BUSCADOR RECOLECTA

Table S6 Candidate genes mapped within the genomic regions associated with the significantly associated SNPs detected in response to U. pisi. Chromosomal linkage disequilibrium (LD) decay was considered to limit the genomic regions were to look for candidate genes., Uromyces pisi ([Pers.] D.C.) Wint. is an important foliar biotrophic pathogen infecting grass pea (Lathyrus sativus L.), compromising their yield stability. To date, few efforts have been made to assess the natural variation in grass pea resistance and to identify the resistance loci operating against this pathogen, limiting its efficient breeding exploitation. To overcome this knowledge gap, the genetic architecture of grass pea resistance to U. pisi was investigated using a worldwide collection of 182 accessions through a genome-wide association approach. The response of the grass pea collection to rust infection under controlled conditions and at the seedling stage did not reveal any hypersensitive response but a continuous variation for disease severity, with the identification of promising sources of partial resistance. A panel of 5,651 high-quality single-nucleotide polymorphism (SNP) markers previously generated was used to test for SNP-trait associations, based on a mixed linear model accounting for population structure. We detected seven SNP markers significantly associated with U. pisi disease severity, suggesting that partial resistance is oligogenic. Six of the associated SNP markers were located in chromosomes 4 and 6, while the remaining SNP markers had no known chromosomal position. Through comparative mapping with the pea reference genome, a total of 19 candidate genes were proposed, encoding for leucine-rich repeat, NB-ARC domain, and TGA transcription factor family, among others. Results presented in this study provided information on the availability of partial resistance in grass pea germplasm and advanced our understanding of the molecular mechanisms of quantitative resistance to rust in grass pea. Moreover, the detected associated SNP markers constitute promising genomic targets for the development of molecular tools to assist disease resistance precision breeding., Peer reviewed

Table S5. Inflation factors observed for the models not accounting (Naïve) or accounting for population structure (Eigen), and familiar relatedness (Kinship) tested in the GWAS related to the inoculation experiments with U. pisi., Uromyces pisi ([Pers.] D.C.) Wint. is an important foliar biotrophic pathogen infecting grass pea (Lathyrus sativus L.), compromising their yield stability. To date, few efforts have been made to assess the natural variation in grass pea resistance and to identify the resistance loci operating against this pathogen, limiting its efficient breeding exploitation. To overcome this knowledge gap, the genetic architecture of grass pea resistance to U. pisi was investigated using a worldwide collection of 182 accessions through a genome-wide association approach. The response of the grass pea collection to rust infection under controlled conditions and at the seedling stage did not reveal any hypersensitive response but a continuous variation for disease severity, with the identification of promising sources of partial resistance. A panel of 5,651 high-quality single-nucleotide polymorphism (SNP) markers previously generated was used to test for SNP-trait associations, based on a mixed linear model accounting for population structure. We detected seven SNP markers significantly associated with U. pisi disease severity, suggesting that partial resistance is oligogenic. Six of the associated SNP markers were located in chromosomes 4 and 6, while the remaining SNP markers had no known chromosomal position. Through comparative mapping with the pea reference genome, a total of 19 candidate genes were proposed, encoding for leucine-rich repeat, NB-ARC domain, and TGA transcription factor family, among others. Results presented in this study provided information on the availability of partial resistance in grass pea germplasm and advanced our understanding of the molecular mechanisms of quantitative resistance to rust in grass pea. Moreover, the detected associated SNP markers constitute promising genomic targets for the development of molecular tools to assist disease resistance precision breeding., Peer reviewed

Table S4. Averaged DS scores (±standard error of the mean) measured from inoculation experiments with U. pisi, in accessions grouped based on their geographical origin, seed color, and seed size. The small letter represents significant differences (P-value ≤ 0.05) among accessions grouped by origin, seed size or color., Uromyces pisi ([Pers.] D.C.) Wint. is an important foliar biotrophic pathogen infecting grass pea (Lathyrus sativus L.), compromising their yield stability. To date, few efforts have been made to assess the natural variation in grass pea resistance and to identify the resistance loci operating against this pathogen, limiting its efficient breeding exploitation. To overcome this knowledge gap, the genetic architecture of grass pea resistance to U. pisi was investigated using a worldwide collection of 182 accessions through a genome-wide association approach. The response of the grass pea collection to rust infection under controlled conditions and at the seedling stage did not reveal any hypersensitive response but a continuous variation for disease severity, with the identification of promising sources of partial resistance. A panel of 5,651 high-quality single-nucleotide polymorphism (SNP) markers previously generated was used to test for SNP-trait associations, based on a mixed linear model accounting for population structure. We detected seven SNP markers significantly associated with U. pisi disease severity, suggesting that partial resistance is oligogenic. Six of the associated SNP markers were located in chromosomes 4 and 6, while the remaining SNP markers had no known chromosomal position. Through comparative mapping with the pea reference genome, a total of 19 candidate genes were proposed, encoding for leucine-rich repeat, NB-ARC domain, and TGA transcription factor family, among others. Results presented in this study provided information on the availability of partial resistance in grass pea germplasm and advanced our understanding of the molecular mechanisms of quantitative resistance to rust in grass pea. Moreover, the detected associated SNP markers constitute promising genomic targets for the development of molecular tools to assist disease resistance precision breeding., Peer reviewed

