Identification of Single Nucleotide Polymorphisms and analysis of Linkage Disequilibrium in sunflower elite inbred lines using the candidate gene approach

Background. Association analysis is a powerful tool to identify gene loci that may contribute to phenotypic variation. This includes the estimation of nucleotide diversity, the assessment of linkage disequilibrium structure (LD) and the evaluation of selection processes. Trait mapping by allele association requires a high-density map, which could be obtained by the addition of Single Nucleotide Polymorphisms (SNPs) and short insertion and/or deletions (indels) to SSR and AFLP genetic maps. Nucleotide diversity analysis of randomly selected candidate regions is a promising approach for the success of association analysis and fine mapping in the sunflower genome. Moreover, knowledge of the distance over which LD persists, in agronomically meaningful sunflower accessions, is important to establish the density of markers and the experimental design for association analysis. Results. A set of 28 candidate genes related to biotic and abiotic stresses were studied in 19 sunflower inbred lines. A total of 14,348 bp of sequence alignment was analyzed per individual. In average, 1 SNP was found per 69 nucleotides and 38 indels were identified in the complete data set. The mean nucleotide polymorphism was moderate ( = 0.0056), as expected for inbred materials. The number of haplotypes per region ranged from 1 to 9 (mean = 3.54 1.88). Model-based population structure analysis allowed detection of admixed individuals within the set of accessions examined. Two putative gene pools were identified (G1 and G2), with a large proportion of the inbred lines being assigned to one of them (G1). Consistent with the absence of population sub-structuring, LD for G1 decayed more rapidly (r 2= 0.48 at 643 bp; trend line, pooled data) than the LD trend line for the entire set of 19 individuals (r2= 0.64 for the same distance). Conclusion. Knowledge about the patterns of diversity and the genetic relationships between breeding materials could be an invaluable aid in crop improvement strategies. The relatively high frequency of SNPs within the elite inbred lines studied here, along with the predicted extent of LD over distances of 100 kbp (r2∼0.1) suggest that high resolution association mapping in sunflower could be achieved with marker densities lower than those usually reported in the literature.Fil:Fusari, C.M. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina.Fil:Lia, V.V. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina.Fil:Hopp, H.E. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina.Fil:Heinz, R.A. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina

Discovery and Functional Categorisation of Expressed Sequence Tags from Flowers of \u3cem\u3eEragrostis Curvula\u3c/em\u3e Genotypes Showing Different Ploidy Levels and Reproductive Modes

Two novel genotypes of weeping lovegrass (Eragrostis curvula) - a dihaploid strain obtained in vitro from an apomictic cultivar and a tetraploid plant derived from the dihaploid after chromosome duplication – have recently been developed. These materials represent an excellent system for the identification, through transcriptional profiling, of genes involved in diplospory and/or ploidy level gene regulation. The aim of this work was the discovery and functional classification of expressed sequence tags (ESTs) from immature inflorescences of the apomictic E. curvula cultivar Tanganyika (2n=4x=40), a dihaploid sexual strain derived from it (2n=2x=20) and a tetraploid sexual strain (2n=4x=40) obtained by colchicine duplication of the dihaploid

Optimizing DDRADseq in non-model species: A Case Study in Eucalyptus dunnii Maiden

