A DNA test for routine prediction in breeding of sweet cherry fruit color,Pav-R<Subscript>f</Subscript>-SSR

Sweet cherry fruit color is a market class-defining trait. The two main market classes in the USA are mahogany, consisting fruit with red skin and flesh, and blush, consisting clear-fleshed fruit with yellow skin and a red overcolor on less than the entire skin surface. Fruit color is a major consideration in sweet cherry breeding as resources and selection thresholds are often differentially applied to each market class. The use of DNA-based information could improve breeding efficiency and accuracy for fruit color, but a predictive DNA test is required. The objective of this study was to develop a reliable, simple DNA test for the prediction of sweet cherry color-based market classes, targeting the major locus, termed here as R _f , associated with fruit color variation. Haplotypes were developed based on 14 SNP markers from the RosBREED cherry 6K SNP array v1 that were associated with the two market classes. To convert the multiple SNP markers to a single, simple PCR-based assay, 11 PCR-based assays targeting microsatellite motifs were designed, using the peach reference genome sequence, and used to screen 20 individuals representing the most common SNP haplotypes. One assay, subsequently named Pav-R_f-SSR, was used to screen 221 phenotyped individuals of the RosBREED sweet cherry reference germplasm set and accurately differentiated individuals with mahogany and blush fruits. Pav-R_f-SSR can be used in DNA-informed breeding schemes to efficiently and accurately predict genetic potential for fruit color and is one of the first DNA tests publicly available for a sweet cherry fruit quality trait.     

Whole-Genome Analysis of Diversity and SNP-Major Gene Association in Peach Germplasm

Peach was domesticated in China more than four millennia ago and from there it spread world-wide. Since the middle of the last century, peach breeding programs have been very dynamic generating hundreds of new commercial varieties, however, in most cases such varieties derive from a limited collection of parental lines (founders). This is one reason for the observed low levels of variability of the commercial gene pool, implying that knowledge of the extent and distribution of genetic variability in peach is critical to allow the choice of adequate parents to confer enhanced productivity, adaptation and quality to improved varieties. With this aim we genotyped 1,580 peach accessions (including a few closely related Prunus species) maintained and phenotyped in five germplasm collections (four European and one Chinese) with the International Peach SNP Consortium 9K SNP peach array. The study of population structure revealed the subdivision of the panel in three main populations, one mainly made up of Occidental varieties from breeding programs (POP1OC_B), one of Occidental landraces (POP2OC_T) and the third of Oriental accessions (POP3OR). Analysis of linkage disequilibrium (LD) identified differential patterns of genome-wide LD blocks in each of the populations. Phenotypic data for seven monogenic traits were integrated in a genome-wide association study (GWAS). The significantly associated SNPs were always in the regions predicted by linkage analysis, forming haplotypes of markers. These diagnostic haplotypes could be used for marker-assisted selection (MAS) in modern breeding programs.     

Identifying SNP markers tightly associated with six major genes in peach [<Emphasis Type="Italic">Prunus persica</Emphasis> (L.) Batsch] using a high-density SNP array with an objective of marker-assisted selection (MAS)

One of the applications of genomics is to identify genetic markers linked to loci responsible for variation in phenotypic traits, which could be used in breeding programs to select individuals with favorable alleles, particularly at the seedling stage. With this aim, in the framework of the European project FruitBreedomics, we selected five main peach fruit characters and a resistance trait, controlled by major genes with Mendelian inheritance: fruit flesh color Y, fruit skin pubescence G, fruit shape S, sub-acid fruit D, stone adhesion-flesh texture F-M, and resistance to green peach aphid Rm2. They were all previously mapped in Prunus. We then selected three F₁ and three F₂ progenies segregating for these characters and developed genetic maps of the linkage groups including the major genes, using the single nucleotide polymorphism (SNP) genome-wide scans obtained with the International Peach SNP Consortium (IPSC) 9K SNP array v1. We identified SNPs co-segregating with the characters in all cases. Their positions were in agreement with the known positions of the major genes. The number of SNPs linked to each of these, as well as the size of the physical regions encompassing them, varied depending on the maps. As a result, the number of useful SNPs for marker-assisted selection varied accordingly. As a whole, this study establishes a sound basis for further development of MAS on these characters. Additionally, we also discussed some limitations that were observed regarding the SNP array efficiency.     

