined for SNPs and used for the development of PCR allelic competitive extension (PACE) markers (3CR Bioscience, Essex, UK). Alignment sequences were visualized in the Integrative Genomics Viewer (Robinson et al. 2011 ) for the manual selection of SNPs. Variants showing A/T⇄C/G substitutions that did not have another variant within the approximately 100 bp flanking region were considered for marker development. Using the reference genome, the selected sequences were retrieved and submitted to 3CR Bioscience for primer design. For genotyping, genomic DNA from F 2 plants was extracted using the protocol described by Kaur et al. ( 2023 ). The polymerase chain reaction (PCR) for PACE primers was performed using 2.5 µl of premade PACE® genotyping master mix (3CR Bioscience, Essex, UK), 0.07 µl of primers mix (Sigma-Aldrich, St. Louis, MO), and 20–40 ng of genomic DNA, for a 5 µl reaction. PCR reactions were performed in a CFX Opus 384 Real Time PCR System (Bio-Rad, Hercules, CA) with the following parameters:15 min at 94 o C, 10 cycles of 20 sec at 94 o C and 60 sec at 57 o C, and 30 cycles of 20 sec at 94 o C and 60 sec at 57 o C. PCR results were analyzed in the Bio-Rad CFX Manager 3.1 (Bio-Rad, Hercules, CA). Genetic mapping of the rust resistance locus in CNC using PACE markers was performed with JoinMap 4.1 (Van Ooijen 2006 ). Default settings for the Maximum Likelihood Mapping algorithm were used to define the linkage order and distances (cM). The genetic distances were later used to graphically design the linkage groups using MapChart 2.3 (Voorrips 2002 ). Results Genetic segregation analysis of the F 2 mapping population To investigate the genetics of rust resistance in CNC, we performed inheritance analyses, bulked segregant sequencing, and genetic mapping. For the inheritance study, we crossed the susceptible cultivar UI114 with the resistant CNC line. CNC displayed a highly resistant reaction when inoculated with races 13 − 2, 20 − 6, 31 − 1, and 20 − 3, characterized by small pustules and hypersensitive reaction. In contrast, UI114 exhibited a classical susceptible reaction characterized by large, sporulating pustules to all tested rust races. The F 1 plants from the cross UI114 × CNC exhibited a resistant reaction to races 13 − 2, 20 − 6, 31 − 1, and 20 − 3, which was identical to that of the resistant parental line CNC, suggesting that the rust resistance in CNC is a dominant trait (Fig. 1 ). Phenotypic evaluations of 275 F 2 individuals indicated that the disease reactions to races 20 − 6 and 31 − 1 co-segregated, meaning that when an F 2 plant was resistant to race 20 − 6, it was also resistant to race 31 − 1 and vice versa. We observed the segregation of 207 resistant and 68 susceptible plants, fitting a ratio of 3 resistant to 1 susceptible (χ² = 0.011, P value = 0.9168), confirming that the rust resistance in CNC to races 20 − 6 and 31 − 1 is conferred by a single dominant gene (Table S1 , Table 1 ). Table 1 Observed and expected reaction for the F 1 and F 2 populations from the cross UI114 × CNC inoculated with different races of U. appendiculatus Parent/Cross Races Generation Observed ratio (R:S) Expected ratio (R:S) χ 2 P value UI114 13 − 2, 20 − 6, 31 − 1, 20 − 3 SP 0:16 CNC 13 − 2, 20 − 6, 31 − 1, 20 − 3 RP 16:0 UI114 × CNC 13 − 2, 20 − 6, 31 − 1, 20 − 3 F 1 6:0 UI114 × CNC 20 − 6, 31 − 1 F 2 207:68 3:1 0.011 0.9168 RP, resistant parent; SP, susceptible parent; R, resistant; S, susceptible Identification of genomic regions associated with resistance using NGS-BSA To identify the genomic region(s) involved in rust resistance, the bulked segregant analysis was performed using whole genome sequencing. Two bulks representing 18 highly resistant (R) and 18 highly susceptible (S) F 2 plants were generated by pooling equimolar concentrations of genomic DNA of each plant for whole genome sequencing. A total of 19.3 and 18.8 GB of Illumina sequence data were generated for the R and S bulks, respectively, representing an average coverage of 34X for the R bulk and 33X for the S bulk of the assembled common bean genome. For the R bulk, 137,938,390 reads were generated, of which 136,499,352 were mapped to the UI111 v1.1 reference genome; for the susceptible bulk, 134,460,710 reads were generated, of which 132,993,492 were mapped to the reference. A comparison of the R and S bulks to the reference genome identified 1,897,742 SNPs. After filtering, 1,421,905 high-quality SNPs were retained for QTL mapping using QTLseqr (Mansfeld and Grumet, 2018 ). SNPs were more frequently distributed in euchromatic regions, with a higher number observed on chromosomes Pv04, Pv10, and Pv11 (Fig. 2 ). The QTL-seq analysis, based on the delta SNP index and the G’ statistics, consistently showed one major peak on chromosome Pv04 between 4,766 and 10,983,713 bp, suggesting the strong co-segregation between the rust phenotype and genotype in this region (Fig. 2 , Table S2). Four apparent minor peaks on chromosome 4 and a peak on chromosome 8 were also observed. These minor peaks were significant based on G’ statistics only. Therefore, subsequent validation efforts focused on delimiting the resistance locus within the subtelomeric region of Pv04, where the major peak was observed. Genetic mapping of the rust resistance gene in CNC To further validate the genomic region identified by NGS-BSA on Pv04, the SNPs within the region were identified to develop PACE markers for genotyping on the F 2 population. A total of 11 PACE markers were developed between 415,751 and 1,874,106 bp and genotyped on 275 F 2 plants from UI114 × CNC (Table S3). The rust resistance locus in CNC was linked at 0.0 cM to markers Pv859224, Pv1178763, and Pv1243509, and positioned 0.8 cM downstream of marker Pv696542 and 0.2 cM upstream of marker Pv1350344 (Fig. 3 ). The resistance locus was located between 696,542 and 1,350,344 bp, spanning a physical genomic interval of 653.8 Kb (Table 2 ). Table 2 Reaction to Uromyces appendiculatus and genotypes at eleven SNP loci of F 2 plants from the cross UI114 × CNC Number of F 2 plants Phenotype to races 20 − 6 and 31 − 1 Genotyping of F 2 plants with PACE markers 1 Pv567309 Pv576749 Pv626590 Pv696542 Pv859224 Pv1178763 Pv1243509 Pv1350344 Pv1604777 Pv1713242 Pv1776022 567,309 2 576,749 626,590 696,542 859,224 1,178,763 1,243,509 1,350,344 1,604,777 1,713,242 1,776,022 UI114 Susceptible AA AA AA AA AA AA AA AA AA AA AA CNC Resistant BB BB BB BB BB BB BB BB BB BB BB Genomic region containing the rust resistance gene in CNC 3 Resistant AA AA AA AA AB AB AB AB AB AB AB 1 Resistant AA AA AA AB AB AB AB AB AB AB AB 1 Resistant AA AA AB AB AB AB AB AB AB AB AB 2 Resistant BB BB BB BB BB BB BB BB BB AB AB 1 Resistant BB BB BB BB BB BB BB BB BB BB AB 72 Resistant BB BB BB BB BB BB BB BB BB BB BB 125 Resistant AB AB AB AB AB AB AB AB AB AB AB 1 Resistant AB AB AB AB AB AB AB AB AA AA AA 1 Resistant AB AB AB AB AB AB AB AA AA AA AA 1 Susceptible AB AB AB AB AA AA AA AA AA AA AA 1 Susceptible AB AB AA AA AA AA AA AA AA AA AA 65 Susceptible AA AA AA AA AA AA AA AA AA AA AA 1 Susceptible AA AA AA AA AA AA AA AA AA AB AB 1 SNP position based on the reference genome of UI111 v1.1; AA = UI114 allele; AB heterozygous; BB = CNC allele The genomic interval delimited by markers Pv696542 and Pv1350344 contained 56 annotated genes according to the UI111 v1.1 reference genome. Among the annotated genes, 11 encode disease-resistance proteins involved in plant defense against pathogens; eight of these genes encode nucleotide-binding domain leucine-rich repeat (NLR) proteins, and three genes encode kinase proteins (Table S4). To further investigate, we used the NCBI Conserved Domain Search tool ( https://www.ncbi.nlm.nih.gov/Structure/cdd/wrpsb.cgi ) to identify conserved domains within these 11 genes. Two genes, which are annotated as NLRs in the UI111 reference genome v1.1, did not display any domains typically associated with plant resistance. The remaining nine genes exhibited either NLR or kinase domains and are strong candidates for the rust resistance gene present in CNC (Table 3 ). Table 3 Candidate genes encoding proteins related to disease resistance in the genomic region containing the rust resistance gene in CNC on the common bean reference genome UI111 v1.1 Gene Start End Predict protein PvUI111.04G007600 709963 714521 Leucine-rich repeat-containing protein PvUI111.04G007900 723,321 726,702 Leucine-rich repeat-containing protein PvUI111.04G008100 729,478 734,847 Leucine-rich repeat-containing protein PvUI111.04G008800 813,803 817,714 Serine/threonine-protein kinase PvUI111.04G009400 889,655 895,350 Cysteine-rich receptor-like protein kinase PvUI111.04G011000 1,170,791 1,183,177 Protein kinase PvUI111.04G011200 1,213,139 1,218,585 Leucine-rich repeat-containing protein PvUI111.04G011300 1,221,763 1,225,645 Leucine-rich repeat-containing protein PvUI111.04G011400 1,229,108 1,235,266 Leucine-rich repeat-containing protein Discussion U. appendiculatus has high genetic diversity and it is constantly evolving to overcome the resistance genes deployed in bean cultivars. In this study, we characterized a rust resistance locus in the cultivar CNC and mapped it to chromosome Pv4 of P. vulgaris . In addition, we identified closely linked SNP-based markers, which can be used by breeding programs in marker-assisted selection (MAS). To our knowledge, the only rust resistance germplasm derived from CNC are the pintos BelDak-RR-1 and BelDak-RR-2, which also contain Ur-3 and Ur-6 (Stavely and Grafton 1989 ). CNC is resistant to 81 of 88 races maintained in the U. appendiculatus collection at the USDA-ARS in Beltsville MD, and it is particularly highly resistant to races present in the United States (Stavely 1984 ; Stavely et al. 1989 ). In North Dakota, which is the leading U.S. bean producer, bean rust is one of the major yield-limiting factors. Therefore, increased efforts have been made to improve rust resistance in bean germplasm since the mid-1980s (MacQueen et al. 2020 ; Osorno et al. 2021 ; Miklas et al. 2023 ). Until 2002, five U. appendiculatus races (52, 54, 69, 70, and 71) were reported in North Dakota, none of which were virulent on CNC (Gross and Venette 2002 ). More recent surveys showed that 20 − 3 is the most frequent race found in dry bean fields in North Dakota with 50–70% of the rust isolates belonging to this race (Monclova-Santana 2019 ). CNC serves as an important source of resistance to race 20 − 3 and can be used by breeding programs aiming to develop rust resistance cultivars for North Dakota (Monclova-Santana 2019 ). To understand the genetics of rust resistance in CNC, we employed BSA coupled with NGS on an F 2 population from cross UI114 × CNC for the rapid identification of genes/genomic loci in CNC governing the rust resistance to races 20 − 6 and 31 − 1. NGS-BSA resulted in the identification of a genomic region associated with resistance on chromosome Pv04 which was further confirmed by genotyping the complete F 2 population using PACE markers. Chromosome Pv04 comprises one of the most complex clusters of rust resistance in common bean, including genes reported in the Middle American cultivars Mexico 309 ( Ur-5 ), Ouro Negro ( Ur-14 ), Dorado, and PI 310762, and Andean cultivars PI 260418 and G19833 (Miklas et al. 2000 ; Shin et al. 2014 ; Valentini et al. 2020 ; Valentini et al. 2021 ). An allelism test has confirmed the independence of Ur-CNC and Ur-14 , but allelism tests with genotypes containing other genes have never been performed (Souza et al. 2011 ). Although the rust resistance in CNC can be distinguished from the above-mentioned cultivars by its unique spectrum of resistance, the overlap of the CNC region with previously identified genomic locations of other resistance genes add further complexities about its independence. Specifically, the gene in CNC was mapped between 696,542 and 1,350,344 bp (UI111 v1.0) the gene in PI 310762 is positioned between 246,091 and 1,164,610 bp (G19833 v1.0); the Ur-5 gene in Mexico 309 is mapped between 381,360 and 1,407,905 bp (G19833 v1.0); the gene in PI260418 is mapped between 303,697 and 1,299,082 bp (G19833 v2.1); and the gene in G19833 lies between 554,115 and 1,301,156 bp (G19833 v2.1) (Shin et al. 2014 ; Valentini et al. 2020 ; Valentini et al. 2021 ). Therefore, additional allelism tests with large segregating populations and fine mapping will be crucial to confirm the independence of these genes. Several features make the resistance gene cluster on Pv04 particularly interesting for further investigations. Firstly, in contrast to other resistance gene clusters, the Pv04 cluster contains disease-resistance genes from both the Andean and Middle American gene pools, suggesting that this cluster existed prior to the geographic separation of P. vulgaris into the two gene pools (Geffroy et al. 2009 ; Geffroy et al. 1999 ; Vlasova et al. 2016 ). On the other hand, resistance gene clusters on Pv01 and Pv11 seem to be originated independently in each gene pool. For example, Pv01 contains genes of Andean origin for resistance to rust ( Ur-9 ) and anthracnose ( Co-1, Co-x, Co-AC , and Co-Pa ) (Miklas et al. 2006 ; Richard et al. 2014 ; Castro et al. 2017 ; Gilio et al. 2020 ), and Pv11 contains genes of Middle American origin for resistance to rust ( Ur-3, Ur-7 , and Ur-11 ) and anthracnose ( Co-2 ) (Miklas et al. 2006 ), suggesting that the resistance gene clusters might be derived from specific gene pools. Secondly, the majority of the resistance-associated genes in the Pv04 cluster encode typical NLR domains, although genes encoding kinase domains are also present (Schmutz et al. 2014 ; Vas Bisneta and Gonçalves-Vidigal 2020). The combination of these two pathogen recognition systems provides the host with greater flexibility, allowing for the rapid evolution of resistance genes and enabling the plant to adapt to new or evolving pathogens over time. Although it has not been elucidated in common beans, the