Restriction site-associated DNA sequencing (RADseq) and its derived protocols, such as double digest RADseq (ddRADseq), offer a flexible and highly cost-effective strategy for efficient plant genome sampling. This has become one of the most popular genotyping approaches for breeding, conservation, and evolution studies in model and non-model plant species. However, universal protocols do not always adapt well to non-model species. Herein, this study reports the development of an optimized and detailed ddRADseq protocol in Eucalyptus dunnii, a non-model species, which combines different aspects of published methodologies. The initial protocol was established using only two samples by selecting the best combination of enzymes and through optimal size selection and simplifying lab procedures. Both single nucleotide polymorphisms (SNPs) and simple sequence repeats (SSRs) were determined with high accuracy after applying stringent bioinformatics settings and quality filters, with and without a reference genome. To scale it up to 24 samples, we added barcoded adapters. We also applied automatic size selection, and therefore obtained an optimal number of loci, the expected SNP locus density, and genome-wide distribution. Reliability and cross-sequencing platform compatibility were verified through dissimilarity coefficients of 0.05 between replicates. To our knowledge, this optimized ddRADseq protocol will allow users to go from the DNA sample to genotyping data in a highly accessible and reproducible way.Instituto de BiotecnologíaFil: Aguirre, Natalia Cristina. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Biotecnología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Filippi, Carla Valeria. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Biotecnología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Zaina, Giusi. University of Udine. Department of Agricultural, Food, Environmental and Animal Sciences; ItaliaFil: Rivas, Juan Gabriel. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Biotecnología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Acuña, Cintia Vanesa. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Biotecnología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Villalba, Pamela Victoria. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Biotecnología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Garcia, Martin Nahuel. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Biotecnología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Gonzalez, Sergio Alberto. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Biotecnología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Rivarola, Maximo Lisandro. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Biotecnología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Martinez, Maria Carolina. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Biotecnología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Puebla, Andrea Fabiana. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Biotecnología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Morgante, Michele. University of Udine. Department of Agricultural, Food, Environmental and Animal Sciences; ItaliaFil: Hopp, Horacio Esteban. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Biotecnología; Argentina. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Paniego, Norma Beatriz. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Biotecnología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Marcucci Poltri, Susana Noemi. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Biotecnología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentin

Population structure and genetic diversity characterization of a sunflower association mapping population using SSR and SNP markers

BACKGROUND: Argentina has a long tradition of sunflower breeding, and its germplasm is a valuable genetic resource worldwide. However, knowledge of the genetic constitution and variability levels of the Argentinean germplasm is still scarce, rendering the global map of cultivated sunflower diversity incomplete. In this study, 42 microsatellite loci and 384 single nucleotide polymorphisms (SNPs) were used to characterize the first association mapping population used for quantitative trait loci mapping in sunflower, along with a selection of allied open-pollinated and composite populations from the germplasm bank of the National Institute of Agricultural Technology of Argentina. The ability of different kinds of markers to assess genetic diversity and population structure was also evaluated.Fil: Filippi, Carla Valeria. Instituto Nacional de Tecnología Agropecuaria. Centro de Investigación en Ciencias Veterinarias y Agronómicas. Instituto de Biotecnología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Aguirre, Natalia Cristina. Instituto Nacional de Tecnología Agropecuaria. Centro de Investigación en Ciencias Veterinarias y Agronómicas. Instituto de Biotecnología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Rivas, Juan Gabriel. Instituto Nacional de Tecnología Agropecuaria. Centro de Investigación en Ciencias Veterinarias y Agronómicas. Instituto de Biotecnología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Zubrzycki, Jeremías Enrique. Instituto Nacional de Tecnología Agropecuaria. Centro de Investigación en Ciencias Veterinarias y Agronómicas. Instituto de Biotecnología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Puebla, Andrea. Instituto Nacional de Tecnología Agropecuaria. Centro de Investigación en Ciencias Veterinarias y Agronómicas. Instituto de Biotecnología; ArgentinaFil: Cordes, Diego. Instituto Nacional de Tecnología Agropecuaria. Centro Regional Córdoba. Estación Experimental Agropecuaria Manfredi; ArgentinaFil: Moreno, Maria V.. Instituto Nacional de Tecnología Agropecuaria. Centro Regional Córdoba. Estación Experimental Agropecuaria Manfredi; ArgentinaFil: Fusari, Corina M.. Instituto Nacional de Tecnología Agropecuaria. Centro de Investigación en Ciencias Veterinarias y Agronómicas. Instituto de Biotecnología; ArgentinaFil: Alvarez, Daniel. Instituto Nacional de Tecnología Agropecuaria. Centro Regional Córdoba. Estación Experimental Agropecuaria Manfredi; ArgentinaFil: Heinz, Ruth Amelia. Instituto Nacional de Tecnología Agropecuaria. Centro de Investigación en Ciencias Veterinarias y Agronómicas. Instituto de Biotecnología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Hopp, Horacio Esteban. Instituto Nacional de Tecnología Agropecuaria. Centro de Investigación en Ciencias Veterinarias y Agronómicas. Instituto de Biotecnología; ArgentinaFil: Paniego, Norma Beatriz. Instituto Nacional de Tecnología Agropecuaria. Centro de Investigación en Ciencias Veterinarias y Agronómicas. Instituto de Biotecnología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Lia, Verónica Viviana. Instituto Nacional de Tecnología Agropecuaria. Centro de Investigación en Ciencias Veterinarias y Agronómicas. Instituto de Biotecnología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentin

Identification of Single Nucleotide Polymorphisms and analysis of Linkage Disequilibrium in sunflower elite inbred lines using the candidate gene approach

Abstract Background Association analysis is a powerful tool to identify gene loci that may contribute to phenotypic variation. This includes the estimation of nucleotide diversity, the assessment of linkage disequilibrium structure (LD) and the evaluation of selection processes. Trait mapping by allele association requires a high-density map, which could be obtained by the addition of Single Nucleotide Polymorphisms (SNPs) and short insertion and/or deletions (indels) to SSR and AFLP genetic maps. Nucleotide diversity analysis of randomly selected candidate regions is a promising approach for the success of association analysis and fine mapping in the sunflower genome. Moreover, knowledge of the distance over which LD persists, in agronomically meaningful sunflower accessions, is important to establish the density of markers and the experimental design for association analysis. Results A set of 28 candidate genes related to biotic and abiotic stresses were studied in 19 sunflower inbred lines. A total of 14,348 bp of sequence alignment was analyzed per individual. In average, 1 SNP was found per 69 nucleotides and 38 indels were identified in the complete data set. The mean nucleotide polymorphism was moderate (θ = 0.0056), as expected for inbred materials. The number of haplotypes per region ranged from 1 to 9 (mean = 3.54 ± 1.88). Model-based population structure analysis allowed detection of admixed individuals within the set of accessions examined. Two putative gene pools were identified (G1 and G2), with a large proportion of the inbred lines being assigned to one of them (G1). Consistent with the absence of population sub-structuring, LD for G1 decayed more rapidly (r2 = 0.48 at 643 bp; trend line, pooled data) than the LD trend line for the entire set of 19 individuals (r2 = 0.64 for the same distance). Conclusion Knowledge about the patterns of diversity and the genetic relationships between breeding materials could be an invaluable aid in crop improvement strategies. The relatively high frequency of SNPs within the elite inbred lines studied here, along with the predicted extent of LD over distances of 100 kbp (r2~0.1) suggest that high resolution association mapping in sunflower could be achieved with marker densities lower than those usually reported in the literature.</p

Stone-working and the earliest steel in Iberia: Scientific analyses and experimental replications of final bronze age stelae and tools