Identification of SNPs in Closely Related Temperate Japonica Rice Cultivars Using Restriction Enzyme-Phased Sequencing

Very low polymorphism in the germplasm typically used by breeding programs poses a significant bottleneck with regards to molecular breeding and the exploitation of breeding materials for quantitative trait analyses. California rice cultivars, derived from a very small base of temperate japonica germplasm and having a relatively brief breeding history, are a good example. In this study, we employed a reduced representation sequencing approach called Restriction Enzyme Site Comparative Analysis (RESCAN) to simultaneously identify and genotype single nucleotide polymorphisms (SNPs) in forty-five rice cultivars representing the majority of the 100 year-old breeding history in California. Over 20,000 putative SNPs were detected relative to the Nipponbare reference genome which enabled the identification and analysis of inheritance of pedigree haplotypes. Haplotype blocks distinguishing modern California cultivars from each other and from the ancestral short grain temperate japonica cultivars were easily identified. Reduced representation sequencing methods such as RESCAN are a valuable alternative to SNP chip genotyping and low coverage whole genome sequencing.     

Application of Genomic and Quantitative Genetic Tools to Identify Candidate Resistance Genes for Brown Rot Resistance in Peach

The availability of a complete peach genome assembly and three different peach genome sequences created by our group provide new opportunities for application of genomic data and can improve the power of the classical Quantitative Trait Loci (QTL) approaches to identify candidate genes for peach disease resistance. Brown rot caused by Monilinia spp., is the most important fungal disease of stone fruits worldwide. Improved levels of peach fruit rot resistance have been identified in some cultivars and advanced selections developed in the UC Davis and USDA breeding programs. Whole genome sequencing of the Pop-DF parents lead to discovery of high-quality SNP markers for QTL genome scanning in this experimental population. Pop-DF created by crossing a brown rot moderately resistant cultivar ‘Dr. Davis’ and a brown rot resistant introgression line, ‘F8,1–42’, derived from an initial almond × peach interspecific hybrid, was evaluated for brown rot resistance in fruit of harvest maturity over three seasons. Using the SNP linkage map of Pop-DF and phenotypic data collected with inoculated fruit, a genome scan for QTL identified several SNP markers associated with brown rot resistance. Two of these QTLs were placed on linkage group 1, covering a large (physical) region on chromosome 1. The genome scan for QTL and SNP effects predicted several candidate genes associated with disease resistance responses in other host-pathogen systems. Two potential candidate genes, ppa011763m and ppa026453m, may be the genes primarily responsible for M. fructicola recognition in peach, activating both PAMP-triggered immunity (PTI) and effector-triggered immunity (ETI) responses. Our results provide a foundation for further genetic dissection, marker assisted breeding for brown rot resistance, and development of peach cultivars resistant to brown rot.     

QTL mapping and breeding value estimation through pedigree-based analysis of fruit size and weight in four diverse peach breeding programs

The narrow genetic base of peach (Prunus persica L. Batsch) challenges efforts to accurately dissect the genetic architecture of complex traits. Standardized phenotypic assessment of pedigree-linked breeding germplasm and new molecular strategies and analytical approaches developed and conducted during the RosBREED project for enabling marker-assisted breeding (MAB) in Rosaceae crops has overcome several aspects of this challenge. The genetic underpinnings of fruit size (fruit equatorial diameter (FD)) and weight (fresh weight (FW)), two most important components of yield, were investigated using the pedigree-based analysis (PBA) approach under a Bayesian framework which has emerged as an alternative strategy to study the genetics of quantitative traits within diverse breeding germplasm across breeding programs. In this study, a complex pedigree with the common founder “Orange Cling” was identified and FD and FW data from 2011 and 2012 analyzed. A genetic model including genetic additive and dominance effects was considered, and its robustness was evaluated by using various prior and initial values in the Markov chain Monte Carlo procedure. Five QTLs were identified which accounted for up to 29 and 17 % of the phenotypic variation for FD and FW, respectively. Additionally, genomic breeding values were obtained for both traits, with accuracies >85 %. This approach serves as a model study for performing PBA across diverse pedigrees. By incorporating multiple breeding programs, the method and results presented support and highlight the ability of this strategy to identify genomic resources as targets for DNA marker development and subsequent MAB within each program.     