Table S3. Variance components and broad-sense heritability for the DS scores of U. pisi inoculation experiments., Uromyces pisi ([Pers.] D.C.) Wint. is an important foliar biotrophic pathogen infecting grass pea (Lathyrus sativus L.), compromising their yield stability. To date, few efforts have been made to assess the natural variation in grass pea resistance and to identify the resistance loci operating against this pathogen, limiting its efficient breeding exploitation. To overcome this knowledge gap, the genetic architecture of grass pea resistance to U. pisi was investigated using a worldwide collection of 182 accessions through a genome-wide association approach. The response of the grass pea collection to rust infection under controlled conditions and at the seedling stage did not reveal any hypersensitive response but a continuous variation for disease severity, with the identification of promising sources of partial resistance. A panel of 5,651 high-quality single-nucleotide polymorphism (SNP) markers previously generated was used to test for SNP-trait associations, based on a mixed linear model accounting for population structure. We detected seven SNP markers significantly associated with U. pisi disease severity, suggesting that partial resistance is oligogenic. Six of the associated SNP markers were located in chromosomes 4 and 6, while the remaining SNP markers had no known chromosomal position. Through comparative mapping with the pea reference genome, a total of 19 candidate genes were proposed, encoding for leucine-rich repeat, NB-ARC domain, and TGA transcription factor family, among others. Results presented in this study provided information on the availability of partial resistance in grass pea germplasm and advanced our understanding of the molecular mechanisms of quantitative resistance to rust in grass pea. Moreover, the detected associated SNP markers constitute promising genomic targets for the development of molecular tools to assist disease resistance precision breeding., Peer reviewed

Table S2. Primer sequences used for RT-qPCR analysis., Uromyces pisi ([Pers.] D.C.) Wint. is an important foliar biotrophic pathogen infecting grass pea (Lathyrus sativus L.), compromising their yield stability. To date, few efforts have been made to assess the natural variation in grass pea resistance and to identify the resistance loci operating against this pathogen, limiting its efficient breeding exploitation. To overcome this knowledge gap, the genetic architecture of grass pea resistance to U. pisi was investigated using a worldwide collection of 182 accessions through a genome-wide association approach. The response of the grass pea collection to rust infection under controlled conditions and at the seedling stage did not reveal any hypersensitive response but a continuous variation for disease severity, with the identification of promising sources of partial resistance. A panel of 5,651 high-quality single-nucleotide polymorphism (SNP) markers previously generated was used to test for SNP-trait associations, based on a mixed linear model accounting for population structure. We detected seven SNP markers significantly associated with U. pisi disease severity, suggesting that partial resistance is oligogenic. Six of the associated SNP markers were located in chromosomes 4 and 6, while the remaining SNP markers had no known chromosomal position. Through comparative mapping with the pea reference genome, a total of 19 candidate genes were proposed, encoding for leucine-rich repeat, NB-ARC domain, and TGA transcription factor family, among others. Results presented in this study provided information on the availability of partial resistance in grass pea germplasm and advanced our understanding of the molecular mechanisms of quantitative resistance to rust in grass pea. Moreover, the detected associated SNP markers constitute promising genomic targets for the development of molecular tools to assist disease resistance precision breeding., Peer reviewed