interaction of an NLR and Kinases was recently demonstrated in wheat, showing that an NLR with tandem NB-ARC domains (WTN1) cooperates with a tandem kinase (WTK3 and its allelic variant Rwt4 to constitute a tandem kinase-NLR immune signaling pathway (Lu et al. 2025 ). In another example, Chen et al. ( 2025 ) discovered that the Sr62 locus consists of a digenic module encoding the tandem kinase Sr62 TK and an NLR (Sr62 NLR ). The pathogen effector AvrSr62 triggers the activation of the wheat Sr62 TK , which then activates the corresponding NLR. Due to the low recombination rates in certain Pv04 regions, it is particularly challenging to narrow down a genomic interval containing the resistance gene. The region containing the gene cluster associated with disease resistance on top of Pv04 originates from ectopic recombination between subtelomeric regions of nonhomologous chromosomes. Specifically, genes encoding Coiled-Coil-Nucleotide-Binding-Site-Leucine-Rich-Repeat (CNL) on the Pv04 gene cluster were derived from CNL of the Pv11 cluster through an ectopic recombination event that occurred more than 19 MYA before the divergence of P. vulgaris and Glycine max (David et al. 2009 ; McClean et al. 2010 ). This ectopic recombination resulted in the occurrence of heterochromatic blocks (David et al. 2009 ), which may be associated with recombination suppression in this region as reported in other studies (Meziadi et al. 2016 , Valentini et al. 2017 ). In summary, we have genetically characterized and mapped the rust resistance in CNC, which exhibits broad resistance against multiple races of U. appendiculatus . The strong resistance conferred by CNC highlights its potential value for enhancing common bean germplasm. Furthermore, the development of molecular markers in this study offers valuable tools for diversifying the deployment of rust resistance in common bean breeding programs. Declarations Conflict of interest The authors declare that there is no conflict of interest. Ethical approval The study does not require ethics approval as we have not used human or animal subject. Author contributions GV and UG conceived the original idea and designed the research., GV developed mapping populations, performed phenotypic and genotypic analysis, and collected data. GV, AS, JS, and UG performed the data analysis. GV and UG prepared the manuscript with inputs from AS and JS. All authors have read and approved the manuscript. UG secured the funding for this research. Acknowledgments This work was supported by the Specialty Crop Block Grant Program, North Dakota Department of Agriculture (NOGA-22-235), USDA-National Institute of Food and Agriculture (NIFA) Hatch project ND02243, and USDA-NIFA Research Capacity Fund project 7006432. Data availability All data and research materials are available upon reasonable request from the corresponding author. References Andrews S (2010) A quality control tool for high throughput sequence data. 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Annu Rep Bean Improv Coop 39:25–31 Liebenberg MM, Pretorius ZA (2004) Inheritance of resistance to Uromyces appendiculatus in the South African dry bean cultivar Kranskop. S Afr J Plant Soil 21:245–250. https://doi.org/10.1080/02571862.2004.10635057 Lu P, Zhang G, Li J, Gong Z, Wang G, Dong L, Zhang H, Guo G, Su M, Wang K, Wang Y, Zhu K, Wu Q, Chen Y, Li M, Huang B, Li B, Li W, Dong L, Hou Y, Cui X, Fu H, Qiu D, Yuan C, Li H, Zhou J-M, Han G-Z, Chen Y, Liu Z (2025) A wheat tandem kinase and NLR pair confers resistance to multiple fungal pathogens. Science 387:1418–1424. https://doi.org/10.1126/science.adp5469 Luannfinke ML, Coyne DP, Steadman JR (1986) The inheritance and association of resistance to rust, common bacterial blight, plant habit and foliar abnormalities in Phaseolus vulgaris . L Euphytica 35:969–982. https://doi.org/10.1007/BF00028607 MacQueen AH, White JW, Lee R, Osorno JM, Schmutz J, Miklas PN, Myers J, McClean PE, Juenger TE (2020) Genetic associations in four decades of multienvironment trials reveal agronomic trait evolution in common bean. Genetics 215:267–284. https://doi.org/10.1534/genetics.120.303038 Magwene PM, Willis JH, Kelly JK (2011) The statistics of bulk segregant analysis using next generation sequencing. PLoS Comput Biol 7:e1002255. https://doi.org/10.1371/journal.pcbi.1002255 Mansfeld BN, Grumet R (2018) QTLseqr: An R Package for Bulk Segregant Analysis with Next-Generation Sequencing. Plant Genome 11(2). https://doi.org/10.3835/plantgenome2018.01.0006 McClean PE, Mamidi S, McConnell M, Chikara S, Lee R (2010) Synteny mapping between common bean and soybean reveals extensive blocks of shared loci. BMC Genomics 11:184. https://doi.org/10.1186/1471-2164-11-184 McKenna A, Hanna M, Banks E, Sivachenko A, Cibulskis K, Kernytsky A, Garimella K, Altshuler D, Gabriel S, Daly M, DePristo MA (2010) The Genome Analysis Toolkit: A MapReduce framework for analyzing next-generation DNA sequencing data. Genome Res 20:1297–1303. https://doi.org/10.1101/gr.107524.110 McMillan MS, Schwartz HF, Otto KL (2003) Sexual stage development of Uromyces appendiculatus and its potential use for disease resistance screening of Phaseolus vulgaris . Plant Dis 87:1133–1138 Meziadi C, Richard MS, Derquennes A, Thareau V, Blanchet S, Gratias A, Pflieger S, Geffroy V (2016) Development of molecular markers linked to disease resistance genes in common bean based on whole genome sequence. Plant Sci 242:351–357. https://doi.org/10.1016/j.plantsci.2015.09.006 Miklas PN, Kelly JD, Beebe SE, Blair MW (2006) Common bean breeding for resistance against biotic and abiotic stresses: from classical to MAS breeding. Euphytica 147:105–131. https://doi.org/10.1007/s10681-006-4600-5 Miklas PN, Soler-Garzón A, Valentini G, Pastor-Corrales MA (2023) Registration of ‘USDA Rattler’ pinto bean. J Plant Regist 17:271–279. https://doi.org/10.1002/plr2.20289 Miklas PN, Stone V, Daly MJ, Stavely JR, Steadman JR, Bassett MJ, Delorme R, Beaver JS (2000) Bacterial, fungal, and viral disease resistance loci mapped in a recombinant inbred common bean population (‘Dorado’/XAN 176). J Am Soc Hort Sci 125:476–481 Monclova-Santana C (2019) Population structure of Uromyces appendiculatus in North Dakota. Ph.D. thesis, North Dakota State University, Fargo, ND, USA Osorno JM, Vander Wal AJ, Posch J, Simons K, Grafton KF, Pasche JS, Valentini G, Pastor-Corrales MA (2021) A new black bean with resistance to bean rust: Registration of ‘ND Twilight’. J Plant Registrations 15:28–36 Pastor-Corrales MA (2002) Apparent vulnerability of certain rust-resistance gene combinations in common bean for the management of Uromyces appendiculatus . Ann Rep Bean Improv Coop 45:42–43 Pastor-Corrales MA (2003) Sources, genes for resistance, and pedigrees of 52 rust and mosaic resistant dry bean germplasm lines released by the USDA Beltsville Bean Project in collaboration with the Michigan, Nebraska, and North Dakota Agricultural Experiment Stations. Ann Rep Bean Improv Coop 46:235–241 Pastor-Corrales MA (2006) Diversity of the rust pathogen and common bean guides gene deployment for development of bean cultivars with durable rust resistance. Ann Rep Bean Improv Coop 49:51–52 Rasmussen JB, Grafton KF, Gross PL, Donohue CM (2002) Genetics of rust resistance in Compuesto Negro Chimaltenango (CNC). Ann Rep Bean Improv Coop 45:94–95 Richard MM, Pflieger S, Sévignac M, Thareau V, Blanchet S, Li Y, Jackson SA, Geffroy V (2014) Fine mapping of Co-x , an anthracnose resistance gene to a highly virulent strain of Colletotrichum lindemuthianum in common bean. Theor Appl Genet 127:1653–1666. https://doi.org/10.1007/s00122-014-2328-5 Robinson JT, Thorvaldsdóttir H, Winckler W, Guttman M, Lander ES, Getz G, Mesirov JP (2011) Integrative genomics viewer. Nat Biotechnol 29(1):24–26. https://doi.org/10.1038/nbt.1754 Schmutz J, McClean PE, Mamidi S, Wu GA, Cannon SB et al (2014) A reference genome for common bean and genome-wide analysis of dual domestications. Nat Genet 46:707–713. https://doi.org/10.1038/ng.3008 Schwartz HF, Pastor-Corrales MA (1989) Bean production problems in the tropics. CIAT, Cali, Colombia, p 654 Shin SH, Song Q, Cregan PB, Pastor-Corrales MA (2014) SSR DNA markers linked with broad-spectrum rust resistance in common bean discovered by bulk segregant analysis using a large set of SNP markers. Ann Rep Bean Improv Coop 57:187–188 Souza TLPO, Dessaune SN, Sanglard DA, Moreira, Barros EB (2011) Characterization of the rust resistance gene present in the common bean cultivar Ouro Negro, the main rust resistance source used in Brazil. Plant Pathol 60:839–845. https://doi.org/10.1111/j.1365-3059.2011.02456.x Stavely JR (1984) Pathogenic specialization in Uromyces phaseoli in the United States and rust resistance in beans. Plant Dis 68:95–99. https://doi.org/10.1094/PD-68-95 Stavely JR (1990) Genetics of rust resistance in Phaseolus vulgaris plant introduction PI 181996. Phytopathology 80:1056 Stavely JR, Freytag GF, Steadman JR, Schwartz HF (1983) The 1983 bean rust workshop. Ann Rep Bean Improv Coop 26:4–6 Stavely JR, Grafton KF (1989) Registration of Beldak-rust resistance – 1 and – 2 dry bean germplasm. Crop Sci 29(3):834–835 Stavely JR, Pastor-Corrales MA, Cali (1989) Colombia, 159–194 Stavely JR, Steadman JR, McMillan RT Jr (1989) New pathogenic variability in Uromyces appendiculatus in North America. Plant Dis 73:428–432. https://doi.org/10.1094/PD-73-0428 Steadman JR, Pastor-Corrales MA, Beaver JS (2002) An overview of the 3rd Bean Rust and 2nd Bean Common Bacterial Blight International Workshops, March 4–8, 2002, Pietermaritzburg, South Africa. Ann Rep Bean Improv Coop 45:120–125 Takagi H, Abe A, Yoshida K, Kosugi S, Natsume S, Mitsuoka C, Uemura A, Utsushi H, Tamiru M, Takuno S, Innan H, Cano LM, Kamoun S, Terauchi R (2013) QTL-seq: Rapid mapping of quantitative trait loci in rice by whole genome resequencing of DNA from two bulked populations. Plant J 74:174–183. https://doi.org/10.1111/tpj.12105 Valentini G, Gonçalves-Vidigal MC, Hurtado-Gonzales OP, de Lima Castro SA, Cregan PB, Song Q, Pastor-Corrales MA (2017) High-resolution mapping reveals linkage between genes in common bean cultivar Ouro Negro conferring resistance to the rust, anthracnose, and angular leaf spot diseases. Theor Appl Genet 130:1705–1722. https://doi.org/10.1007/s00122-017-2920-6 Valentini G, Xavier LFS, Hurtado-Gonzales OP, Song Q, Pastor-Corrales MA (2021) Mapping of a rust resistance locus in Andean common bean landrace G19833. Ann Rep Bean Improv Coop 64:13–14 Valentini G, Xavier LFS, Pastor-Corrales MA (2020) Mapping of a broad-spectrum rust resistance locus in Andean common bean PI 260418. APS Plant Health 2020 Annual Meeting (Online, USA) Van Ooijen JW (2006) JoinMap 4, Software for the Calculation of Genetic Linkage Maps in Experimental Populations. Kyazma B.V., Wageningen, Netherlands Vasimuddin Md, Misra S, Li H, Aluru S (2019) Efficient architecture-aware acceleration of BWA-MEM for multicore systems. In: 2019 IEEE International Parallel and Distributed Processing Symposium (IPDPS). pp. 314–324. https://doi.org/10.1109/IPDPS.2019.00041 Vaz Bisneta MV, Gonçalves-Vidigal MC (2020) Integration of anthracnose resistance loci and RLK and NBS-LRR-encoding genes in the Phaseolus vulgaris L. genome. Plant Physiol 60(6):2901–2918 Vlasova A, Capella-Gutiérrez S, Rendón-Anaya M et al (2016) Genome and transcriptome analysis of the Mesoamerican common bean and the role of gene duplications in establishing tissue and temporal specialization of genes. Genome Biol 17:32. https://doi.org/10.1186/s13059-016-0883-6 Voorrips RE (2002) MapChart: Software for the graphical presentation of linkage maps and QTLs. J Hered 93:77–78. https://doi.org/10.1093/jhered/93.1.77 Supplementary Files Supplementarymaterial.xlsx Table S1. Phenotype and genotype of 275 F 2 plants from the UI114 x CNC cross used for the genetic linkage analysis of the rust resistance gene in CNC Table S2. Statistics for the significant QTLs identified between resistant and susceptible bulks from the UI114 x CNC F2 population obtained via QTLseqR analysis. Table S3. PCR allelic competitive extension (PACE) markers designed for the region containing the rust resistance locus in the common bean cultivar CNC Table S4. Predicted gene models in the interval containing the rust resistance gene in CNC on Pv04 according to the reference genome UI111 Cite Share Download PDF Status: Posted Version 1 posted You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. 