Genetic structure of a QTL hotspot on chromosome 2 in sweet cherry indicates positive selection for favorable haplotypes

A genomic region of particular interest for sweet cherry (Prunus avium L.) breeding is a quantitative trait locus (QTL) “hotspot” on chromosome 2. QTLs for fruit size, firmness, sweetness, and flowering time are reported to map to this region. An understanding of genetic diversity, allele sources, linkage relationships, and historical recombinations is critical to enable the combining of favorable alleles at multiple loci. The objectives of this study were to characterize, visualize, and interpret the genetic structure of this previously identified QTL hotspot within North American sweet cherry breeding germplasm, using a pedigree-based haploblocking approach. Across the 29.4 cM (6.3 Mbp) region defined by single nucleotide polymorphism (SNP) information from the RosBREED cherry 6K SNP array v1, a total of 12 recombination events falling into six inter-marker regions were traced within the pedigree of elite and wild germplasm (n = 55). These recombinations defined five haploblocks containing 5–15 markers and exhibiting 7–11 haplotypes each. Over the entire QTL hotspot, 30 extended haplotypes were identified for which parental gametes could be determined. When the haploblocks and their haplotypes were used to explore genetic diversity, ancestry, and recombination patterns, and then integrated with previous QTL results for fruit size, the results indicated that favorable alleles at this QTL hotspot are under positive selection in breeding. The genetic framework provided by a haploblock approach and knowledge of haplotype-level diversity sets the stage for assigning breeding utility to these haplotypes.     

Development and evaluation of a 9K SNP array for peach by internationally coordinated SNP detection and validation in breeding germplasm

Although a large number of single nucleotide polymorphism (SNP) markers covering the entire genome are needed to enable molecular breeding efforts such as genome wide association studies, fine mapping, genomic selection and marker-assisted selection in peach [Prunus persica (L.) Batsch] and related Prunus species, only a limited number of genetic markers, including simple sequence repeats (SSRs), have been available to date. To address this need, an international consortium (The International Peach SNP Consortium; IPSC) has pursued a coordinated effort to perform genome-scale SNP discovery in peach using next generation sequencing platforms to develop and characterize a high-throughput Illumina Infinium® SNP genotyping array platform. We performed whole genome re-sequencing of 56 peach breeding accessions using the Illumina and Roche/454 sequencing technologies. Polymorphism detection algorithms identified a total of 1,022,354 SNPs. Validation with the Illumina GoldenGate® assay was performed on a subset of the predicted SNPs, verifying ∼75% of genic (exonic and intronic) SNPs, whereas only about a third of intergenic SNPs were verified. Conservative filtering was applied to arrive at a set of 8,144 SNPs that were included on the IPSC peach SNP array v1, distributed over all eight peach chromosomes with an average spacing of 26.7 kb between SNPs. Use of this platform to screen a total of 709 accessions of peach in two separate evaluation panels identified a total of 6,869 (84.3%) polymorphic SNPs.The almost 7,000 SNPs verified as polymorphic through extensive empirical evaluation represent an excellent source of markers for future studies in genetic relatedness, genetic mapping, and dissecting the genetic architecture of complex agricultural traits. The IPSC peach SNP array v1 is commercially available and we expect that it will be used worldwide for genetic studies in peach and related stone fruit and nut species.     

The polymorphisms of bovine <Emphasis Type="Italic">VEGF</Emphasis> gene and their associations with growth traits in Chinese cattle

PCR-SSCP and DNA sequencing methods were employed to screen the genetic variation of VEGF gene in 671 individuals belonging to three Chinese indigenous cattle breeds including Nanyang, Jiaxian Red and Qinchuan. Three haplotypes (A, B and C), four observed genotypes (AA, AB, BB and AC) and three new SNPs (6765T>C ss130456744, 6860A>G ss130456745, 6893T>C ss130456746) were detected. The analysis suggested that one SNP (ss130456744) in the bovine VEGF gene had significant effects on birth weight, body weight and heart girth at 6 months old in the Nanyang breed (P < 0.05). The results showed that the SNP (ss130456744) in intron 2 of the VEGF gene is associated with early development and growth of Chinese cattle. These findings raise hope that this polymorphism can be a molecular breeding marker in breeding strategies through marker assisted selection (MAS) in Chinese domestic cattle.     

Gene-associated single nucleotide polymorphism discovery in perennial ryegrass (Lolium perenne L.)

Molecular genetic marker development in perennial ryegrass has largely been dependent on anonymous sequence variation. The availability of a large-scale EST resource permits the development of functionally-associated genetic markers based on SNP variation in candidate genes. Genic SNP loci and associated haplotypes are suitable for implementation in molecular breeding of outbreeding forage species. Strategies for in vitro SNP discovery through amplicon cloning and sequencing have been designed and implemented. Putative SNPs were identified within and between the parents of the F₁(NA₆ × AU₆) genetic mapping family and were validated among progeny individuals. Proof-of-concept for the process was obtained using the drought tolerance-associated LpASRa2 gene. SNP haplotype structures were determined and correlated with predicted amino acid changes. Gene-length LD was evaluated across diverse germplasm collections. A survey of SNP variation across 100 candidate genes revealed a high frequency of SNP incidence (c. 1 per 54 bp), with similar proportions in exons and introns. A proportion (c. 50%) of the validated genic SNPs were assigned to the F₁(NA₆ × AU₆) genetic map, showing high levels of coincidence with previously mapped RFLP loci. The perennial ryegrass SNP resource will enable genetic map integration, detailed LD studies and selection of superior allele content during varietal development.     

Two loss-of-function alleles of the glutathione S-transferase (GST) gene cause anthocyanin deficiency in flower and fruit skin of peach (Prunus persica)

Flower and fruit colors are important agronomic traits. To date, there is no forward genetic evidence that the glutathione S-transferase (GST) gene is responsible for the white flower color in peach (Prunus persica). In this study, genetic analysis indicated that the white-flower trait is monogenetic, is recessive to the non-white allele, and shows pleiotropic effects with non-white-flowered types. The genetic locus underpinning this trait was mapped onto chromosome 3 between 0.421951 and 3.227115 Mb by using bulked segregant analysis in conjunction with whole-genome sequencing, and was further mapped between 0 and 1.178149 Mb by using the backcross 1 (BC₁) population. Finally, the locus was fine-mapped within 535.974- and 552.027-kb intervals by using 151 F₂ individuals and 75 individuals from a BC₁ self-pollinated (BC₁S₁) population, respectively. Pp3G013600, encoding a GST that is known to transport anthocyanin, was identified within the mapping interval. The analysis of genome sequence data showed Pp3G013600 in white flowers has a 2-bp insertion or a 5-bp deletion in the third exon. These variants likely render the GST non-functional because of early stop codons that reduce the protein length from 215 amino acids to 167 and 175 amino acids, respectively. Genetic markers based on these variants validated a complete correlation between the GST loss-of-function alleles and white flower in 128 peach accessions. This correlation was further confirmed by silencing of Pp3G013600 using virus-induced gene silencing technology, which reduced anthocyanin accumulation in peach fruit. The new knowledge from this study is useful for designing peach breeding programs to generate cultivars with white flower and fruit skin.     

桃种质资源亲缘关系的研究进展

种质资源是现代育种和生物技术研究的物质基础。桃种质资源亲缘关系的研究将为探讨桃的起源、进化、分类、育种和资源利用提供科学依据。本文从形态学、细胞学、孢粉学、生物化学及DNA分子标记等几个方面综述了桃种质资源亲缘关系的研究进展,探讨了桃种质资源亲缘关系的研究现状、前景及尚需解决的问题,并就进一步开展桃种质资源亲缘关系的研究进行了分析,提出桃种质野生种、近缘野生种厦不同栽培品种群间的分子系统学关系是今后研究的重点。     

A strategy for developing representative germplasm sets for systematic QTL validation,demonstrated for apple,peach, and sweet cherry

Horticultural crop improvement would benefit from a standardized, systematic, and statistically robust procedure for validating quantitative trait loci (QTLs) in germplasm relevant to breeding programs. Here, we describe and demonstrate a strategy for developing reference germplasm sets of perennial, clonally propagated crops, especially those with long juvenile periods. Germplasm is chosen to efficiently represent important members of larger pedigree-connected genepools. To facilitate validation of multiple QTLs, genome-wide representation of alleles is optimized for designated important breeding parents (IBPs) by estimating average allelic representation in relatives. The strategy and arising principles were demonstrated in a simulated germplasm set. Strong statistical power can be achieved with a carefully chosen germplasm set composed of IBPs, their numerous unselected progenies and close relatives, and all available founders and intermediate ancestors. Crop Reference Sets were developed in the marker-assisted breeding (MAB)-enabling “RosBREED” project as a base resource for QTL validation in US breeding germplasm of apple (Malus × domestica), peach (Prunus persica), and sweet cherry (Prunus avium) consisting of 467, 452, and 268 individuals, respectively. These sets adequately represent the most designated IBPs, have distinct advantages for QTL validation over other germplasm arrangements of equal size, and are recommended as a base resource for QTL validation by breeders of these US crops. The strategy described here can be used to develop efficient reference germplasm sets suiting other breeding genepools or to calculate the statistical power for QTL validation of germplasm sets already established.     

Mapping quantitative trait loci associated with blush in peach [Prunus persica (L.) Batsch]

Blush is an important trait for marketing peaches. The red skin pigmentation develops through the flavonoid and anthocyanin pathways, and both genetic and environmental stimuli, and their interaction, control the regulation of these pathways. The molecular basis of blush development in peach is yet to be understood. An F₂ blush population (ZC²) derived from a cross between two peach cultivars with contrasting phenotypes for blush, “Zin Dai” (～30 % red) and “Crimson Lady” (～100 % red), was used for linkage map construction and quantitative trait loci (QTLs) mapping. The segregating population was phenotyped for blush for 4 years using a visual rating scale and quantified using a colorimeter (L*, a*, and b*) 1 year. The ZC² population was genotyped with the IPSC 9 K peach single-nucleotide polymorphism (SNP) array v1, and a high-density ZC² genetic linkage map was constructed. The map covers genetic a distance of ～452.51 cM with an average marker spacing of 2.38 cM/marker. Four QTLs were detected: one major QTL on LG3 (Blush.Pp.ZC-3.1) and three minor QTLs on LG 4 and 7 (Blush.Pp.ZC-4.1; Blush.Pp.ZC-4.2; Blush.Pp.ZC-7.1), indicating the presence of major and minor genes involved in blush development. Candidate genes involved in skin and flesh coloration of peach (PprMYB10), cherry (PavMYB10), and apple (MdMYB1/MdMYBA/MdMYB10) are located within the interval of the major QTL on LG3, suggesting the same genetic control for color development in the Rosaceae family. Marker-assisted selection (MAS) for blush is discussed.     

Characterization of genetic diversity and population structure in wheat using array based SNP markers

Genetic diversity is crucial for successful adaptation and sustained improvement in crops. India is bestowed with diverse agro-climatic conditions which makes it rich in wheat germplasm adapted to various niches. Germplasm repository consists of local landraces, trait specific genetic stocks including introgressions from wild relatives, exotic collections, released varieties, and improved germplasm. Characterization of genetic diversity is done using morpho-physiological characters as well as by analyzing variations at DNA level. However, there are not many reports on array based high throughput SNP markers having characteristics of genome wide coverage employed in Indian spring wheat germplasm. Amongst wheat SNP arrays, 35K Axiom Wheat Breeder’s Array has the highest SNP polymorphism efficiency suitable for genetic mapping and genetic diversity characterization. Therefore, genotyping was done using 35K in 483 wheat genotypes resulting in 14,650 quality filtered SNPs, that were distributed across the B (~?50%), A (~?39%), and D (~?10%) genomes. The total genetic distance coverage was 4477.85 cM with 3.27 SNP/cM and 0.49 cM/SNP as average marker density and average inter-marker distance, respectively. The PIC ranged from 0.09 to 0.38 with an average of 0.29 across genomes. Population structure and Principal Coordinate Analysis resulted in two subpopulations (SP1 and SP2). The analysis of molecular variance revealed the genetic variation of 2% among and 98% within subpopulations indicating high gene flow between SP1 and SP2. The subpopulation SP2 showed high level of genetic diversity based on genetic diversity indices viz. Shannon’s information index (I)?=?0.648, expected heterozygosity (He)?=?0.456 and unbiased expected heterozygosity (uHe)?=?0.456. To the best of our knowledge, this study is the first to include the largest set of Indian wheat genotypes studied exclusively for genetic diversity. These findings may serve as a potential source for the identification of uncharacterized QTL/gene using genome wide association studies and marker assisted selection in wheat breeding programs.

     

Development of an integrated 200K SNP genotyping array and application for genetic mapping,genome assembly improvement and genome wide association studies in pear (Pyrus)

Pear (Pyrus; 2n = 34), the third most important temperate fruit crop, has great nutritional and economic value. Despite the availability of many genomic resources in pear, it is challenging to genotype novel germplasm resources and breeding progeny in a timely and cost‐effective manner. Genotyping arrays can provide fast, efficient and high‐throughput genetic characterization of diverse germplasm, genetic mapping and breeding populations. We present here 200K AXIOM^® PyrSNP, a large‐scale single nucleotide polymorphism (SNP) genotyping array to facilitate genotyping of Pyrus species. A diverse panel of 113 re‐sequenced pear genotypes was used to discover SNPs to promote increased adoption of the array. A set of 188 diverse accessions and an F₁ population of 98 individuals from ‘Cuiguan’ × ‘Starkrimson’ was genotyped with the array to assess its effectiveness. A large majority of SNPs (166 335 or 83%) are of high quality. The high density and uniform distribution of the array SNPs facilitated prediction of centromeric regions on 17 pear chromosomes, and significantly improved the genome assembly from 75.5% to 81.4% based on genetic mapping. Identification of a gene associated with flowering time and candidate genes linked to size of fruit core via genome wide association studies showed the usefulness of the array in pear genetic research. The newly developed high‐density SNP array presents an important tool for rapid and high‐throughput genotyping in pear for genetic map construction, QTL identification and genomic selection.     

High density SNP mapping and QTL analysis for fruit quality characteristics in peach (Prunus persica L.)

Single nucleotide polymorphisms (SNPs) were used to construct an integrated SNP linkage map of peach (Prunus persica (L.) Batsch). A set of 1,536 SNPs were evaluated with the GoldenGate® Genotyping assay in two mapping populations, Pop-DF, and Pop-DG. After genotyping and filtering, a final set of 1,400 high quality SNPs in Pop-DF and 962 in Pop-DG with full map coverage were selected and used to construct two linkage maps with JoinMap®4.0. The Pop-DF map covered 422 cM of the peach genome and included 1,037 SNP markers, and Pop-DG map covered 369 cM and included 738 SNPs. A consensus map was constructed with 588 SNP markers placed in eight linkage groups (n?=?8 for peach), with map coverage of 454 cM and an average distance of 0.81 cM/marker site. Placements of SNPs on the “peach v1.0” physical map were compared to placement on the linkage maps and several differences were observed. Using the SNP linkage map of Pop-DG and phenotypic data collected for three harvest seasons, a QTL analysis for fruit quality traits and chilling injury symptoms was carried out with the mapped SNPs. Significant QTL effects were detected for mealiness (M) and flesh bleeding (FBL) QTLs on linkage group 4 and flesh browning (FBr) on linkage group 5. This study represents one of the first examples of QTL detection for quality traits and chilling injury symptoms using a high-density SNP map in a single peach F1 family.     

Polymorphism analysis of genomic regions associated with broad-spectrum effective blast resistance genes for marker development in rice

Cultivated European rice germplasm is generally characterized by moderate to high sensitivity to blast, and blast resistance is therefore one of the most important traits to improve in rice breeding. We collected a panel of 25 rice genotypes containing 13 broad range rice resistance genes that are commonly used in breeding programs around the world: Pi1, Pi2, Pi5, Pi7, Pi9, Pi33, Pib, Pik, Pik-p, Pita, Pita ² , Piz and Piz-t. The efficiency of the selected Pi genes towards Italian blast pathotypes was tested via artificial inoculation and under natural field infection conditions. To characterize haplotypes present in the chromosomal regions of the blast resistance genes, a polymorphism search was conducted in the sequence regions adjacent to the blast resistance by examining DNA from the Pi gene donors with a panel of 5–7 potential receivers (cultivated European rice genotypes). Seven InDel and 8 presence/absence polymorphisms were directly detected by gel analysis after DNA amplification, while sequencing of 12.870 bp through 32 loci in different genotypes revealed 85 SNP (one SNP every 151 bp). Seven SSRs were additionally tested revealing 5 polymorphic markers between donors and receivers. Polymorphisms were used to develop 35 PCR-based molecular markers suitable for introgressing of Pi genes into a set of the European rice germplasm. For this last purpose, allelic molecular marker variation was evaluated within a representative collection of about 95 rice genotypes. Polymorphic combinations allowing introgression of the broad spectrum resistance genes into a susceptible genetic background have been identified, thus confirming the potential of the identified markers for molecular-assisted breeding.