Table S1. Characterization of the grass pea accessions evaluated for the response to U. pisi based on seed color, size, and geographical origin, and respective BLUEs data., Uromyces pisi ([Pers.] D.C.) Wint. is an important foliar biotrophic pathogen infecting grass pea (Lathyrus sativus L.), compromising their yield stability. To date, few efforts have been made to assess the natural variation in grass pea resistance and to identify the resistance loci operating against this pathogen, limiting its efficient breeding exploitation. To overcome this knowledge gap, the genetic architecture of grass pea resistance to U. pisi was investigated using a worldwide collection of 182 accessions through a genome-wide association approach. The response of the grass pea collection to rust infection under controlled conditions and at the seedling stage did not reveal any hypersensitive response but a continuous variation for disease severity, with the identification of promising sources of partial resistance. A panel of 5,651 high-quality single-nucleotide polymorphism (SNP) markers previously generated was used to test for SNP-trait associations, based on a mixed linear model accounting for population structure. We detected seven SNP markers significantly associated with U. pisi disease severity, suggesting that partial resistance is oligogenic. Six of the associated SNP markers were located in chromosomes 4 and 6, while the remaining SNP markers had no known chromosomal position. Through comparative mapping with the pea reference genome, a total of 19 candidate genes were proposed, encoding for leucine-rich repeat, NB-ARC domain, and TGA transcription factor family, among others. Results presented in this study provided information on the availability of partial resistance in grass pea germplasm and advanced our understanding of the molecular mechanisms of quantitative resistance to rust in grass pea. Moreover, the detected associated SNP markers constitute promising genomic targets for the development of molecular tools to assist disease resistance precision breeding., Peer reviewed

Figure S3. Heat map depicting mean log2-fold changes of gene expression levels. Relative gene expression analysis of candidate gene related to U. pisi disease response (A-C) was analyzed in accessions susceptible (PI283574, and PI221467_B) and partially resistant (PI165528, PI283566, and PI577138_A). Relative expression values were normalized to the mean of the non-inoculated control (0 HAI) of the most susceptible accession (PI221467_B). Higher levels of relative gene expression (log2-fold changes > 0) are indicated in red and lower levels (log2-fold changes < 0) are indicated in blue. Small letters represent significant differences (P-value <0.05), among accessions or time-points (0 HAI, 12 HAI, 24 HAI, 48 HAI)., Uromyces pisi ([Pers.] D.C.) Wint. is an important foliar biotrophic pathogen infecting grass pea (Lathyrus sativus L.), compromising their yield stability. To date, few efforts have been made to assess the natural variation in grass pea resistance and to identify the resistance loci operating against this pathogen, limiting its efficient breeding exploitation. To overcome this knowledge gap, the genetic architecture of grass pea resistance to U. pisi was investigated using a worldwide collection of 182 accessions through a genome-wide association approach. The response of the grass pea collection to rust infection under controlled conditions and at the seedling stage did not reveal any hypersensitive response but a continuous variation for disease severity, with the identification of promising sources of partial resistance. A panel of 5,651 high-quality single-nucleotide polymorphism (SNP) markers previously generated was used to test for SNP-trait associations, based on a mixed linear model accounting for population structure. We detected seven SNP markers significantly associated with U. pisi disease severity, suggesting that partial resistance is oligogenic. Six of the associated SNP markers were located in chromosomes 4 and 6, while the remaining SNP markers had no known chromosomal position. Through comparative mapping with the pea reference genome, a total of 19 candidate genes were proposed, encoding for leucine-rich repeat, NB-ARC domain, and TGA transcription factor family, among others. Results presented in this study provided information on the availability of partial resistance in grass pea germplasm and advanced our understanding of the molecular mechanisms of quantitative resistance to rust in grass pea. Moreover, the detected associated SNP markers constitute promising genomic targets for the development of molecular tools to assist disease resistance precision breeding., Peer reviewed

Figure S2. Allele frequency distribution at the DS range scale, of the associated SNPs detected in the GWAS in response to U. pisi. Allele missing data refers to the absence of genotypic data., Uromyces pisi ([Pers.] D.C.) Wint. is an important foliar biotrophic pathogen infecting grass pea (Lathyrus sativus L.), compromising their yield stability. To date, few efforts have been made to assess the natural variation in grass pea resistance and to identify the resistance loci operating against this pathogen, limiting its efficient breeding exploitation. To overcome this knowledge gap, the genetic architecture of grass pea resistance to U. pisi was investigated using a worldwide collection of 182 accessions through a genome-wide association approach. The response of the grass pea collection to rust infection under controlled conditions and at the seedling stage did not reveal any hypersensitive response but a continuous variation for disease severity, with the identification of promising sources of partial resistance. A panel of 5,651 high-quality single-nucleotide polymorphism (SNP) markers previously generated was used to test for SNP-trait associations, based on a mixed linear model accounting for population structure. We detected seven SNP markers significantly associated with U. pisi disease severity, suggesting that partial resistance is oligogenic. Six of the associated SNP markers were located in chromosomes 4 and 6, while the remaining SNP markers had no known chromosomal position. Through comparative mapping with the pea reference genome, a total of 19 candidate genes were proposed, encoding for leucine-rich repeat, NB-ARC domain, and TGA transcription factor family, among others. Results presented in this study provided information on the availability of partial resistance in grass pea germplasm and advanced our understanding of the molecular mechanisms of quantitative resistance to rust in grass pea. Moreover, the detected associated SNP markers constitute promising genomic targets for the development of molecular tools to assist disease resistance precision breeding., Peer reviewed

Figure S1. Quantile-Quantile (QQ) plots of the observed versus expected p-values of the GWAS results related to U. pisi inoculation experiments, using a model accounting for population structure (Eigen), a model accounting for familiar relatedness (Kinship), and a naïve model., Uromyces pisi ([Pers.] D.C.) Wint. is an important foliar biotrophic pathogen infecting grass pea (Lathyrus sativus L.), compromising their yield stability. To date, few efforts have been made to assess the natural variation in grass pea resistance and to identify the resistance loci operating against this pathogen, limiting its efficient breeding exploitation. To overcome this knowledge gap, the genetic architecture of grass pea resistance to U. pisi was investigated using a worldwide collection of 182 accessions through a genome-wide association approach. The response of the grass pea collection to rust infection under controlled conditions and at the seedling stage did not reveal any hypersensitive response but a continuous variation for disease severity, with the identification of promising sources of partial resistance. A panel of 5,651 high-quality single-nucleotide polymorphism (SNP) markers previously generated was used to test for SNP-trait associations, based on a mixed linear model accounting for population structure. We detected seven SNP markers significantly associated with U. pisi disease severity, suggesting that partial resistance is oligogenic. Six of the associated SNP markers were located in chromosomes 4 and 6, while the remaining SNP markers had no known chromosomal position. Through comparative mapping with the pea reference genome, a total of 19 candidate genes were proposed, encoding for leucine-rich repeat, NB-ARC domain, and TGA transcription factor family, among others. Results presented in this study provided information on the availability of partial resistance in grass pea germplasm and advanced our understanding of the molecular mechanisms of quantitative resistance to rust in grass pea. Moreover, the detected associated SNP markers constitute promising genomic targets for the development of molecular tools to assist disease resistance precision breeding., Peer reviewed

Supplementary Table 1: Primer sequences used for RT-qPCR analysis. Supplementary Table 2. Analysis of variance (ANOVA) for the effects of Genotype, Repetition, interaction between Genotype and Repetition. Supplementary Table 3. List of candidate genes underlying Lathyrus cicera pQTLs for rust resistance based on orthologous positions in the Pisum sativum genome., Rusts are among the most important foliar biotrophic fungal diseases in legumes. Lathyrus cicera crop can be severely damaged by Uromyces pisi, to which partial resistance has been identified. Nevertheless, the underlying genetic basis and molecular mechanisms of this resistance are poorly understood in L. cicera. To prioritise the causative variants controlling partial resistance to rust in L. cicera, a recombinant inbred line (RIL) population, segregating for response to this pathogen, was used to combine the detection of related phenotypic- and expression-quantitative trait loci (pQTLs and eQTLs, respectively). RILs’ U. pisi disease severity (DS) was recorded in three independent screenings at seedling (growth chamber) and in one season of exploratory screening at adult plant stage (semi-controlled field conditions). A continuous DS range was observed in both conditions and used for pQTL mapping. Different pQTLs were identified under the growth chamber and semi-controlled field conditions, indicating a distinct genetic basis depending on the plant developmental stage and/or the environment. Additionally, the expression of nine genes related to U. pisi resistance in L. cicera was quantified for each RIL individual and used for eQTL mapping. One cis-eQTL and one trans-eQTL were identified controlling the expression variation of one gene related to rust resistance – a member of glycosyl hydrolase family 17. Integrating phenotyping, gene expression and linkage mapping allowed prioritising four candidate genes relevant for disease-resistance precision breeding involved in adaptation to biotic stress, cellular, and organelle homeostasis, and proteins directly involved in plant defence., Peer reviewed