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Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {\"props\":{\"pageProps\":{\"initialData\":{\"identity\":\"rs-6572527\",\"acceptedTermsAndConditions\":true,\"allowDirectSubmit\":true,\"archivedVersions\":[],\"articleType\":\"Research Article\",\"associatedPublications\":[],\"authors\":[{\"id\":473160806,\"identity\":\"8e9416c6-30fc-47fb-bad5-05c04b91a8ba\",\"order_by\":0,\"name\":\"Giseli Valentini\",\"email\":\"\",\"orcid\":\"\",\"institution\":\"North Dakota State University\",\"correspondingAuthor\":false,\"prefix\":\"\",\"firstName\":\"Giseli\",\"middleName\":\"\",\"lastName\":\"Valentini\",\"suffix\":\"\"},{\"id\":473160807,\"identity\":\"cfcf5aba-5690-47de-8c26-2da9fb50a873\",\"order_by\":1,\"name\":\"Anuj Sharma\",\"email\":\"\",\"orcid\":\"\",\"institution\":\"University of Florida\",\"correspondingAuthor\":false,\"prefix\":\"\",\"firstName\":\"Anuj\",\"middleName\":\"\",\"lastName\":\"Sharma\",\"suffix\":\"\"},{\"id\":473160808,\"identity\":\"aa931ee7-2510-455c-a996-6cfed311b5b3\",\"order_by\":2,\"name\":\"Jatinder Singh\",\"email\":\"\",\"orcid\":\"\",\"institution\":\"North Dakota State University\",\"correspondingAuthor\":false,\"prefix\":\"\",\"firstName\":\"Jatinder\",\"middleName\":\"\",\"lastName\":\"Singh\",\"suffix\":\"\"},{\"id\":473160809,\"identity\":\"86300925-8d15-4c36-b44c-1007162d0e96\",\"order_by\":3,\"name\":\"Upinder Gill\",\"email\":\"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAAA6UlEQVRIiWNgGAWjYFACxgYGHhDNzNx44AMDM1hMgkgtjA0HZxCnBQh4oHoP8xCjxeB2c9uDNxX3GOTdgVpsd1jbGxxgPnibB5+WOwfbDeecKWYwPAzUknsmPXHDAbZka7xabiS2SfO2JTAYNoO0tB1OMDjAYyZNWMs/qBbLtsNAh/F/I0JLQwKDPDDEDjO2HWbccICHDa8WyTsH2yTnHEvgMQAFcm9beuLMw2zGlnPwaOG73f5M4k1Ngpx8/+GDD362WdvzHW9+eOMNHi2wKOAxOAATYcanHEkLg3wDIZWjYBSMglEwYgEAo0NOhYhjq0wAAAAASUVORK5CYII=\",\"orcid\":\"https://orcid.org/0000-0003-4572-5243\",\"institution\":\"North Dakota State University\",\"correspondingAuthor\":true,\"prefix\":\"\",\"firstName\":\"Upinder\",\"middleName\":\"\",\"lastName\":\"Gill\",\"suffix\":\"\"}],\"badges\":[],\"createdAt\":\"2025-05-01 14:32:27\",\"currentVersionCode\":1,\"declarations\":\"\",\"doi\":\"10.21203/rs.3.rs-6572527/v1\",\"doiUrl\":\"https://doi.org/10.21203/rs.3.rs-6572527/v1\",\"draftVersion\":[],\"editorialEvents\":[],\"editorialNote\":\"\",\"failedWorkflow\":false,\"files\":[{\"id\":85067535,\"identity\":\"a4a52250-794e-4a1c-bd5c-8adcdefaaa5b\",\"added_by\":\"auto\",\"created_at\":\"2025-06-20 15:14:26\",\"extension\":\"png\",\"order_by\":1,\"title\":\"Figure 1\",\"display\":\"\",\"copyAsset\":false,\"role\":\"figure\",\"size\":5471944,\"visible\":true,\"origin\":\"\",\"legend\":\"\\u003cp\\u003ePhenotypes of bean plants inoculated with races 13-2, 20-6, 31-1, and 20-3 of \\u003cem\\u003eU. appendiculatus\\u003c/em\\u003e. (a) Susceptible parental line UI114; (b) Resistant parental line CNC; (c) F\\u003csub\\u003e1\\u003c/sub\\u003e plant from the cross UI114 × CNC. Scheme of inoculation: cut leaf tip (left leaf): races 13-2 (top) and 20-6 (bottom); uncut leaf tip (right leaf): 20-3 (top) and 31-1 (bottom).\\u003c/p\\u003e\",\"description\":\"\",\"filename\":\"Fig1.png\",\"url\":\"https://assets-eu.researchsquare.com/files/rs-6572527/v1/660aeddf4ba48f8004f42fc0.png\"},{\"id\":85067532,\"identity\":\"71370f11-85d5-459e-b049-d19de8bbe3df\",\"added_by\":\"auto\",\"created_at\":\"2025-06-20 15:14:26\",\"extension\":\"png\",\"order_by\":2,\"title\":\"Figure 2\",\"display\":\"\",\"copyAsset\":false,\"role\":\"figure\",\"size\":2511531,\"visible\":true,\"origin\":\"\",\"legend\":\"\\u003cp\\u003eQTL mapping of rust resistance in CNC via NGS-BSA. a) SNP distribution on the 11 linkage groups performed using a 2 Mb sliding window; b) QTLseq analysis using Δ(SNP-index); c) G’ value (c); and d) −log10(p-value) methods.\\u003c/p\\u003e\",\"description\":\"\",\"filename\":\"Fig2.png\",\"url\":\"https://assets-eu.researchsquare.com/files/rs-6572527/v1/ec070fc091af85e8091fdda5.png\"},{\"id\":85067977,\"identity\":\"22830fb9-19a5-4e00-9d72-365d5780efbc\",\"added_by\":\"auto\",\"created_at\":\"2025-06-20 15:22:26\",\"extension\":\"png\",\"order_by\":3,\"title\":\"Figure 3\",\"display\":\"\",\"copyAsset\":false,\"role\":\"figure\",\"size\":10739,\"visible\":true,\"origin\":\"\",\"legend\":\"\\u003cp\\u003eGenetic mapping of the rust resistance gene in CNC on the common bean chromosome Pv04. Genetic map developed from 275 F\\u003csub\\u003e2\\u003c/sub\\u003e plants phenotyped with races 20-6 and 31-1 of the rust pathogen and genotyped with 11 PACE markers.\\u003c/p\\u003e\",\"description\":\"\",\"filename\":\"Fig3.png\",\"url\":\"https://assets-eu.researchsquare.com/files/rs-6572527/v1/e801d8fda1536a1ff92d533d.png\"},{\"id\":91725346,\"identity\":\"b9d04ffe-b490-4bc5-8c43-2d45c3815103\",\"added_by\":\"auto\",\"created_at\":\"2025-09-19 14:55:43\",\"extension\":\"pdf\",\"order_by\":0,\"title\":\"\",\"display\":\"\",\"copyAsset\":false,\"role\":\"manuscript-pdf\",\"size\":7740218,\"visible\":true,\"origin\":\"\",\"legend\":\"\",\"description\":\"\",\"filename\":\"manuscript.pdf\",\"url\":\"https://assets-eu.researchsquare.com/files/rs-6572527/v1/b44ecc85-f43d-4ffd-9fad-1ed9b4afe409.pdf\"},{\"id\":85067533,\"identity\":\"e80ca33f-7d40-4247-a1e4-5cc8b1799904\",\"added_by\":\"auto\",\"created_at\":\"2025-06-20 15:14:26\",\"extension\":\"xlsx\",\"order_by\":1,\"title\":\"\",\"display\":\"\",\"copyAsset\":false,\"role\":\"supplement\",\"size\":56510,\"visible\":true,\"origin\":\"\",\"legend\":\"\\u003cp\\u003eTable S1. Phenotype and genotype of 275 F\\u003csub\\u003e2\\u003c/sub\\u003e plants from the UI114 x CNC cross used for the genetic linkage analysis of the rust resistance gene in CNC\\u003c/p\\u003e\\n\\u003cp\\u003eTable S2. Statistics for the significant QTLs identified between resistant and susceptible bulks from the UI114 x CNC F2 population obtained via QTLseqR analysis.\\u003c/p\\u003e\\n\\u003cp\\u003eTable S3. PCR allelic competitive extension (PACE) markers designed for the region containing the rust resistance locus in the common bean cultivar CNC\\u003c/p\\u003e\\n\\u003cp\\u003eTable S4. Predicted gene models in the interval containing the rust resistance gene in CNC on Pv04 according to the reference genome UI111\\u003c/p\\u003e\",\"description\":\"\",\"filename\":\"Supplementarymaterial.xlsx\",\"url\":\"https://assets-eu.researchsquare.com/files/rs-6572527/v1/9eb9b7fb94d4b4dca28f2195.xlsx\"}],\"financialInterests\":\"\",\"formattedTitle\":\"Identification and genetic mapping of a rust resistance gene on chromosome Pv04 in the common bean cultivar CNC using bulked segregant sequencing\",\"fulltext\":[{\"header\":\"Key Message\",\"content\":\"\\u003cp\\u003eGenetic mapping of the rust resistance gene in the common bean cultivar Compuesto Negro Chimaltenango (CNC) using NGS-BSA.\\u003c/p\\u003e\"},{\"header\":\"Introduction\",\"content\":\"\\u003cp\\u003eCommon bean (\\u003cem\\u003ePhaseolus vulgaris\\u003c/em\\u003e L.) is a highly nutritious food and part of the daily diet of about 400\\u0026nbsp;million people in the tropics (CIAT \\u003cspan citationid=\\\"CR8\\\" class=\\\"CitationRef\\\"\\u003e2024\\u003c/span\\u003e). Especially in Latin America and Africa, beans play an important role as the primary source of protein for direct human consumption (Broughton et al. \\u003cspan citationid=\\\"CR4\\\" class=\\\"CitationRef\\\"\\u003e2003\\u003c/span\\u003e). Diseases are often the most severe constraint in the common bean production, negatively impacting the quality and yield. Rust, caused by the fungus \\u003cem\\u003eUromyces appendiculatus\\u003c/em\\u003e, is a disease with worldwide distribution, aligning with the distribution of its hosts. Although the disease occurs in many parts of the world, the yield losses are often more severe when common beans are cultivated under warm and humid conditions (Schwartz and Pastor-Corrales \\u003cspan citationid=\\\"CR39\\\" class=\\\"CitationRef\\\"\\u003e1989\\u003c/span\\u003e; Stavely and Pastor-Corrales 1989). The adoption of cultivars with genetic resistance is the most cost-effective and practical strategy to control rust (Miklas et al. \\u003cspan citationid=\\\"CR27\\\" class=\\\"CitationRef\\\"\\u003e2006\\u003c/span\\u003e). In the US, the introduction of rust resistance genes in the mid-1980s was crucial to reduce chemical applications primarily used to control rust in common bean (MacQueen et al. \\u003cspan citationid=\\\"CR20\\\" class=\\\"CitationRef\\\"\\u003e2020\\u003c/span\\u003e). Indeed, using disease-resistant cultivars lowers production costs while averting yield loss, and reduces the dependence on chemical control and the negative environmental impact caused by their application (Miklas et al. \\u003cspan citationid=\\\"CR27\\\" class=\\\"CitationRef\\\"\\u003e2006\\u003c/span\\u003e). However, the wide and complex virulence diversity of \\u003cem\\u003eU. appendiculatus\\u003c/em\\u003e, distributed into several races, hinders the development and durability of resistance in common bean cultivars (Stavely et al. \\u003cspan citationid=\\\"CR46\\\" class=\\\"CitationRef\\\"\\u003e1989\\u003c/span\\u003e). \\u003cem\\u003eU. appendiculatus\\u003c/em\\u003e is an autoecious fungus that can potentially undergo sexual reproduction to produce new virulent strains via recombination (McMillan et al. \\u003cspan citationid=\\\"CR25\\\" class=\\\"CitationRef\\\"\\u003e2003\\u003c/span\\u003e). The complexity of the \\u003cem\\u003eU. appendiculatus\\u003c/em\\u003e races coupled with the potential for the breakdown of monogenic resistances in bean cultivars necessitates the identification of additional resistance (\\u003cem\\u003eR\\u003c/em\\u003e) genes and their pyramiding in elite germplasm (Pastor-Corrales \\u003cspan citationid=\\\"CR32\\\" class=\\\"CitationRef\\\"\\u003e2002\\u003c/span\\u003e; \\u003cspan citationid=\\\"CR33\\\" class=\\\"CitationRef\\\"\\u003e2003\\u003c/span\\u003e; \\u003cspan citationid=\\\"CR34\\\" class=\\\"CitationRef\\\"\\u003e2006\\u003c/span\\u003e). The deployment of a combination of multiple \\u003cem\\u003eR\\u003c/em\\u003e genes in a single cultivar is an important strategy for obtaining effective and durable genetic resistance to rust (Stavely et al. \\u003cspan citationid=\\\"CR46\\\" class=\\\"CitationRef\\\"\\u003e1989\\u003c/span\\u003e; Pastor-Corrales \\u003cspan citationid=\\\"CR34\\\" class=\\\"CitationRef\\\"\\u003e2006\\u003c/span\\u003e).\\u003c/p\\u003e \\u003cp\\u003eGenes conditioning rust resistance are recognized by the Bean Improvement Cooperative (BIC) Committee and identified by the \\u003cem\\u003eUr-\\u003c/em\\u003esymbol (Kelly et al. \\u003cspan citationid=\\\"CR16\\\" class=\\\"CitationRef\\\"\\u003e1996\\u003c/span\\u003e). The \\u003cem\\u003eUr-3\\u003c/em\\u003e, \\u003cem\\u003eUr-5, Ur-7\\u003c/em\\u003e, \\u003cem\\u003eUr-11\\u003c/em\\u003e, and \\u003cem\\u003eUr-14\\u003c/em\\u003e are from the Middle American bean gene pool (Augustin et al. \\u003cspan citationid=\\\"CR2\\\" class=\\\"CitationRef\\\"\\u003e1972\\u003c/span\\u003e; Ballantyne \\u003cspan citationid=\\\"CR3\\\" class=\\\"CitationRef\\\"\\u003e1978\\u003c/span\\u003e; Stavely \\u003cspan citationid=\\\"CR42\\\" class=\\\"CitationRef\\\"\\u003e1984\\u003c/span\\u003e; Stavely \\u003cspan citationid=\\\"CR43\\\" class=\\\"CitationRef\\\"\\u003e1990\\u003c/span\\u003e; Souza et al. \\u003cspan citationid=\\\"CR41\\\" class=\\\"CitationRef\\\"\\u003e2011\\u003c/span\\u003e), and \\u003cem\\u003eUr-4, Ur-6, Ur-9, Ur-12\\u003c/em\\u003e, and \\u003cem\\u003eUr-13\\u003c/em\\u003e are from the Andean gene pool (Ballantyne \\u003cspan citationid=\\\"CR3\\\" class=\\\"CitationRef\\\"\\u003e1978\\u003c/span\\u003e; Luannfinke et al. \\u003cspan citationid=\\\"CR19\\\" class=\\\"CitationRef\\\"\\u003e1986\\u003c/span\\u003e; Jung et al. \\u003cspan citationid=\\\"CR14\\\" class=\\\"CitationRef\\\"\\u003e1998\\u003c/span\\u003e; Liebenberg and Pretorius \\u003cspan citationid=\\\"CR17\\\" class=\\\"CitationRef\\\"\\u003e2004\\u003c/span\\u003e). These genes have been mapped on six chromosomes of common bean (Pv01, Pv04, Pv06, Pv07, Pv08, and Pv11). All \\u003cem\\u003eUr-\\u003c/em\\u003e genes, but \\u003cem\\u003eUr-14\\u003c/em\\u003e, were mapped in the 12 differential cultivars for \\u003cem\\u003eU. appendiculatus\\u003c/em\\u003e race characterization. This set was proposed by Steadman et al. (\\u003cspan citationid=\\\"CR48\\\" class=\\\"CitationRef\\\"\\u003e2002\\u003c/span\\u003e) and comprises six Andean and six Middle American cultivars. Compuesto Negro Chimaltenango (CNC), which is one of 12 differentials and presents broad resistance to multiple rust races, still needs to be genetically characterized in detail.\\u003c/p\\u003e \\u003cp\\u003eCNC is a black bean cultivar from the Middle American gene pool and originally from Guatemala. This cultivar was included in the first set of differential lines developed by Stavely et al. (\\u003cspan citationid=\\\"CR44\\\" class=\\\"CitationRef\\\"\\u003e1983\\u003c/span\\u003e), and was retained as part of the updated set of rust differential lines proposed in 2002 (Steadman et al. \\u003cspan citationid=\\\"CR48\\\" class=\\\"CitationRef\\\"\\u003e2002\\u003c/span\\u003e). The resistance in CNC is effective against most races of bean rust (Stavely \\u003cspan citationid=\\\"CR42\\\" class=\\\"CitationRef\\\"\\u003e1984\\u003c/span\\u003e; Stavely et al. \\u003cspan citationid=\\\"CR46\\\" class=\\\"CitationRef\\\"\\u003e1989\\u003c/span\\u003e). Despite its rust resistance, CNC has not been extensively used in cultivar development, and the genetic basis of its resistance remains poorly understood. Previous reports indicate CNC possesses a single dominant gene that conditions rust resistance against race 49 of \\u003cem\\u003eU. appendiculatus\\u003c/em\\u003e (Rasmussen et al. \\u003cspan citationid=\\\"CR35\\\" class=\\\"CitationRef\\\"\\u003e2002\\u003c/span\\u003e). Although other races were not tested on CNC, the presence of multiple resistance genes in the line has been suggested (Rasmussen et al. \\u003cspan citationid=\\\"CR35\\\" class=\\\"CitationRef\\\"\\u003e2002\\u003c/span\\u003e). Given the importance of highly effective rust resistance in CNC, we present its genetic characterization, mapping, and the development of molecular markers to facilitate its potential use in future bean cultivars.\\u003c/p\\u003e\"},{\"header\":\"Material and Methods\",\"content\":\"\\u003cdiv id=\\\"Sec3\\\" class=\\\"Section2\\\"\\u003e \\u003ch2\\u003ePlant material\\u003c/h2\\u003e \\u003cp\\u003eThe F\\u003csub\\u003e1\\u003c/sub\\u003e and F\\u003csub\\u003e2\\u003c/sub\\u003e populations were developed from the cross between UI114 and CNC at North Dakota State University, Fargo, ND. CNC is resistant to most of the known races of \\u003cem\\u003eU. appendiculatus\\u003c/em\\u003e, while UI114, a pinto bean cultivar, is highly susceptible. The dominant violet flower trait, inherited from the male parent CNC, was observed in the F\\u003csub\\u003e1\\u003c/sub\\u003e plants, confirming that they were hybrids. A total of 275 F\\u003csub\\u003e2\\u003c/sub\\u003e plants were obtained by F\\u003csub\\u003e1\\u003c/sub\\u003e self-pollination and used for the inheritance test.