The south-western Iberian stelae from the Final Bronze Age (FBA) and Early Iron Age (EIA) have long been the centre of archaeological interest. These monuments show representations of human and animal figures, objects such as weapons, ornaments and chariots. Moreover, they provide insights into prehistoric stone working and sculpting techniques. On the downside, petrological studies of the rocks and consequent reflections on suitable tools are still the exception. Due to the lack of analysis and technological studies, this research will put emphasis on a multidisciplinary approach involving petrographic and metallurgic analyses, complemented by experimental archaeology. Firstly, an accurate lithological determination of stelae from Capilla (Spain) has been established with petrographic methods. According to the results, two slabs for replications of stelae were obtained from the identified mono-mineral quartz-sandstone (“quartzite”) outcrop. The experiment involved the trial of chisels made from all expedient materials that were available in the FBA-EIA by a professional stonemason. Considering that such extremely hard rocks require robust tools for carving, a long ignored iron chisel from the FBA site of Rocha do Vigio (Portugal) was studied with metallography and exposed heterogeneous, however surprisingly high-carbon steel. The analysed rocks are amongst the hardest that can be used for sculpting, and in the course of the experiment, bronze and lithic tools could be discarded. The only tool that showed an effect was the replication of the steel chisel from Rocha do Vigio with a hardened edge. The distinct work traces were compared to the original monuments. We hypothesize that the production of carbon steel as well as its hardening were possibly already known at the FBA-EIA transition in Iberia. Hence, only the access to iron technology allowed for the making of stelae from the lithotypes that were frequently used in the Zújar basin around the municipality of Capilla.This research received funding from the German Research Foundation (DFG, project AR 1305/2–1).Peer reviewe

Identification of Single Nucleotide Polymorphisms and analysis of Linkage Disequilibrium in sunflower elite inbred lines using the candidate gene approach-1

E data calculated for the G1 subset of individuals identified in the STRUCTURE analysis (HA52, HA61, HA89, HA370, HAR3, HAR5, KLM280, PAC2, RHA266, RHA274, RHA293 and RHA374).<p><b>Copyright information:</b></p><p>Taken from "Identification of Single Nucleotide Polymorphisms and analysis of Linkage Disequilibrium in sunflower elite inbred lines using the candidate gene approach"</p><p>http://www.biomedcentral.com/1471-2229/8/7</p><p>BMC Plant Biology 2008;8():7-7.</p><p>Published online 23 Jan 2008</p><p>PMCID:PMC2266750.</p><p></p

Seed dormancy QTL identification across a Sorghum bicolor segregating population

Pre-harvest sprouting (PHS) in Sorghum bicolor is one of the main constrains for its production in the central region of Argentina, as grain maturation often coincides with rainy or high environmental humidity conditions. The obtention of more dormant genotypes with higher PHS resistance has always been a desirable trait for breeders but the typical quantitative nature of seed dormancy makes its manipulation difficult through classical breeding. Dissecting this quantitative variability into quantitative trait loci (QTL) is a main concern especially in cereal species. In this work, a sorghum segregating population including 190 families was genotyped with microsatellite markers and the SbABI5 candidate gene. A genetic map encompassing 96 markers and a total length of 1331 cM was built. Seed dormancy was phenotyped in F3 and F4 panicles in two contrasting Argentinean environments (Castelar and Manfredi). Six seed dormancy QTL for mature grains were identified (qGI-1, qGI-3, qGI-4, qGI-6, qGI-7 and qGI-9) with the aid of QTL Cartographer and QTLNetwork, three of them (qGI-3, qGI-7 and qGI- 9) being co-localised by both approaches. No epistasis was detected for the identified QTL but QTL-byenvironment interaction was significant for qGI-7 and qGI-9. Interestingly, seed dormancy candidate genes SbABI3/VP1 and SbGA20ox3 were located within qGI-3, which makes them noteworthy candidate genes for this QTL.Inst. de BiotecnologíaFil: Cantoro, Renata. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Fisiológicas y Ecológicas Vinculadas A la Agricultura; Argentina. Universidad de Buenos Aires. Facultad de Agronomía. Departamento de Producción Vegetal. Cátedra de Cultivos Industriales; ArgentinafIL: Fernandez, Luis German. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Biotecnología; ArgentinaFil: Cervigni, Gerardo Domingo Lucio Universidad Nacional de Rosario. Facultad de Ciencias Bioquímicas y Farmacéuticas. Centro de Estudios Fotosintéticos y Bioquímicos; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Rodriguez, María V. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Fisiológicas y Ecológicas Vinculadas A la Agricultura; Argentina. Universidad de Buenos Aires. Facultad de Agronomía. Departamento de Producción Vegetal. Cátedra de Cultivos Industriales; ArgentinaFil: Paniego, Norma Beatriz. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Biotecnología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad Nacional de Buenos Aires. Facultad de Ciencias Exactas y Naturales; ArgentinaFil: Heinz, Ruth Amelia. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Biotecnología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad Nacional de Buenos Aires. Facultad de Ciencias Exactas y Naturales; ArgentinaFil: Gieco, Jorge Omar. Instituto Nacional de Tecnología Agropecuaria (INTA). Estación Experimental Agropecuaria Manfredi; ArgentinaFil: Benech-Arnold, Roberto Luis. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Fisiológicas y Ecológicas Vinculadas a la Agricultura; Argentin