\\u003c/p\\u003e \\u003c/div\\u003e\\n\\u003ch3\\u003eRust phenotyping\\u003c/h3\\u003e\\n\\u003cp\\u003eThe races used in this study were obtained from the collection maintained at North Dakota State University. Six F\\u003csub\\u003e1\\u003c/sub\\u003e plants from the cross UI114 \\u0026times; CNC were inoculated with races 13\\u0026thinsp;\\u0026minus;\\u0026thinsp;2, 20\\u0026thinsp;\\u0026minus;\\u0026thinsp;6, 31\\u0026thinsp;\\u0026minus;\\u0026thinsp;1, and 20\\u0026thinsp;\\u0026minus;\\u0026thinsp;3 of \\u003cem\\u003eU. appendiculatus\\u003c/em\\u003e. Later, races 20\\u0026thinsp;\\u0026minus;\\u0026thinsp;6 and 31\\u0026thinsp;\\u0026minus;\\u0026thinsp;1 of \\u003cem\\u003eU. appendiculatus\\u003c/em\\u003e were selected to study the genetic inheritance on the F\\u003csub\\u003e2\\u003c/sub\\u003e population.\\u003c/p\\u003e \\u003cp\\u003eSeeds of the F\\u003csub\\u003e1\\u003c/sub\\u003e and F\\u003csub\\u003e2\\u003c/sub\\u003e populations were planted in 12.7-cm diameter pots containing Pro-mix BX general purpose growing mix (Premier Tech) and maintained under greenhouse conditions at 20\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;5\\u0026deg;C. Inoculations of \\u003cem\\u003eU. appendiculatus\\u003c/em\\u003e were performed nine days after planting when the primary leaves were about 2/3 expanded. Parental lines and the cultivars Aurora (\\u003cem\\u003eUr-3\\u003c/em\\u003e), Early Gallatin (\\u003cem\\u003eUr-4\\u003c/em\\u003e), Mexico 309 (\\u003cem\\u003eUr-5\\u003c/em\\u003e), Golden Gate Wax (\\u003cem\\u003eUr-6\\u003c/em\\u003e), and ND Falcon (\\u003cem\\u003eUr-11\\u003c/em\\u003e) were used as internal controls.\\u003c/p\\u003e \\u003cp\\u003eThe inoculum solution was prepared by suspending urediniospores of \\u003cem\\u003eU. appendiculatus\\u003c/em\\u003e in distilled water containing Tween 20 (0.01%) to obtain a concentration of 2 \\u0026times; 10\\u003csup\\u003e4\\u003c/sup\\u003e uredospores mL\\u003csup\\u003e\\u0026minus;\\u0026thinsp;1\\u003c/sup\\u003e. The F\\u003csub\\u003e1\\u003c/sub\\u003e and F\\u003csub\\u003e2\\u003c/sub\\u003e populations were simultaneously inoculated with four and two races of \\u003cem\\u003eU. appendiculatus\\u003c/em\\u003e, respectively, on the abaxial side of the primary leaves using a cotton swab containing the spore solution, avoiding contamination between races.\\u003c/p\\u003e \\u003cp\\u003eAfter inoculation, plants were transferred to a mist chamber (20\\u0026thinsp;\\u0026plusmn;\\u0026thinsp;1\\u0026deg;C, relative humidity\\u0026thinsp;\\u0026gt;\\u0026thinsp;95%) for 18 hours, under darkness. After this period, plants were relocated to the greenhouse and kept for approximately 10\\u0026ndash;12 days, until disease symptoms appeared. Disease evaluation were performed using a 1\\u0026ndash;6 scale proposed by Stavely et al. (\\u003cspan citationid=\\\"CR44\\\" class=\\\"CitationRef\\\"\\u003e1983\\u003c/span\\u003e), where 1\\u0026thinsp;=\\u0026thinsp;without any visible rust reactions; 2\\u0026thinsp;=\\u0026thinsp;necrotic spots without sporulation (hypersensitive reaction or HR); 2\\u0026thinsp;+\\u0026thinsp;=\\u0026thinsp;necrotic spots 300\\u0026ndash;1000 \\u0026micro;m (1 mm) in diameter; 2++ = necrotic spots 1\\u0026ndash;3 mm in diameter; 2+++ = necrotic spots larger than 3 mm in diameter; 3\\u0026thinsp;=\\u0026thinsp;tiny sporulating uredinia (rust pustules) less than 300 \\u0026micro;m in diameter; 4\\u0026thinsp;=\\u0026thinsp;sporulating uredinia 300\\u0026ndash;500 \\u0026micro;m in diameter; 5\\u0026thinsp;=\\u0026thinsp;sporulating uredinia 500\\u0026ndash;800 \\u0026micro;m in diameter; and 6\\u0026thinsp;=\\u0026thinsp;sporulating uredinia larger than 800 \\u0026micro;m in diameter. Plants with grades 1, 2 and 3 were classified as resistant, whereas plants with grades 4, 5, and 6 were considered susceptible. Evaluations including more than one grade were recorded and separated by \\u0026ldquo;,\\u0026rdquo; with the most predominant type of reactions listed first. The chi-square (χ\\u003csup\\u003e2\\u003c/sup\\u003e) test was performed to study the genetic segregation (resistant/susceptible) ratios.\\u003c/p\\u003e\\n\\u003ch3\\u003eNext generation sequencing based bulked segregant analysis (NGS-BSA)\\u003c/h3\\u003e\\n\\u003cp\\u003eBased on the disease phenotype of the F\\u003csub\\u003e2\\u003c/sub\\u003e population, plants were selected for NGS-BSA. Two bulks were developed: a resistant bulk containing 18 F\\u003csub\\u003e2\\u003c/sub\\u003e plants resistant to races 20\\u0026thinsp;\\u0026minus;\\u0026thinsp;6 and 31\\u0026thinsp;\\u0026minus;\\u0026thinsp;1 and a susceptible bulk containing 18 F\\u003csub\\u003e2\\u003c/sub\\u003e plants susceptible to races 20\\u0026thinsp;\\u0026minus;\\u0026thinsp;6 and 31\\u0026thinsp;\\u0026minus;\\u0026thinsp;1. Young leaves were collected from each plant and lyophilized for further DNA extraction using the OmniPrep DNA extraction kit (G-Biosciences, St. Louis, MO) following the manufacture\\u0026rsquo;s protocol with a few modifications. DNA purity and concentration were verified using a NanoDrop (Thermo Fisher Scientific, Waltham, MA). For the two bulks, DNA of plants were bulked separately in equimolar concentrations based on the NanoDrop readings. The paired-end DNA sequencing data (2\\u0026times;150 bp) from bulks were generated using Illumina NovaSeq X Plus by outsourcing to Novogene (\\u003cspan class=\\\"ExternalRef\\\"\\u003e\\u003cspan class=\\\"RefSource\\\"\\u003ewww.novogene.com\\u003c/a\\u003e\\u003c/span\\u003e\\u003cspan address=\\\"http://www.novogene.com\\\" targettype=\\\"URL\\\" class=\\\"RefTarget\\\"\\u003e\\u003c/span\\u003e\\u003c/span\\u003e).\\u003c/p\\u003e \\u003cp\\u003eFor data analysis \\u003cem\\u003eviz.\\u003c/em\\u003e, quality check, adapter trimming, read mapping and variant calling, Bulk2SNPs pipeline (\\u003cspan class=\\\"ExternalRef\\\"\\u003e\\u003cspan class=\\\"RefSource\\\"\\u003ehttps://github.com/NDSUrustlab/Bulk2SNPs\\u003c/span\\u003e\\u003cspan address=\\\"https://github.com/NDSUrustlab/Bulk2SNPs\\\" targettype=\\\"URL\\\" class=\\\"RefTarget\\\"\\u003e\\u003c/span\\u003e\\u003c/span\\u003e) was used. This pipeline uses a set of bioinformatics software, such as fastqc (Andrews \\u003cspan citationid=\\\"CR1\\\" class=\\\"CitationRef\\\"\\u003e2010\\u003c/span\\u003e), fastp (Chen \\u003cspan citationid=\\\"CR7\\\" class=\\\"CitationRef\\\"\\u003e2023\\u003c/span\\u003e), bwa-mem2 (Vasimuddin et al. 2019), and GATK (McKenna et al. \\u003cspan citationid=\\\"CR24\\\" class=\\\"CitationRef\\\"\\u003e2010\\u003c/span\\u003e) for analyzing raw sequencing data from two bulks and output a Single Nucleotide Polymorphism (SNP) table file for downstream QTL analysis. The reference genome \\u003cem\\u003eP. vulgaris\\u003c/em\\u003e UI111 v1.1 (\\u003cspan class=\\\"ExternalRef\\\"\\u003e\\u003cspan class=\\\"RefSource\\\"\\u003ehttps://phytozome-next.jgi.doe.gov/\\u003c/span\\u003e\\u003cspan address=\\\"https://phytozome-next.jgi.doe.gov/\\\" targettype=\\\"URL\\\" class=\\\"RefTarget\\\"\\u003e\\u003c/span\\u003e\\u003c/span\\u003e) was used for mapping the sequence reads for each bulk. For QTL analysis, we used an R package, QTLseqr (Mansfeld and Grumet \\u003cspan citationid=\\\"CR22\\\" class=\\\"CitationRef\\\"\\u003e2018\\u003c/span\\u003e) available at \\u003cspan class=\\\"ExternalRef\\\"\\u003e\\u003cspan class=\\\"RefSource\\\"\\u003ehttps://github.com/bmansfeld/QTLseqr\\u003c/span\\u003e\\u003cspan address=\\\"https://github.com/bmansfeld/QTLseqr\\\" targettype=\\\"URL\\\" class=\\\"RefTarget\\\"\\u003e\\u003c/span\\u003e\\u003c/span\\u003e. QTLseqr implements the bulked segregant analysis via G\\u0026rsquo; and Δ (SNP-index), described by Magwene et al. (\\u003cspan citationid=\\\"CR21\\\" class=\\\"CitationRef\\\"\\u003e2011\\u003c/span\\u003e) and Takagi et al. (\\u003cspan citationid=\\\"CR49\\\" class=\\\"CitationRef\\\"\\u003e2013\\u003c/span\\u003e), respectively. Low-confidence SNPs were filtered out based on the reference allele frequency (refAlleleFreq\\u0026thinsp;=\\u0026thinsp;0.05), and minimum and maximum read depth (minTotalDepth\\u0026thinsp;=\\u0026thinsp;40 and maxTotalDepth\\u0026thinsp;=\\u0026thinsp;400). Identification of candidate QTL regions was performed using a 2 Mb sliding window, whereby the confidence intervals for the ΔSNP-indices was determined using 10,000 simulations.\\u003c/p\\u003e\\n\\u003ch3\\u003eMarker development and genetic mapping\\u003c/h3\\u003e\\n\\u003cp\\u003eIndividual F\\u003csub\\u003e2\\u003c/sub\\u003e plants were genotyped to confirm marker-trait association. The targeted genomic region containing the gene of interest identified by NGS-BSA was examined for SNPs and used for the development of PCR allelic competitive extension (PACE) markers (3CR Bioscience, Essex, UK). Alignment sequences were visualized in the Integrative Genomics Viewer (Robinson et al. \\u003cspan citationid=\\\"CR37\\\" class=\\\"CitationRef\\\"\\u003e2011\\u003c/span\\u003e) for the manual selection of SNPs. Variants showing A/T⇄C/G substitutions that did not have another variant within the approximately 100 bp flanking region were considered for marker development. Using the reference genome, the selected sequences were retrieved and submitted to 3CR Bioscience for primer design.\\u003c/p\\u003e \\u003cp\\u003eFor genotyping, genomic DNA from F\\u003csub\\u003e2\\u003c/sub\\u003e plants was extracted using the protocol described by Kaur et al. (\\u003cspan citationid=\\\"CR15\\\" class=\\\"CitationRef\\\"\\u003e2023\\u003c/span\\u003e). The polymerase chain reaction (PCR) for PACE primers was performed using 2.5 \\u0026micro;l of premade PACE\\u0026reg; genotyping master mix (3CR Bioscience, Essex, UK), 0.07 \\u0026micro;l of primers mix (Sigma-Aldrich, St. Louis, MO), and 20\\u0026ndash;40 ng of genomic DNA, for a 5 \\u0026micro;l reaction. PCR reactions were performed in a CFX Opus 384 Real Time PCR System (Bio-Rad, Hercules, CA) with the following parameters:15 min at 94 \\u003csup\\u003eo\\u003c/sup\\u003eC, 10 cycles of 20 sec at 94 \\u003csup\\u003eo\\u003c/sup\\u003eC and 60 sec at 57 \\u003csup\\u003eo\\u003c/sup\\u003eC, and 30 cycles of 20 sec at 94\\u003csup\\u003eo\\u003c/sup\\u003eC and 60 sec at 57\\u003csup\\u003eo\\u003c/sup\\u003eC. PCR results were analyzed in the Bio-Rad CFX Manager 3.1 (Bio-Rad, Hercules, CA).\\u003c/p\\u003e \\u003cp\\u003eGenetic mapping of the rust resistance locus in CNC using PACE markers was performed with JoinMap 4.1 (Van Ooijen \\u003cspan citationid=\\\"CR53\\\" class=\\\"CitationRef\\\"\\u003e2006\\u003c/span\\u003e). Default settings for the Maximum Likelihood Mapping algorithm were used to define the linkage order and distances (cM). The genetic distances were later used to graphically design the linkage groups using MapChart 2.3 (Voorrips \\u003cspan citationid=\\\"CR57\\\" class=\\\"CitationRef\\\"\\u003e2002\\u003c/span\\u003e).\\u003c/p\\u003e\"},{\"header\":\"Results\",\"content\":\"\\u003cdiv id=\\\"Sec8\\\" class=\\\"Section2\\\"\\u003e \\u003ch2\\u003eGenetic segregation analysis of the F\\u003csub\\u003e2\\u003c/sub\\u003e mapping population\\u003c/h2\\u003e \\u003cp\\u003eTo investigate the genetics of rust resistance in CNC, we performed inheritance analyses, bulked segregant sequencing, and genetic mapping. For the inheritance study, we crossed the susceptible cultivar UI114 with the resistant CNC line. CNC displayed a highly resistant reaction when inoculated with races 13\\u0026thinsp;\\u0026minus;\\u0026thinsp;2, 20\\u0026thinsp;\\u0026minus;\\u0026thinsp;6, 31\\u0026thinsp;\\u0026minus;\\u0026thinsp;1, and 20\\u0026thinsp;\\u0026minus;\\u0026thinsp;3, characterized by small pustules and hypersensitive reaction. In contrast, UI114 exhibited a classical susceptible reaction characterized by large, sporulating pustules to all tested rust races. The F\\u003csub\\u003e1\\u003c/sub\\u003e plants from the cross UI114 \\u0026times; CNC exhibited a resistant reaction to races 13\\u0026thinsp;\\u0026minus;\\u0026thinsp;2, 20\\u0026thinsp;\\u0026minus;\\u0026thinsp;6, 31\\u0026thinsp;\\u0026minus;\\u0026thinsp;1, and 20\\u0026thinsp;\\u0026minus;\\u0026thinsp;3, which was identical to that of the resistant parental line CNC, suggesting that the rust resistance in CNC is a dominant trait (Fig.\\u0026nbsp;\\u003cspan refid=\\\"Fig1\\\" class=\\\"InternalRef\\\"\\u003e1\\u003c/span\\u003e).\\u003c/p\\u003e \\u003cp\\u003e \\u003c/p\\u003e \\u003cp\\u003ePhenotypic evaluations of 275 F\\u003csub\\u003e2\\u003c/sub\\u003e individuals indicated that the disease reactions to races 20\\u0026thinsp;\\u0026minus;\\u0026thinsp;6 and 31\\u0026thinsp;\\u0026minus;\\u0026thinsp;1 co-segregated, meaning that when an F\\u003csub\\u003e2\\u003c/sub\\u003e plant was resistant to race 20\\u0026thinsp;\\u0026minus;\\u0026thinsp;6, it was also resistant to race 31\\u0026thinsp;\\u0026minus;\\u0026thinsp;1 and vice versa. We observed the segregation of 207 resistant and 68 susceptible plants, fitting a ratio of 3 resistant to 1 susceptible (χ\\u0026sup2; = 0.011, P value\\u0026thinsp;=\\u0026thinsp;0.9168), confirming that the rust resistance in CNC to races 20\\u0026thinsp;\\u0026minus;\\u0026thinsp;6 and 31\\u0026thinsp;\\u0026minus;\\u0026thinsp;1 is conferred by a single dominant gene (Table \\u003cspan refid=\\\"MOESM1\\\" class=\\\"InternalRef\\\"\\u003eS1\\u003c/span\\u003e, Table\\u0026nbsp;\\u003cspan refid=\\\"Tab1\\\" class=\\\"InternalRef\\\"\\u003e1\\u003c/span\\u003e).\\u003c/p\\u003e \\u003cp\\u003e \\u003cdiv class=\\\"gridtable\\\"\\u003e\\u003ctable float=\\\"Yes\\\" id=\\\"Tab1\\\" border=\\\"1\\\"\\u003e \\u003ccaption language=\\\"En\\\"\\u003e \\u003cdiv class=\\\"CaptionNumber\\\"\\u003eTable 1\\u003c/div\\u003e \\u003cdiv class=\\\"CaptionContent\\\"\\u003e \\u003cp\\u003eObserved and expected reaction for the F\\u003csub\\u003e1\\u003c/sub\\u003e and F\\u003csub\\u003e2\\u003c/sub\\u003e populations from the cross UI114 \\u0026times; CNC inoculated with different races of \\u003cem\\u003eU. appendiculatus\\u003c/em\\u003e\\u003c/p\\u003e \\u003c/div\\u003e \\u003c/caption\\u003e \\u003ccolgroup cols=\\\"7\\\"\\u003e \\u003cdiv align=\\\"left\\\" class=\\\"colspec\\\" colname=\\\"c1\\\" colnum=\\\"1\\\"\\u003e\\u003c/div\\u003e \\u003cdiv align=\\\"left\\\" class=\\\"colspec\\\" colname=\\\"c2\\\" colnum=\\\"2\\\"\\u003e\\u003c/div\\u003e \\u003cdiv align=\\\"left\\\" class=\\\"colspec\\\" colname=\\\"c3\\\" colnum=\\\"3\\\"\\u003e\\u003c/div\\u003e \\u003cdiv align=\\\"left\\\" class=\\\"colspec\\\" colname=\\\"c4\\\" colnum=\\\"4\\\"\\u003e\\u003c/div\\u003e \\u003cdiv align=\\\"left\\\" class=\\\"colspec\\\" colname=\\\"c5\\\" colnum=\\\"5\\\"\\u003e\\u003c/div\\u003e \\u003cdiv align=\\\"char\\\" char=\\\".\\\" class=\\\"colspec\\\" colname=\\\"c6\\\" colnum=\\\"6\\\"\\u003e\\u003c/div\\u003e \\u003cdiv align=\\\"char\\\" char=\\\".\\\" class=\\\"colspec\\\" colname=\\\"c7\\\" colnum=\\\"7\\\"\\u003e\\u003c/div\\u003e \\u003cthead\\u003e \\u003ctr\\u003e \\u003cth align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003eParent/Cross\\u003c/p\\u003e \\u003c/th\\u003e \\u003cth align=\\\"left\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003eRaces\\u003c/p\\u003e \\u003c/th\\u003e \\u003cth align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003eGeneration\\u003c/p\\u003e \\u003c/th\\u003e \\u003cth align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003eObserved ratio (R:S)\\u003c/p\\u003e \\u003c/th\\u003e \\u003cth align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003eExpected ratio (R:S)\\u003c/p\\u003e \\u003c/th\\u003e \\u003cth align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003eχ\\u003csup\\u003e2\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/th\\u003e \\u003cth align=\\\"left\\\" colname=\\\"c7\\\"\\u003e \\u003cp\\u003eP value\\u003c/p\\u003e \\u003c/th\\u003e \\u003c/tr\\u003e \\u003c/thead\\u003e \\u003ctbody\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003eUI114\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"char\\\" char=\\\"\\u0026minus;\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e13\\u0026thinsp;\\u0026minus;\\u0026thinsp;2, 20\\u0026thinsp;\\u0026minus;\\u0026thinsp;6, 31\\u0026thinsp;\\u0026minus;\\u0026thinsp;1, 20\\u0026thinsp;\\u0026minus;\\u0026thinsp;3\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003eSP\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e0:16\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e\\u0026nbsp;\\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e\\u0026nbsp;\\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c7\\\"\\u003e\\u0026nbsp;\\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003eCNC\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"char\\\" char=\\\"\\u0026minus;\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e13\\u0026thinsp;\\u0026minus;\\u0026thinsp;2, 20\\u0026thinsp;\\u0026minus;\\u0026thinsp;6, 31\\u0026thinsp;\\u0026minus;\\u0026thinsp;1, 20\\u0026thinsp;\\u0026minus;\\u0026thinsp;3\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003eRP\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e16:0\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e\\u0026nbsp;\\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e\\u0026nbsp;\\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c7\\\"\\u003e\\u0026nbsp;\\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003eUI114 \\u0026times; CNC\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"char\\\" char=\\\"\\u0026minus;\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e13\\u0026thinsp;\\u0026minus;\\u0026thinsp;2, 20\\u0026thinsp;\\u0026minus;\\u0026thinsp;6, 31\\u0026thinsp;\\u0026minus;\\u0026thinsp;1, 20\\u0026thinsp;\\u0026minus;\\u0026thinsp;3\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003eF\\u003csub\\u003e1\\u003c/sub\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e6:0\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e\\u0026nbsp;\\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e\\u0026nbsp;\\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c7\\\"\\u003e\\u0026nbsp;\\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003eUI114 \\u0026times; CNC\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"char\\\" char=\\\"\\u0026minus;\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e20\\u0026thinsp;\\u0026minus;\\u0026thinsp;6, 31\\u0026thinsp;\\u0026minus;\\u0026thinsp;1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003eF\\u003csub\\u003e2\\u003c/sub\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e207:68\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003e3:1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"char\\\" char=\\\".\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003e0.011\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"char\\\" char=\\\".\\\" colname=\\\"c7\\\"\\u003e \\u003cp\\u003e0.9168\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003c/tbody\\u003e \\u003c/colgroup\\u003e \\u003ctfoot\\u003e \\u003ctr\\u003e\\u003ctd colspan=\\\"7\\\"\\u003eRP, resistant parent; SP, susceptible parent; R, resistant; S, susceptible\\u003c/td\\u003e\\u003c/tr\\u003e \\u003c/tfoot\\u003e \\u003c/table\\u003e\\u003c/div\\u003e \\u003c/p\\u003e \\u003c/div\\u003e\\n\\u003ch3\\u003eIdentification of genomic regions associated with resistance using NGS-BSA\\u003c/h3\\u003e\\n\\u003cp\\u003eTo identify the genomic region(s) involved in rust resistance, the bulked segregant analysis was performed using whole genome sequencing. Two bulks representing 18 highly resistant (R) and 18 highly susceptible (S) F\\u003csub\\u003e2\\u003c/sub\\u003e plants were generated by pooling equimolar concentrations of genomic DNA of each plant for whole genome sequencing. A total of 19.3 and 18.8 GB of Illumina sequence data were generated for the R and S bulks, respectively, representing an average coverage of 34X for the R bulk and 33X for the S bulk of the assembled common bean genome. For the R bulk, 137,938,390 reads were generated, of which 136,499,352 were mapped to the UI111 v1.1 reference genome; for the susceptible bulk, 134,460,710 reads were generated, of which 132,993,492 were mapped to the reference.\\u003c/p\\u003e \\u003cp\\u003eA comparison of the R and S bulks to the reference genome identified 1,897,742 SNPs. After filtering, 1,421,905 high-quality SNPs were retained for QTL mapping using QTLseqr (Mansfeld and Grumet, \\u003cspan citationid=\\\"CR22\\\" class=\\\"CitationRef\\\"\\u003e2018\\u003c/span\\u003e). SNPs were more frequently distributed in euchromatic regions, with a higher number observed on chromosomes Pv04, Pv10, and Pv11 (Fig.\\u0026nbsp;\\u003cspan refid=\\\"Fig2\\\" class=\\\"InternalRef\\\"\\u003e2\\u003c/span\\u003e). The QTL-seq analysis, based on the delta SNP index and the G\\u0026rsquo; statistics, consistently showed one major peak on chromosome Pv04 between 4,766 and 10,983,713 bp, suggesting the strong co-segregation between the rust phenotype and genotype in this region (Fig.\\u0026nbsp;\\u003cspan refid=\\\"Fig2\\\" class=\\\"InternalRef\\\"\\u003e2\\u003c/span\\u003e, Table S2). Four apparent minor peaks on chromosome 4 and a peak on chromosome 8 were also observed. These minor peaks were significant based on G\\u0026rsquo; statistics only. Therefore, subsequent validation efforts focused on delimiting the resistance locus within the subtelomeric region of Pv04, where the major peak was observed.\\u003c/p\\u003e \\u003cp\\u003e \\u003c/p\\u003e\\n\\u003ch3\\u003eGenetic mapping of the rust resistance gene in CNC\\u003c/h3\\u003e\\n\\u003cp\\u003eTo further validate the genomic region identified by NGS-BSA on Pv04, the SNPs within the region were identified to develop PACE markers for genotyping on the F\\u003csub\\u003e2\\u003c/sub\\u003e population. A total of 11 PACE markers were developed between 415,751 and 1,874,106 bp and genotyped on 275 F\\u003csub\\u003e2\\u003c/sub\\u003e plants from UI114 \\u0026times; CNC (Table S3). The rust resistance locus in CNC was linked at 0.0 cM to markers Pv859224, Pv1178763, and Pv1243509, and positioned 0.8 cM downstream of marker Pv696542 and 0.2 cM upstream of marker Pv1350344 (Fig.\\u0026nbsp;\\u003cspan refid=\\\"Fig3\\\" class=\\\"InternalRef\\\"\\u003e3\\u003c/span\\u003e). The resistance locus was located between 696,542 and 1,350,344 bp, spanning a physical genomic interval of 653.8 Kb (Table\\u0026nbsp;\\u003cspan refid=\\\"Tab2\\\" class=\\\"InternalRef\\\"\\u003e2\\u003c/span\\u003e).\\u003c/p\\u003e \\u003cp\\u003e \\u003c/p\\u003e \\u003cp\\u003e \\u003cdiv class=\\\"gridtable\\\"\\u003e\\u003ctable float=\\\"Yes\\\" id=\\\"Tab2\\\" border=\\\"1\\\"\\u003e \\u003ccaption language=\\\"En\\\"\\u003e \\u003cdiv class=\\\"CaptionNumber\\\"\\u003eTable 2\\u003c/div\\u003e \\u003cdiv class=\\\"CaptionContent\\\"\\u003e \\u003cp\\u003eReaction to \\u003cem\\u003eUromyces appendiculatus\\u003c/em\\u003e and genotypes at eleven SNP loci of F\\u003csub\\u003e2\\u003c/sub\\u003e plants from the cross UI114 \\u0026times; CNC\\u003c/p\\u003e \\u003c/div\\u003e \\u003c/caption\\u003e \\u003ccolgroup cols=\\\"14\\\"\\u003e \\u003cdiv align=\\\"left\\\" class=\\\"colspec\\\" colname=\\\"c1\\\" colnum=\\\"1\\\"\\u003e\\u003c/div\\u003e \\u003cdiv align=\\\"left\\\" class=\\\"colspec\\\" colname=\\\"c2\\\" colnum=\\\"2\\\"\\u003e\\u003c/div\\u003e \\u003cdiv align=\\\"left\\\" class=\\\"colspec\\\" colname=\\\"c3\\\" colnum=\\\"3\\\"\\u003e\\u003c/div\\u003e \\u003cdiv align=\\\"left\\\" class=\\\"colspec\\\" colname=\\\"c4\\\" colnum=\\\"4\\\"\\u003e\\u003c/div\\u003e \\u003cdiv align=\\\"left\\\" class=\\\"colspec\\\" colname=\\\"c5\\\" colnum=\\\"5\\\"\\u003e\\u003c/div\\u003e \\u003cdiv align=\\\"left\\\" class=\\\"colspec\\\" colname=\\\"c6\\\" colnum=\\\"6\\\"\\u003e\\u003c/div\\u003e \\u003cdiv align=\\\"left\\\" class=\\\"colspec\\\" colname=\\\"c7\\\" colnum=\\\"7\\\"\\u003e\\u003c/div\\u003e \\u003cdiv align=\\\"left\\\" class=\\\"colspec\\\" colname=\\\"c8\\\" colnum=\\\"8\\\"\\u003e\\u003c/div\\u003e \\u003cdiv align=\\\"left\\\" class=\\\"colspec\\\" colname=\\\"c9\\\" colnum=\\\"9\\\"\\u003e\\u003c/div\\u003e \\u003cdiv align=\\\"left\\\" class=\\\"colspec\\\" colname=\\\"c10\\\" colnum=\\\"10\\\"\\u003e\\u003c/div\\u003e \\u003cdiv align=\\\"left\\\" class=\\\"colspec\\\" colname=\\\"c11\\\" colnum=\\\"11\\\"\\u003e\\u003c/div\\u003e \\u003cdiv align=\\\"left\\\" class=\\\"colspec\\\" colname=\\\"c12\\\" colnum=\\\"12\\\"\\u003e\\u003c/div\\u003e \\u003cdiv align=\\\"left\\\" class=\\\"colspec\\\" colname=\\\"c13\\\" colnum=\\\"13\\\"\\u003e\\u003c/div\\u003e \\u003cdiv align=\\\"left\\\" class=\\\"colspec\\\" colname=\\\"c14\\\" colnum=\\\"14\\\"\\u003e\\u003c/div\\u003e \\u003cthead\\u003e \\u003ctr\\u003e \\u003cth align=\\\"left\\\" colname=\\\"c1\\\" morerows=\\\"2\\\" rowspan=\\\"3\\\"\\u003e \\u003cp\\u003eNumber of F\\u003csub\\u003e2\\u003c/sub\\u003e plants\\u003c/p\\u003e \\u003c/th\\u003e \\u003cth align=\\\"left\\\" colname=\\\"c2\\\" morerows=\\\"2\\\" rowspan=\\\"3\\\"\\u003e \\u003cp\\u003ePhenotype to races 20\\u0026thinsp;\\u0026minus;\\u0026thinsp;6 and 31\\u0026thinsp;\\u0026minus;\\u0026thinsp;1\\u003c/p\\u003e \\u003c/th\\u003e \\u003cth align=\\\"left\\\" colname=\\\"c3\\\"\\u003e\\u0026nbsp;\\u003c/th\\u003e \\u003cth align=\\\"left\\\" colspan=\\\"11\\\" nameend=\\\"c14\\\" namest=\\\"c4\\\"\\u003e \\u003cp\\u003eGenotyping of F\\u003csub\\u003e2\\u003c/sub\\u003e plants with PACE markers\\u003csup\\u003e1\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/th\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003cth align=\\\"left\\\" colname=\\\"c3\\\"\\u003e\\u0026nbsp;\\u003c/th\\u003e \\u003cth align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003ePv567309\\u003c/p\\u003e \\u003c/th\\u003e \\u003cth align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003ePv576749\\u003c/p\\u003e \\u003c/th\\u003e \\u003cth align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003ePv626590\\u003c/p\\u003e \\u003c/th\\u003e \\u003cth align=\\\"left\\\" colname=\\\"c7\\\"\\u003e \\u003cp\\u003ePv696542\\u003c/p\\u003e \\u003c/th\\u003e \\u003cth align=\\\"left\\\" colname=\\\"c8\\\"\\u003e \\u003cp\\u003ePv859224\\u003c/p\\u003e \\u003c/th\\u003e \\u003cth align=\\\"left\\\" colname=\\\"c9\\\"\\u003e \\u003cp\\u003ePv1178763\\u003c/p\\u003e \\u003c/th\\u003e \\u003cth align=\\\"left\\\" colname=\\\"c10\\\"\\u003e \\u003cp\\u003ePv1243509\\u003c/p\\u003e \\u003c/th\\u003e \\u003cth align=\\\"left\\\" colname=\\\"c11\\\"\\u003e \\u003cp\\u003ePv1350344\\u003c/p\\u003e \\u003c/th\\u003e \\u003cth align=\\\"left\\\" colname=\\\"c12\\\"\\u003e \\u003cp\\u003ePv1604777\\u003c/p\\u003e \\u003c/th\\u003e \\u003cth align=\\\"left\\\" colname=\\\"c13\\\"\\u003e \\u003cp\\u003ePv1713242\\u003c/p\\u003e \\u003c/th\\u003e \\u003cth align=\\\"left\\\" colname=\\\"c14\\\"\\u003e \\u003cp\\u003ePv1776022\\u003c/p\\u003e \\u003c/th\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003cth align=\\\"left\\\" colname=\\\"c3\\\"\\u003e\\u0026nbsp;\\u003c/th\\u003e \\u003cth align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003e567,309\\u003csup\\u003e2\\u003c/sup\\u003e\\u003c/p\\u003e \\u003c/th\\u003e \\u003cth align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003e576,749\\u003c/p\\u003e \\u003c/th\\u003e \\u003cth align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003e626,590\\u003c/p\\u003e \\u003c/th\\u003e \\u003cth align=\\\"left\\\" colname=\\\"c7\\\"\\u003e \\u003cp\\u003e696,542\\u003c/p\\u003e \\u003c/th\\u003e \\u003cth align=\\\"left\\\" colname=\\\"c8\\\"\\u003e \\u003cp\\u003e859,224\\u003c/p\\u003e \\u003c/th\\u003e \\u003cth align=\\\"left\\\" colname=\\\"c9\\\"\\u003e \\u003cp\\u003e1,178,763\\u003c/p\\u003e \\u003c/th\\u003e \\u003cth align=\\\"left\\\" colname=\\\"c10\\\"\\u003e \\u003cp\\u003e1,243,509\\u003c/p\\u003e \\u003c/th\\u003e \\u003cth align=\\\"left\\\" colname=\\\"c11\\\"\\u003e \\u003cp\\u003e1,350,344\\u003c/p\\u003e \\u003c/th\\u003e \\u003cth align=\\\"left\\\" colname=\\\"c12\\\"\\u003e \\u003cp\\u003e1,604,777\\u003c/p\\u003e \\u003c/th\\u003e \\u003cth align=\\\"left\\\" colname=\\\"c13\\\"\\u003e \\u003cp\\u003e1,713,242\\u003c/p\\u003e \\u003c/th\\u003e \\u003cth align=\\\"left\\\" colname=\\\"c14\\\"\\u003e \\u003cp\\u003e1,776,022\\u003c/p\\u003e \\u003c/th\\u003e \\u003c/tr\\u003e \\u003c/thead\\u003e \\u003ctbody\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003eUI114\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003eSusceptible\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e\\u0026nbsp;\\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003eAA\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003eAA\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003eAA\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c7\\\"\\u003e \\u003cp\\u003eAA\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c8\\\"\\u003e \\u003cp\\u003eAA\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c9\\\"\\u003e \\u003cp\\u003eAA\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c10\\\"\\u003e \\u003cp\\u003eAA\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c11\\\"\\u003e \\u003cp\\u003eAA\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c12\\\"\\u003e \\u003cp\\u003eAA\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c13\\\"\\u003e \\u003cp\\u003eAA\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c14\\\"\\u003e \\u003cp\\u003eAA\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003eCNC\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003eResistant\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e\\u0026nbsp;\\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003eBB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003eBB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003eBB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c7\\\"\\u003e \\u003cp\\u003eBB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c8\\\"\\u003e \\u003cp\\u003eBB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c9\\\"\\u003e \\u003cp\\u003eBB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c10\\\"\\u003e \\u003cp\\u003eBB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c11\\\"\\u003e \\u003cp\\u003eBB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c12\\\"\\u003e \\u003cp\\u003eBB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c13\\\"\\u003e \\u003cp\\u003eBB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c14\\\"\\u003e \\u003cp\\u003eBB\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e\\u0026nbsp;\\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e\\u0026nbsp;\\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e\\u0026nbsp;\\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e\\u0026nbsp;\\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e\\u0026nbsp;\\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e\\u0026nbsp;\\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colspan=\\\"5\\\" nameend=\\\"c11\\\" namest=\\\"c7\\\"\\u003e \\u003cp\\u003e\\u003cem\\u003eGenomic region containing the rust resistance gene in CNC\\u003c/em\\u003e\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c12\\\"\\u003e\\u0026nbsp;\\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c13\\\"\\u003e\\u0026nbsp;\\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c14\\\"\\u003e\\u0026nbsp;\\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e3\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003eResistant\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e\\u0026nbsp;\\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003eAA\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003eAA\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003eAA\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c7\\\"\\u003e \\u003cp\\u003eAA\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c8\\\"\\u003e \\u003cp\\u003eAB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c9\\\"\\u003e \\u003cp\\u003eAB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c10\\\"\\u003e \\u003cp\\u003eAB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c11\\\"\\u003e \\u003cp\\u003eAB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c12\\\"\\u003e \\u003cp\\u003eAB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c13\\\"\\u003e \\u003cp\\u003eAB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c14\\\"\\u003e \\u003cp\\u003eAB\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003eResistant\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e\\u0026nbsp;\\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003eAA\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003eAA\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003eAA\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c7\\\"\\u003e \\u003cp\\u003eAB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c8\\\"\\u003e \\u003cp\\u003eAB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c9\\\"\\u003e \\u003cp\\u003eAB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c10\\\"\\u003e \\u003cp\\u003eAB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c11\\\"\\u003e \\u003cp\\u003eAB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c12\\\"\\u003e \\u003cp\\u003eAB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c13\\\"\\u003e \\u003cp\\u003eAB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c14\\\"\\u003e \\u003cp\\u003eAB\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003eResistant\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e\\u0026nbsp;\\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003eAA\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003eAA\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003eAB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c7\\\"\\u003e \\u003cp\\u003eAB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c8\\\"\\u003e \\u003cp\\u003eAB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c9\\\"\\u003e \\u003cp\\u003eAB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c10\\\"\\u003e \\u003cp\\u003eAB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c11\\\"\\u003e \\u003cp\\u003eAB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c12\\\"\\u003e \\u003cp\\u003eAB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c13\\\"\\u003e \\u003cp\\u003eAB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c14\\\"\\u003e \\u003cp\\u003eAB\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e2\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003eResistant\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e\\u0026nbsp;\\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003eBB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003eBB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003eBB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c7\\\"\\u003e \\u003cp\\u003eBB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c8\\\"\\u003e \\u003cp\\u003eBB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c9\\\"\\u003e \\u003cp\\u003eBB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c10\\\"\\u003e \\u003cp\\u003eBB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c11\\\"\\u003e \\u003cp\\u003eBB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c12\\\"\\u003e \\u003cp\\u003eBB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c13\\\"\\u003e \\u003cp\\u003eAB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c14\\\"\\u003e \\u003cp\\u003eAB\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003eResistant\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e\\u0026nbsp;\\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003eBB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003eBB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003eBB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c7\\\"\\u003e \\u003cp\\u003eBB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c8\\\"\\u003e \\u003cp\\u003eBB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c9\\\"\\u003e \\u003cp\\u003eBB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c10\\\"\\u003e \\u003cp\\u003eBB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c11\\\"\\u003e \\u003cp\\u003eBB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c12\\\"\\u003e \\u003cp\\u003eBB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c13\\\"\\u003e \\u003cp\\u003eBB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c14\\\"\\u003e \\u003cp\\u003eAB\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e72\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003eResistant\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e\\u0026nbsp;\\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003eBB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003eBB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003eBB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c7\\\"\\u003e \\u003cp\\u003eBB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c8\\\"\\u003e \\u003cp\\u003eBB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c9\\\"\\u003e \\u003cp\\u003eBB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c10\\\"\\u003e \\u003cp\\u003eBB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c11\\\"\\u003e \\u003cp\\u003eBB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c12\\\"\\u003e \\u003cp\\u003eBB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c13\\\"\\u003e \\u003cp\\u003eBB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c14\\\"\\u003e \\u003cp\\u003eBB\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e125\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003eResistant\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e\\u0026nbsp;\\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003eAB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003eAB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003eAB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c7\\\"\\u003e \\u003cp\\u003eAB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c8\\\"\\u003e \\u003cp\\u003eAB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c9\\\"\\u003e \\u003cp\\u003eAB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c10\\\"\\u003e \\u003cp\\u003eAB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c11\\\"\\u003e \\u003cp\\u003eAB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c12\\\"\\u003e \\u003cp\\u003eAB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c13\\\"\\u003e \\u003cp\\u003eAB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c14\\\"\\u003e \\u003cp\\u003eAB\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003eResistant\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e\\u0026nbsp;\\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003eAB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003eAB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003eAB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c7\\\"\\u003e \\u003cp\\u003eAB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c8\\\"\\u003e \\u003cp\\u003eAB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c9\\\"\\u003e \\u003cp\\u003eAB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c10\\\"\\u003e \\u003cp\\u003eAB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c11\\\"\\u003e \\u003cp\\u003eAB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c12\\\"\\u003e \\u003cp\\u003eAA\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c13\\\"\\u003e \\u003cp\\u003eAA\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c14\\\"\\u003e \\u003cp\\u003eAA\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003eResistant\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e\\u0026nbsp;\\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003eAB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003eAB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003eAB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c7\\\"\\u003e \\u003cp\\u003eAB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c8\\\"\\u003e \\u003cp\\u003eAB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c9\\\"\\u003e \\u003cp\\u003eAB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c10\\\"\\u003e \\u003cp\\u003eAB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c11\\\"\\u003e \\u003cp\\u003eAA\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c12\\\"\\u003e \\u003cp\\u003eAA\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c13\\\"\\u003e \\u003cp\\u003eAA\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c14\\\"\\u003e \\u003cp\\u003eAA\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003eSusceptible\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e\\u0026nbsp;\\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003eAB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003eAB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003eAB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c7\\\"\\u003e \\u003cp\\u003eAB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c8\\\"\\u003e \\u003cp\\u003eAA\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c9\\\"\\u003e \\u003cp\\u003eAA\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c10\\\"\\u003e \\u003cp\\u003eAA\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c11\\\"\\u003e \\u003cp\\u003eAA\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c12\\\"\\u003e \\u003cp\\u003eAA\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c13\\\"\\u003e \\u003cp\\u003eAA\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c14\\\"\\u003e \\u003cp\\u003eAA\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003eSusceptible\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e\\u0026nbsp;\\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003eAB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003eAB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003eAA\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c7\\\"\\u003e \\u003cp\\u003eAA\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c8\\\"\\u003e \\u003cp\\u003eAA\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c9\\\"\\u003e \\u003cp\\u003eAA\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c10\\\"\\u003e \\u003cp\\u003eAA\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c11\\\"\\u003e \\u003cp\\u003eAA\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c12\\\"\\u003e \\u003cp\\u003eAA\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c13\\\"\\u003e \\u003cp\\u003eAA\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c14\\\"\\u003e \\u003cp\\u003eAA\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e65\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003eSusceptible\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e\\u0026nbsp;\\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003eAA\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003eAA\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003eAA\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c7\\\"\\u003e \\u003cp\\u003eAA\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c8\\\"\\u003e \\u003cp\\u003eAA\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c9\\\"\\u003e \\u003cp\\u003eAA\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c10\\\"\\u003e \\u003cp\\u003eAA\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c11\\\"\\u003e \\u003cp\\u003eAA\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c12\\\"\\u003e \\u003cp\\u003eAA\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c13\\\"\\u003e \\u003cp\\u003eAA\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c14\\\"\\u003e \\u003cp\\u003eAA\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003e1\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003eSusceptible\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e\\u0026nbsp;\\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003eAA\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e \\u003cp\\u003eAA\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e \\u003cp\\u003eAA\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c7\\\"\\u003e \\u003cp\\u003eAA\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c8\\\"\\u003e \\u003cp\\u003eAA\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c9\\\"\\u003e \\u003cp\\u003eAA\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c10\\\"\\u003e \\u003cp\\u003eAA\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c11\\\"\\u003e \\u003cp\\u003eAA\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c12\\\"\\u003e \\u003cp\\u003eAA\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c13\\\"\\u003e \\u003cp\\u003eAB\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c14\\\"\\u003e \\u003cp\\u003eAB\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003c/tbody\\u003e \\u003c/colgroup\\u003e \\u003ctfoot\\u003e \\u003ctr\\u003e\\u003ctd colspan=\\\"14\\\"\\u003e\\u003csup\\u003e1\\u003c/sup\\u003eSNP position based on the reference genome of UI111 v1.1; AA\\u0026thinsp;=\\u0026thinsp;UI114 allele; AB heterozygous; BB\\u0026thinsp;=\\u0026thinsp;CNC allele\\u003c/td\\u003e\\u003c/tr\\u003e \\u003c/tfoot\\u003e \\u003c/table\\u003e\\u003c/div\\u003e \\u003c/p\\u003e \\u003cp\\u003eThe genomic interval delimited by markers Pv696542 and Pv1350344 contained 56 annotated genes according to the UI111 v1.1 reference genome. Among the annotated genes, 11 encode disease-resistance proteins involved in plant defense against pathogens; eight of these genes encode nucleotide-binding domain leucine-rich repeat (NLR) proteins, and three genes encode kinase proteins (Table S4). To further investigate, we used the NCBI Conserved Domain Search tool (\\u003cspan class=\\\"ExternalRef\\\"\\u003e\\u003cspan class=\\\"RefSource\\\"\\u003ehttps://www.ncbi.nlm.nih.gov/Structure/cdd/wrpsb.cgi\\u003c/span\\u003e\\u003cspan address=\\\"https://www.ncbi.nlm.nih.gov/Structure/cdd/wrpsb.cgi\\\" targettype=\\\"URL\\\" class=\\\"RefTarget\\\"\\u003e\\u003c/span\\u003e\\u003c/span\\u003e) to identify conserved domains within these 11 genes. Two genes, which are annotated as NLRs in the UI111 reference genome v1.1, did not display any domains typically associated with plant resistance. The remaining nine genes exhibited either NLR or kinase domains and are strong candidates for the rust resistance gene present in CNC (Table\\u0026nbsp;\\u003cspan refid=\\\"Tab3\\\" class=\\\"InternalRef\\\"\\u003e3\\u003c/span\\u003e).\\u003c/p\\u003e \\u003cp\\u003e \\u003cdiv class=\\\"gridtable\\\"\\u003e\\u003ctable float=\\\"Yes\\\" id=\\\"Tab3\\\" border=\\\"1\\\"\\u003e \\u003ccaption language=\\\"En\\\"\\u003e \\u003cdiv class=\\\"CaptionNumber\\\"\\u003eTable 3\\u003c/div\\u003e \\u003cdiv class=\\\"CaptionContent\\\"\\u003e \\u003cp\\u003eCandidate genes encoding proteins related to disease resistance in the genomic region containing the rust resistance gene in CNC on the common bean reference genome UI111 v1.1\\u003c/p\\u003e \\u003c/div\\u003e \\u003c/caption\\u003e \\u003ccolgroup cols=\\\"4\\\"\\u003e \\u003cdiv align=\\\"left\\\" class=\\\"colspec\\\" colname=\\\"c1\\\" colnum=\\\"1\\\"\\u003e\\u003c/div\\u003e \\u003cdiv align=\\\"char\\\" char=\\\".\\\" class=\\\"colspec\\\" colname=\\\"c2\\\" colnum=\\\"2\\\"\\u003e\\u003c/div\\u003e \\u003cdiv align=\\\"char\\\" char=\\\".\\\" class=\\\"colspec\\\" colname=\\\"c3\\\" colnum=\\\"3\\\"\\u003e\\u003c/div\\u003e \\u003cdiv align=\\\"left\\\" class=\\\"colspec\\\" colname=\\\"c4\\\" colnum=\\\"4\\\"\\u003e\\u003c/div\\u003e \\u003cthead\\u003e \\u003ctr\\u003e \\u003cth align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003eGene\\u003c/p\\u003e \\u003c/th\\u003e \\u003cth align=\\\"left\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003eStart\\u003c/p\\u003e \\u003c/th\\u003e \\u003cth align=\\\"left\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003eEnd\\u003c/p\\u003e \\u003c/th\\u003e \\u003cth align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003ePredict protein\\u003c/p\\u003e \\u003c/th\\u003e \\u003c/tr\\u003e \\u003c/thead\\u003e \\u003ctbody\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003ePvUI111.04G007600\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"char\\\" char=\\\".\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e709963\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"char\\\" char=\\\".\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e714521\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003eLeucine-rich repeat-containing protein\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003ePvUI111.04G007900\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"char\\\" char=\\\".\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e723,321\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"char\\\" char=\\\".\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e726,702\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003eLeucine-rich repeat-containing protein\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003ePvUI111.04G008100\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"char\\\" char=\\\".\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e729,478\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"char\\\" char=\\\".\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e734,847\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003eLeucine-rich repeat-containing protein\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003ePvUI111.04G008800\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"char\\\" char=\\\".\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e813,803\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"char\\\" char=\\\".\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e817,714\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003eSerine/threonine-protein kinase\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003ePvUI111.04G009400\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"char\\\" char=\\\".\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e889,655\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"char\\\" char=\\\".\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e895,350\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003eCysteine-rich receptor-like protein kinase\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003ePvUI111.04G011000\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"char\\\" char=\\\".\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e1,170,791\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"char\\\" char=\\\".\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e1,183,177\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003eProtein kinase\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003ePvUI111.04G011200\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"char\\\" char=\\\".\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e1,213,139\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"char\\\" char=\\\".\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e1,218,585\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003eLeucine-rich repeat-containing protein\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003ePvUI111.04G011300\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"char\\\" char=\\\".\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e1,221,763\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"char\\\" char=\\\".\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e1,225,645\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003eLeucine-rich repeat-containing protein\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003ctr\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e \\u003cp\\u003ePvUI111.04G011400\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"char\\\" char=\\\".\\\" colname=\\\"c2\\\"\\u003e \\u003cp\\u003e1,229,108\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"char\\\" char=\\\".\\\" colname=\\\"c3\\\"\\u003e \\u003cp\\u003e1,235,266\\u003c/p\\u003e \\u003c/td\\u003e \\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e \\u003cp\\u003eLeucine-rich repeat-containing protein\\u003c/p\\u003e \\u003c/td\\u003e \\u003c/tr\\u003e \\u003c/tbody\\u003e \\u003c/colgroup\\u003e \\u003c/table\\u003e\\u003c/div\\u003e \\u003c/p\\u003e\"},{\"header\":\"Discussion\",\"content\":\"\\u003cp\\u003e \\u003cem\\u003eU. appendiculatus\\u003c/em\\u003e has high genetic diversity and it is constantly evolving to overcome the resistance genes deployed in bean cultivars. In this study, we characterized a rust resistance locus in the cultivar CNC and mapped it to chromosome Pv4 of \\u003cem\\u003eP. vulgaris\\u003c/em\\u003e. In addition, we identified closely linked SNP-based markers, which can be used by breeding programs in marker-assisted selection (MAS). To our knowledge, the only rust resistance germplasm derived from CNC are the pintos BelDak-RR-1 and BelDak-RR-2, which also contain \\u003cem\\u003eUr-3\\u003c/em\\u003e and \\u003cem\\u003eUr-6\\u003c/em\\u003e (Stavely and Grafton \\u003cspan citationid=\\\"CR45\\\" class=\\\"CitationRef\\\"\\u003e1989\\u003c/span\\u003e).\\u003c/p\\u003e \\u003cp\\u003eCNC is resistant to 81 of 88 races maintained in the \\u003cem\\u003eU. appendiculatus\\u003c/em\\u003e collection at the USDA-ARS in Beltsville MD, and it is particularly highly resistant to races present in the United States (Stavely \\u003cspan citationid=\\\"CR42\\\" class=\\\"CitationRef\\\"\\u003e1984\\u003c/span\\u003e; Stavely et al. \\u003cspan citationid=\\\"CR46\\\" class=\\\"CitationRef\\\"\\u003e1989\\u003c/span\\u003e). In North Dakota, which is the leading U.S. bean producer, bean rust is one of the major yield-limiting factors. Therefore, increased efforts have been made to improve rust resistance in bean germplasm since the mid-1980s (MacQueen et al. \\u003cspan citationid=\\\"CR20\\\" class=\\\"CitationRef\\\"\\u003e2020\\u003c/span\\u003e; Osorno et al. \\u003cspan citationid=\\\"CR31\\\" class=\\\"CitationRef\\\"\\u003e2021\\u003c/span\\u003e; Miklas et al. \\u003cspan citationid=\\\"CR28\\\" class=\\\"CitationRef\\\"\\u003e2023\\u003c/span\\u003e). Until 2002, five \\u003cem\\u003eU. appendiculatus\\u003c/em\\u003e races (52, 54, 69, 70, and 71) were reported in North Dakota, none of which were virulent on CNC (Gross and Venette \\u003cspan citationid=\\\"CR13\\\" class=\\\"CitationRef\\\"\\u003e2002\\u003c/span\\u003e). More recent surveys showed that 20\\u0026thinsp;\\u0026minus;\\u0026thinsp;3 is the most frequent race found in dry bean fields in North Dakota with 50\\u0026ndash;70% of the rust isolates belonging to this race (Monclova-Santana \\u003cspan citationid=\\\"CR30\\\" class=\\\"CitationRef\\\"\\u003e2019\\u003c/span\\u003e). CNC serves as an important source of resistance to race 20\\u0026thinsp;\\u0026minus;\\u0026thinsp;3 and can be used by breeding programs aiming to develop rust resistance cultivars for North Dakota (Monclova-Santana \\u003cspan citationid=\\\"CR30\\\" class=\\\"CitationRef\\\"\\u003e2019\\u003c/span\\u003e).\\u003c/p\\u003e \\u003cp\\u003eTo understand the genetics of rust resistance in CNC, we employed BSA coupled with NGS on an F\\u003csub\\u003e2\\u003c/sub\\u003e population from cross UI114 \\u0026times; CNC for the rapid identification of genes/genomic loci in CNC governing the rust resistance to races 20\\u0026thinsp;\\u0026minus;\\u0026thinsp;6 and 31\\u0026thinsp;\\u0026minus;\\u0026thinsp;1. NGS-BSA resulted in the identification of a genomic region associated with resistance on chromosome Pv04 which was further confirmed by genotyping the complete F\\u003csub\\u003e2\\u003c/sub\\u003e population using PACE markers. Chromosome Pv04 comprises one of the most complex clusters of rust resistance in common bean, including genes reported in the Middle American cultivars Mexico 309 (\\u003cem\\u003eUr-5\\u003c/em\\u003e), Ouro Negro (\\u003cem\\u003eUr-14\\u003c/em\\u003e), Dorado, and PI 310762, and Andean cultivars PI 260418 and G19833 (Miklas et al. \\u003cspan citationid=\\\"CR29\\\" class=\\\"CitationRef\\\"\\u003e2000\\u003c/span\\u003e; Shin et al. \\u003cspan citationid=\\\"CR40\\\" class=\\\"CitationRef\\\"\\u003e2014\\u003c/span\\u003e; Valentini et al. \\u003cspan citationid=\\\"CR52\\\" class=\\\"CitationRef\\\"\\u003e2020\\u003c/span\\u003e; Valentini et al. \\u003cspan citationid=\\\"CR51\\\" class=\\\"CitationRef\\\"\\u003e2021\\u003c/span\\u003e). An allelism test has confirmed the independence of \\u003cem\\u003eUr-CNC\\u003c/em\\u003e and \\u003cem\\u003eUr-14\\u003c/em\\u003e, but allelism tests with genotypes containing other genes have never been performed (Souza et al. \\u003cspan citationid=\\\"CR41\\\" class=\\\"CitationRef\\\"\\u003e2011\\u003c/span\\u003e). Although the rust resistance in CNC can be distinguished from the above-mentioned cultivars by its unique spectrum of resistance, the overlap of the CNC region with previously identified genomic locations of other resistance genes add further complexities about its independence. Specifically, the gene in CNC was mapped between 696,542 and 1,350,344 bp (UI111 v1.0) the gene in PI 310762 is positioned between 246,091 and 1,164,610 bp (G19833 v1.0); the \\u003cem\\u003eUr-5\\u003c/em\\u003e gene in Mexico 309 is mapped between 381,360 and 1,407,905 bp (G19833 v1.0); the gene in PI260418 is mapped between 303,697 and 1,299,082 bp (G19833 v2.1); and the gene in G19833 lies between 554,115 and 1,301,156 bp (G19833 v2.1) (Shin et al. \\u003cspan citationid=\\\"CR40\\\" class=\\\"CitationRef\\\"\\u003e2014\\u003c/span\\u003e; Valentini et al. \\u003cspan citationid=\\\"CR52\\\" class=\\\"CitationRef\\\"\\u003e2020\\u003c/span\\u003e; Valentini et al. \\u003cspan citationid=\\\"CR51\\\" class=\\\"CitationRef\\\"\\u003e2021\\u003c/span\\u003e). Therefore, additional allelism tests with large segregating populations and fine mapping will be crucial to confirm the independence of these genes.\\u003c/p\\u003e \\u003cp\\u003eSeveral features make the resistance gene cluster on Pv04 particularly interesting for further investigations. Firstly, in contrast to other resistance gene clusters, the Pv04 cluster contains disease-resistance genes from both the Andean and Middle American gene pools, suggesting that this cluster existed prior to the geographic separation of \\u003cem\\u003eP. vulgaris\\u003c/em\\u003e into the two gene pools (Geffroy et al. \\u003cspan citationid=\\\"CR10\\\" class=\\\"CitationRef\\\"\\u003e2009\\u003c/span\\u003e; Geffroy et al. \\u003cspan citationid=\\\"CR11\\\" class=\\\"CitationRef\\\"\\u003e1999\\u003c/span\\u003e; Vlasova et al. \\u003cspan citationid=\\\"CR56\\\" class=\\\"CitationRef\\\"\\u003e2016\\u003c/span\\u003e). On the other hand, resistance gene clusters on Pv01 and Pv11 seem to be originated independently in each gene pool. For example, Pv01 contains genes of Andean origin for resistance to rust (\\u003cem\\u003eUr-9\\u003c/em\\u003e) and anthracnose (\\u003cem\\u003eCo-1, Co-x, Co-AC\\u003c/em\\u003e, and \\u003cem\\u003eCo-Pa\\u003c/em\\u003e) (Miklas et al. \\u003cspan citationid=\\\"CR27\\\" class=\\\"CitationRef\\\"\\u003e2006\\u003c/span\\u003e; Richard et al. \\u003cspan citationid=\\\"CR36\\\" class=\\\"CitationRef\\\"\\u003e2014\\u003c/span\\u003e; Castro et al. \\u003cspan citationid=\\\"CR5\\\" class=\\\"CitationRef\\\"\\u003e2017\\u003c/span\\u003e; Gilio et al. \\u003cspan citationid=\\\"CR12\\\" class=\\\"CitationRef\\\"\\u003e2020\\u003c/span\\u003e), and Pv11 contains genes of Middle American origin for resistance to rust (\\u003cem\\u003eUr-3, Ur-7\\u003c/em\\u003e, and \\u003cem\\u003eUr-11\\u003c/em\\u003e) and anthracnose (\\u003cem\\u003eCo-2\\u003c/em\\u003e) (Miklas et al. \\u003cspan citationid=\\\"CR27\\\" class=\\\"CitationRef\\\"\\u003e2006\\u003c/span\\u003e), suggesting that the resistance gene clusters might be derived from specific gene pools. Secondly, the majority of the resistance-associated genes in the Pv04 cluster encode typical NLR domains, although genes encoding kinase domains are also present (Schmutz et al. \\u003cspan citationid=\\\"CR38\\\" class=\\\"CitationRef\\\"\\u003e2014\\u003c/span\\u003e; Vas Bisneta and Gon\\u0026ccedil;alves-Vidigal 2020). The combination of these two pathogen recognition systems provides the host with greater flexibility, allowing for the rapid evolution of resistance genes and enabling the plant to adapt to new or evolving pathogens over time. Although it has not been elucidated in common beans, the interaction of an NLR and Kinases was recently demonstrated in wheat, showing that an NLR with tandem NB-ARC domains (WTN1) cooperates with a tandem kinase (WTK3 and its allelic variant Rwt4 to constitute a tandem kinase-NLR immune signaling pathway (Lu et al. \\u003cspan citationid=\\\"CR18\\\" class=\\\"CitationRef\\\"\\u003e2025\\u003c/span\\u003e). In another example, Chen et al. (\\u003cspan citationid=\\\"CR6\\\" class=\\\"CitationRef\\\"\\u003e2025\\u003c/span\\u003e) discovered that the \\u003cem\\u003eSr62\\u003c/em\\u003e locus consists of a digenic module encoding the tandem kinase Sr62\\u003csup\\u003eTK\\u003c/sup\\u003e and an NLR (Sr62\\u003csup\\u003eNLR\\u003c/sup\\u003e). The pathogen effector AvrSr62 triggers the activation of the wheat Sr62\\u003csup\\u003eTK\\u003c/sup\\u003e, which then activates the corresponding NLR.\\u003c/p\\u003e \\u003cp\\u003eDue to the low recombination rates in certain Pv04 regions, it is particularly challenging to narrow down a genomic interval containing the resistance gene. The region containing the gene cluster associated with disease resistance on top of Pv04 originates from ectopic recombination between subtelomeric regions of nonhomologous chromosomes. Specifically, genes encoding Coiled-Coil-Nucleotide-Binding-Site-Leucine-Rich-Repeat (CNL) on the Pv04 gene cluster were derived from CNL of the Pv11 cluster through an ectopic recombination event that occurred more than 19 MYA before the divergence of \\u003cem\\u003eP. vulgaris\\u003c/em\\u003e and \\u003cem\\u003eGlycine max\\u003c/em\\u003e (David et al. \\u003cspan citationid=\\\"CR9\\\" class=\\\"CitationRef\\\"\\u003e2009\\u003c/span\\u003e; McClean et al. \\u003cspan citationid=\\\"CR23\\\" class=\\\"CitationRef\\\"\\u003e2010\\u003c/span\\u003e). This ectopic recombination resulted in the occurrence of heterochromatic blocks (David et al. \\u003cspan citationid=\\\"CR9\\\" class=\\\"CitationRef\\\"\\u003e2009\\u003c/span\\u003e), which may be associated with recombination suppression in this region as reported in other studies (Meziadi et al. \\u003cspan citationid=\\\"CR26\\\" class=\\\"CitationRef\\\"\\u003e2016\\u003c/span\\u003e, Valentini et al. \\u003cspan citationid=\\\"CR50\\\" class=\\\"CitationRef\\\"\\u003e2017\\u003c/span\\u003e).\\u003c/p\\u003e \\u003cp\\u003eIn summary, we have genetically characterized and mapped the rust resistance in CNC, which exhibits broad resistance against multiple races of \\u003cem\\u003eU. appendiculatus\\u003c/em\\u003e. The strong resistance conferred by CNC highlights its potential value for enhancing common bean germplasm. Furthermore, the development of molecular markers in this study offers valuable tools for diversifying the deployment of rust resistance in common bean breeding programs.\\u003c/p\\u003e\"},{\"header\":\"Declarations\",\"content\":\" \\u003ch2\\u003eConflict of interest\\u003c/h2\\u003e \\u003cp\\u003eThe authors declare that there is no conflict of interest.\\u003c/p\\u003e \\u003ch2\\u003eEthical approval\\u003c/strong\\u003e \\u003cp\\u003eThe study does not require ethics approval as we have not used human or animal subject.\\u003c/p\\u003e\\u003ch2\\u003eAuthor contributions\\u003c/h2\\u003e \\u003cp\\u003eGV and UG conceived the original idea and designed the research., GV developed mapping populations, performed phenotypic and genotypic analysis, and collected data. GV, AS, JS, and UG performed the data analysis. GV and UG prepared the manuscript with inputs from AS and JS. All authors have read and approved the manuscript. UG secured the funding for this research.\\u003c/p\\u003e\\u003ch2\\u003eAcknowledgments\\u003c/h2\\u003e \\u003cp\\u003eThis work was supported by the Specialty Crop Block Grant Program, North Dakota Department of Agriculture (NOGA-22-235), USDA-National Institute of Food and Agriculture (NIFA) Hatch project ND02243, and USDA-NIFA Research Capacity Fund project 7006432.\\u003c/p\\u003e\\u003ch2\\u003eData availability\\u003c/h2\\u003e \\u003cp\\u003eAll data and research materials are available upon reasonable request from the corresponding author.\\u003c/p\\u003e\"},{\"header\":\"References\",\"content\":\"\\u003col\\u003e\\u003cli\\u003e\\u003cspan\\u003eAndrews S (2010) A quality control tool for high throughput sequence data. 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J Hered 93:77\\u0026ndash;78. \\u003cspan class=\\\"ExternalRef\\\"\\u003e\\u003cspan class=\\\"RefSource\\\"\\u003ehttps://doi.org/10.1093/jhered/93.1.77\\u003c/span\\u003e\\u003cspan address=\\\"10.1093/jhered/93.1.77\\\" targettype=\\\"DOI\\\" class=\\\"RefTarget\\\"\\u003e\\u003c/span\\u003e\\u003c/span\\u003e\\u003c/span\\u003e\\u003c/li\\u003e\\u003c/ol\\u003e\"}],\"fulltextSource\":\"\",\"fullText\":\"\",\"funders\":[],\"hasAdminPriorityOnWorkflow\":false,\"hasManuscriptDocX\":true,\"hasOptedInToPreprint\":true,\"hasPassedJournalQc\":\"\",\"hasAnyPriority\":false,\"hideJournal\":true,\"highlight\":\"\",\"institution\":\"\",\"isAcceptedByJournal\":false,\"isAuthorSuppliedPdf\":false,\"isDeskRejected\":\"\",\"isHiddenFromSearch\":false,\"isInQc\":false,\"isInWorkflow\":false,\"isPdf\":false,\"isPdfUpToDate\":true,\"isWithdrawnOrRetracted\":false,\"journal\":{\"display\":true,\"email\":\"info@researchsquare.com\",\"identity\":\"researchsquare\",\"isNatureJournal\":false,\"hasQc\":true,\"allowDirectSubmit\":true,\"externalIdentity\":\"\",\"sideBox\":\"\",\"snPcode\":\"\",\"submissionUrl\":\"/submission\",\"title\":\"Research Square\",\"twitterHandle\":\"researchsquare\",\"acdcEnabled\":true,\"dfaEnabled\":false,\"editorialSystem\":\"\",\"reportingPortfolio\":\"\",\"inReviewEnabled\":false,\"inReviewRevisionsEnabled\":true},\"keywords\":\"Phaseolus vulgaris L., Uromyces appendiculatus, Compuesto Negro Chimaltenango, Bulked segregant analysis, resistance mapping\",\"lastPublishedDoi\":\"10.21203/rs.3.rs-6572527/v1\",\"lastPublishedDoiUrl\":\"https://doi.org/10.21203/rs.3.rs-6572527/v1\",\"license\":{\"name\":\"CC BY 4.0\",\"url\":\"https://creativecommons.org/licenses/by/4.0/\"},\"manuscriptAbstract\":\"\\u003cp\\u003eRust, caused by the fungus \\u003cem\\u003eUromyces appendiculatus\\u003c/em\\u003e, is one of the most significant diseases of common beans in the United States and worldwide. The identification and characterization of new rust resistance genes are essential for developing cultivars with broad and durable resistance. The black common bean cultivar Compuesto Negro Chimaltenango (CNC) exhibits broad resistance to most known races of the rust pathogen, still its resistance has not been fully characterized. To genetically characterize and map the resistance loci in this cultivar, we crossed it with susceptible cultivar UI114 and evaluated the F\\u003csub\\u003e2\\u003c/sub\\u003e population for resistance against races 20-6 and 31-1 of \\u003cem\\u003eU. appendiculatus\\u003c/em\\u003e. Our results showed that rust resistance in CNC for races 20-6 and 31-1 co-segregates and is conditioned by a single dominant gene. Whole genome sequencing-based bulked segregant analysis identified a genomic region on the proximal end of chromosome 4 associated with rust resistance in CNC. To further validate the identified region, molecular markers spanning this region were mapped on the complete F\\u003csub\\u003e2\\u003c/sub\\u003e population. This further delimited the resistance locus in a genomic interval of 653.8 kb. The genetic mapping information and the molecular markers developed in this study will be helpful in developing rust-resistant cultivars of common bean.\\u003c/p\\u003e\",\"manuscriptTitle\":\"Identification and genetic mapping of a rust resistance gene on chromosome Pv04 in the common bean cultivar CNC using bulked segregant sequencing\",\"msid\":\"\",\"msnumber\":\"\",\"nonDraftVersions\":[{\"code\":1,\"date\":\"2025-06-20 15:14:21\",\"doi\":\"10.21203/rs.3.rs-6572527/v1\",\"editorialEvents\":[{\"type\":\"communityComments\",\"content\":0}],\"status\":\"published\",\"journal\":{\"display\":true,\"email\":\"info@researchsquare.com\",\"identity\":\"researchsquare\",\"isNatureJournal\":false,\"hasQc\":true,\"allowDirectSubmit\":true,\"externalIdentity\":\"\",\"sideBox\":\"\",\"snPcode\":\"\",\"submissionUrl\":\"/submission\",\"title\":\"Research Square\",\"twitterHandle\":\"researchsquare\",\"acdcEnabled\":true,\"dfaEnabled\":false,\"editorialSystem\":\"\",\"reportingPortfolio\":\"\",\"inReviewEnabled\":false,\"inReviewRevisionsEnabled\":true}}],\"origin\":\"\",\"ownerIdentity\":\"98d689ba-cf7e-4291-afcc-2bb3481faf5f\",\"owner\":[],\"postedDate\":\"June 20th, 2025\",\"published\":true,\"recentEditorialEvents\":[],\"rejectedJournal\":[],\"revision\":\"\",\"amendment\":\"\",\"status\":\"posted\",\"subjectAreas\":[],\"tags\":[],\"updatedAt\":\"2025-09-19T14:47:33+00:00\",\"versionOfRecord\":[],\"versionCreatedAt\":\"2025-06-20 15:14:21\",\"video\":\"\",\"vorDoi\":\"\",\"vorDoiUrl\":\"\",\"workflowStages\":[]},\"version\":\"v1\",\"identity\":\"rs-6572527\",\"journalConfig\":\"researchsquare\"},\"__N_SSP\":true},\"page\":\"/article/[identity]/[[...version]]\",\"query\":{\"redirect\":\"/article/rs-6572527\",\"identity\":\"rs-6572527\",\"version\":[\"v1\"]},\"buildId\":\"XKTyCvWXoU3ODBz1xrDgd\",\"isFallback\":false,\"isExperimentalCompile\":false,\"dynamicIds\":[84888],\"gssp\":true,\"scriptLoader\":[]}","source_license":"CC-BY-4.0","license_restricted":false}