Genetic characterization of sunflower breeding resources from Argentina: assessing diversity in key open-pollinated and composite populations

Open-pollinated (OPs) and composite populations (CPs) represent a valuable resource for sunflower breeding programmes. However, little is known about the levels and distribution of genetic variation within each of these populations. In this study, quantitative and qualitative traits along with molecular markers were used to evaluate 14 populations from the Instituto Nacional de Tecnología Agropecuaria (INTA) sunflower germplasm collection. These populations were chosen to represent historically important accessions that still play a central role within the INTA sunflower breeding programme due to their extensive variation in diverse agronomically important traits. Nine quantitative and eight qualitative agro-morphological descriptors were recorded and compared with those of a larger set of accessions representative of the phenotypic diversity of the sunflower collection. Molecular characterization was conducted on a total of 311 individuals using 16 microsatellite markers. Overall, the average gene diversity was 0.56 and the average number of alleles per locus was 6.25. No statistically significant differences in genetic diversity were detected between the OPs and CPs. Global estimates of F ST revealed very high levels of differentiation among accessions (F ST= 0.413, P< 0.05). Population structure analyses were consistent with the observed levels of differentiation and identified two major groups. The results of this work show that high global diversity is preserved within the accessions analysed here. Additionally, this study provides a set of reliable and discriminant markers for the cost-effective molecular characterization of sunflower accessions, along with the guidelines for the delineation of sampling strategies for OPs and CPs, thus aiding the efficient management and exploitation of sunflower germplasm collections.Fil: Moreno, M. V.. Instituto Nacional de Tecnología Agropecuaria. Centro Regional Córdoba. Estación Experimental Agropecuaria Manfredi; ArgentinaFil: Nishinakamasu, V.. Instituto Nacional de Tecnología Agropecuaria. Centro de Investigación en Ciencias Veterinarias y Agronómicas. Instituto de Biotecnología; ArgentinaFil: Loray, M. A.. Instituto Nacional de Semillas; ArgentinaFil: Alvarez, D.. Instituto Nacional de Tecnología Agropecuaria. Centro Regional Córdoba. Estación Experimental Agropecuaria Manfredi; ArgentinaFil: Gieco, J.. Instituto Nacional de Tecnología Agropecuaria. Centro Regional Córdoba. Estación Experimental Agropecuaria Manfredi; ArgentinaFil: Vicario, A.. Instituto Nacional de Semillas; ArgentinaFil: Hopp, Horacio Esteban. Instituto Nacional de Tecnología Agropecuaria. Centro de Investigación en Ciencias Veterinarias y Agronómicas. Instituto de Biotecnología; ArgentinaFil: Heinz, Ruth Amelia. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Instituto Nacional de Tecnología Agropecuaria. Centro de Investigación en Ciencias Veterinarias y Agronómicas. Instituto de Biotecnología; ArgentinaFil: Paniego, Norma Beatriz. Instituto Nacional de Tecnología Agropecuaria. Centro de Investigación en Ciencias Veterinarias y Agronómicas. Instituto de Biotecnología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Lia, Verónica Viviana. Instituto Nacional de Tecnología Agropecuaria. Centro de Investigación en Ciencias Veterinarias y Agronómicas. Instituto de Biotecnología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentin