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1. Development of whole-genome multiplex assays and construction of an integrated genetic map using SSR markers in Senegalese sole

Author

Israel Guerrero-Cózar, María Jesús Zamorano, Pedro Seoane, Cathaysa Pérez-García, Fernando Cruz, M. Gonzalo Claros, Hicham Benzekri, Manuel Manchado, Marta Gut, Juan José García Sánchez, [Guerrero-Cózar,I, Manchado,M] IFAPA Centro El Toruño, Junta de Andalucía, El Puerto de Santa María, Cádiz, Spain. [Perez-Garcia,C, Zamorano,MJ] Aquaculture Research Group (GIA), IU-ECOAQUA, Universidad de Las Palmas de Gran Canaria, Telde, Spain. [Benzekri,H, Seoane,P, Claros,MG] Department of Molecular Biology and Biochemistry, Universidad de Málaga, Málaga, Spain. [Sánchez,JJ] Instituto Nacional de Toxicología Y Ciencias Forenses (INT), La Laguna, Sta. Cruz de Tenerife, Spain. [Cruz,F, Gut,M] CNAG‑CRG, Centre for Genomic Regulation (CRG), The Barcelona Institute of Science and Technology (BIST), Barcelona, Spain. [Claros,MG] CIBER de Enfermedades Raras (CIBERER), Málaga, Spain. [Claros,MG] Institute of Biomedical Research in Málaga (IBIMA), IBIMA-RARE, Málaga, Spain. [Claros,MG] Instituto de Hortofruticultura Subtropical Y Mediterránea (IHSM-UMA-CSIC), Málaga, Spain. [Manchado,M] 'Crecimiento Azul', Centro IFAPA El Toruño, Unidad Asociada Al CSIC', El Puerto de Sta. María, Spain., and This study was funded by project RTA2017-00054-C03-01, RTA2017-00054-C03-02 and RTA2017-00054-C03-03 funded from MCIU/AEI/FEDER, UE. IGC is funded by a predoctoral fellowship from INIA.

Subjects
0106 biological sciences, 0301 basic medicine, Genetic Linkage, Multiplex polymerase chain reaction, Analytical, Diagnostic and Therapeutic Techniques and Equipment::Investigative Techniques::Genetic Techniques::Pedigree [Medical Subject Headings], Llenguados, 01 natural sciences, Genome, Organisms::Eukaryota::Animals [Medical Subject Headings], Multiplex, Phylogeny, Multidisciplinary, Mapeo cromosómico, Contig, Organisms::Eukaryota::Animals::Chordata::Vertebrates::Fishes::Flatfishes [Medical Subject Headings], Femenino, Chromosome Mapping, food and beverages, Sintenía, Pedigree, Flatfishes, Microsatellite, Medicine, Female, Analytical, Diagnostic and Therapeutic Techniques and Equipment::Investigative Techniques::Genetic Techniques::Chromosome Mapping::Physical Chromosome Mapping [Medical Subject Headings], Phenomena and Processes::Genetic Phenomena::Genetic Structures::Genome::Genome Components::Tandem Repeat Sequences::Microsatellite Repeats [Medical Subject Headings], Science, Computational biology, Biology, Variación genética, 010603 evolutionary biology, Synteny, Article, 03 medical and health sciences, Phenomena and Processes::Genetic Phenomena::Genetic Linkage [Medical Subject Headings], Linaje, Reacción en cadena de la polimerasa multiplex, Animals, Genetic variation, Genotyping, Genoma, Animal breeding, Genetic diversity, Analytical, Diagnostic and Therapeutic Techniques and Equipment::Investigative Techniques::Genetic Techniques::Genetic Association Studies::Genome-Wide Association Study [Medical Subject Headings], Phenomena and Processes::Genetic Phenomena::Sequence Homology::Sequence Homology, Nucleic Acid::Synteny [Medical Subject Headings], Genòmica, 030104 developmental biology, Genetic marker, Phenomena and Processes::Genetic Phenomena::Phylogeny [Medical Subject Headings], Genetic markers, Peces planos, Genètica, Genome-Wide Association Study, Microsatellite Repeats
Abstract

The Senegalese sole (Solea senegalensis) is an economically important flatfish species. In this study, a genome draft was analyzed to identify microsatellite (SSR) markers for whole-genome genotyping. A subset of 224 contigs containing SSRs were preselected and validated by using a de novo female hybrid assembly. Overall, the SSR density in the genome was 886.7 markers per megabase of genomic sequences and the dinucleotide motif was the most abundant (52.4%). In silico comparison identified a set of 108 SSRs (with di-, tetra- or pentanucleotide motifs) widely distributed in the genome and suitable for primer design. A total of 106 markers were structured in thirteen multiplex PCR assays (with up to 10-plex) and the amplification conditions were optimized with a high-quality score. Main genetic diversity statistics and genotyping reliability were assessed. A subset of 40 high polymorphic markers were selected to optimize four supermultiplex PCRs (with up to 11-plex) for pedigree analysis. Theoretical exclusion probabilities and real parentage allocation tests using parent-offspring information confirmed their robustness and effectiveness for parental assignment. These new SSR markers were combined with previously published SSRs (in total 229 makers) to construct a new and improved integrated genetic map containing 21 linkage groups that matched with the expected number of chromosomes. Synteny analysis with respect to C. semilaevis provided new clues on chromosome evolution in flatfish and the formation of metacentric and submetacentric chromosomes in Senegalese sole. This study was funded by project RTA2017-00054-C03-01, RTA2017-00054-C03-02 and RTA2017-00054-C03-03 funded from MCIU/AEI/FEDER, UE. IGC is funded by a predoctoral fellowship from INIA

Published
2020

2. Chromosome anchoring in Senegalese sole (Solea senegalensis) reveals sex-associated markers and genome rearrangements in flatfish

Author

M. Gonzalo Claros, Concha Berbel, Manuel Manchado, Juan F. Martinez-Blanch, Jèssica Gómez-Garrido, Israel Guerrero-Cózar, Pierre-Alexandre Gagnaire, Tyler Alioto, Institut des Sciences de l'Evolution de Montpellier (UMR ISEM), Centre de Coopération Internationale en Recherche Agronomique pour le Développement (Cirad)-École Pratique des Hautes Études (EPHE), Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Université de Montpellier (UM)-Institut de recherche pour le développement [IRD] : UR226-Centre National de la Recherche Scientifique (CNRS), École pratique des hautes études (EPHE), Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Université de Montpellier (UM)-Centre de Coopération Internationale en Recherche Agronomique pour le Développement (Cirad)-Centre National de la Recherche Scientifique (CNRS)-Institut de recherche pour le développement [IRD] : UR226, Guerrero-Cózar, Israel, Gómez-Garrido, Jèssica, Berbel, Concha, Martinez-Blanch, Juan F., Alioto, Tyler, Claros, M. Gonzalo, Gagnaire, Pierre-Alexandre, Manchado, Manuel, [Guerrero-Cózar,I, Berbel,C, Manchado,M] IFAPA Centro El Toruño, Junta de Andalucía, Puerto de Santa María, Cádiz, Spain. [Gomez-Garrido,J, Alioto,T] CNAG-CRG, Centre for Genomic Regulation (CRG), Barcelona Institute of Science and Technology (BIST), Barcelona, Spain. [Martinez-Blanch,JF] Biopolis S.L.-ADM, Parc Cientifc Universidad De Valencia, Paterna, Spain. [Alioto,T] Universitat Pompeu Fabra (UPF), Barcelona, Spain. [Claros,MG] Department of Molecular Biology and Biochemistry, Universidad de Málaga, Málaga, Spain. [Claros,MG] CIBER de Enfermedades Raras (CIBERER), Málaga, Spain. [Claros,MG] Institute of Biomedical Research in Málaga (IBIMA), IBIMA-RARE, Málaga, Spain. [Claros,MG] Instituto de Hortofruticultura Subtropical Y Mediterránea (IHSM-UMA-CSIC), Málaga, Spain. [Gagnaire,PA] ISEM, Univ Montpellier, CNRS, EPHE, IRD, Montpellier, France. [Manchado,M] Crecimiento Azul, Centro IFAPA El Toruño, Unidad Asociada al CSIC, El Puerto de Santa María, Spain., and This study was funded by project RTA2017-00054-C03-01 and RTA2017-00054-C03-funded from MCIU/AEI/FEDER, UE and cofunded 80% by Programa Operativo FEDER de Andalucía 2014-2020, project PP.AVA.AVA201601.9 SOLEALGAE. Moreover, the study has received funding from EU H2020 research and innovation program under grant agreement 817992 ERANET-BLUEBIO COFUND project PCI2020-111994 BestBrood/AEI/10.13039/501100011033. IGC is funded by a predoctoral fellowship from INIA. This work would not have been possible without the computer resources and the technical support provided by the Plataforma Andaluza de Bioinformática of the University of Málaga and CNAG. We acknowledge the support of the Spanish Ministry of Science, Innovation and Universities to the EMBL partnership, the Centro de Excelencia Severo Ochoa and the CERCA Programme/Generalitat de Catalunya, the Spanish Ministry of Science and Innovation through the Instituto de Salud Carlos III, Generalitat de Catalunya through Departament de Salut and Departament d’Empresa i Coneixement and co-fnancing with funds from the European Regional Development Fund by the Spanish Ministry of Science and Innovation corresponding to the Programa Operativo FEDER Plurirregional de España (POPE) 2014-2020 and by the Secretaria d’Universitats i Recerca, Departament d’Empresa i Coneixe ment of the Generalitat de Catalunya corresponding to the Programa Operatiu FEDER de Catalunya 2014-20.

Subjects
Male, Genetic Linkage, Genome-wide association study, Genome, 0302 clinical medicine, Organisms::Eukaryota::Animals [Medical Subject Headings], Diferenciación sexual, Ligamiento genético, Genetics, Gene Rearrangement, 0303 health sciences, Multidisciplinary, Contig, Organisms::Eukaryota::Animals::Chordata::Vertebrates::Fishes::Flatfishes [Medical Subject Headings], [SDV.BID.EVO]Life Sciences [q-bio]/Biodiversity/Populations and Evolution [q-bio.PE], Genomics, Penetrance, Repeticiones de microsatélite, Fatfish, Phenomena and Processes::Genetic Phenomena::Genetic Structures::Base Sequence::Repetitive Sequences, Nucleic Acid::Tandem Repeat Sequences::Microsatellite Repeats [Medical Subject Headings], Flatfishes, Medicine, Sex, Female, Heterogametic sex, Phenomena and Processes::Genetic Phenomena::Genetic Processes::Sex Determination Processes [Medical Subject Headings], Science, Check Tags::Male [Medical Subject Headings], Locus (genetics), Biology, Chromosome, Article, Phenomena and Processes::Genetic Phenomena::Genetic Structures::Genome [Medical Subject Headings], 03 medical and health sciences, Genetic linkage, Phenomena and Processes::Genetic Phenomena::Genetic Linkage [Medical Subject Headings], Phenomena and Processes::Genetic Phenomena::Genetic Processes::Gene Rearrangement [Medical Subject Headings], Animals, Genoma, 030304 developmental biology, Animal breeding, Cromosomas, Sex Determination Processes, [SDV.GEN.GA]Life Sciences [q-bio]/Genetics/Animal genetics, Genes, Check Tags::Female [Medical Subject Headings], 030217 neurology & neurosurgery, Genome-Wide Association Study, Microsatellite Repeats
Abstract

The integration of physical and high-density genetic maps is a very useful approach to achieve chromosome-level genome assemblies. Here, the genome of a male Senegalese sole (Solea senegalensis) was de novo assembled and the contigs were anchored to a high-quality genetic map for chromosome-level scaffolding. Hybrid assembled genome was 609.3 Mb long and contained 3403 contigs with a N50 of 513 kb. The linkage map was constructed using 16,287 informative SNPs derived from ddRAD sequencing in 327 sole individuals from five families. Markers were assigned to 21 linkage groups with an average number of 21.9 markers per megabase. The anchoring of the physical to the genetic map positioned 1563 contigs into 21 pseudo-chromosomes covering 548.6 Mb. Comparison of genetic and physical distances indicated that the average genome-wide recombination rate was 0.23 cM/Mb and the female-to-male ratio 1.49 (female map length: 2,698.4 cM, male: 2,036.6 cM). Genomic recombination landscapes were different between sexes with crossovers mainly concentrated toward the telomeres in males while they were more uniformly distributed in females. A GWAS analysis using seven families identified 30 significant sex-associated SNP markers located in linkage group 18. The follicle-stimulating hormone receptor appeared as the most promising locus associated with sex within a region with very low recombination rates. An incomplete penetrance of sex markers with males as the heterogametic sex was determined. An interspecific comparison with other Pleuronectiformes genomes identified a high sequence similarity between homologous chromosomes, and several chromosomal rearrangements including a lineage-specific Robertsonian fusion in S. senegalensis. This study was funded by project RTA2017-00054-C03-01 and RTA2017-00054-C03-funded from MCIU/AEI/FEDER, UE and cofunded 80% by Programa Operativo FEDER de Andalucía 2014-2020, project PP.AVA.AVA201601.9 SOLEALGAE. Moreover, the study has received funding from EU H2020 research and innovation program under grant agreement 817992 ERANET-BLUEBIO COFUND project PCI2020-111994 BestBrood/AEI/10.13039/501100011033. IGC is funded by a predoctoral fellowship from INIA. This work would not have been possible without the computer resources and the technical support provided by the Plataforma Andaluza de Bioinformática of the University of Málaga and CNAG.

Published
2021
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3. Genetic Estimates for Growth and Shape-Related Traits in the Flatfish Senegalese Sole

Author

Israel Guerrero-Cózar, Concha Berbel, Elena Espinosa, Manuel Gonzalo Claros, Ricardo Zerolo, Eduardo Jiménez-Fernández, Manuel Manchado, [Guerrero-Cozar,I, Berbel,C, Manchado,M] IFAPA Centro El Toruño, Junta de Andalucía, El Puerto de Santa María Cádiz, Spain. [Jimenez-Fernandez,E] Free Radical Research Group, Centre for Health Sciences, University of the Highlands and Islands, UK. [Espinosa,E, Claros,MG] Molecular Biology and Biochemistry Department, University of Málaga, Málaga, Spain. [Claros,MG] CIBER de Enfermedades Raras (CIBERER), Málaga, Spain. [Claros,MG] Institute of Biomedical Research in Málaga (IBIMA), IBIMA-RARE, Málaga, Spain. [Claros,MG] Instituto de Hortofruticultura Subtropical y Mediterránea (IHSM-UMA-CSIC), Málaga, Spain. [Zerolo,R] CUPIMAR, San Fernando, Cádiz, Spain. [Manchado,M] 'Crecimiento Azul', Centro IFAPA El Toruño, Unidad Asociada al CSIC, El Puerto de Sta. María, Spain., and This research was funded by MCIU/AEI/FEDER, UE, grant numbers RTA2017-00054-C03- 01 and RTA2017-00054-C03-03. Moreover, this research was funded by the EU H2020 research and innovation program under grant agreement 817992 ERANET-BLUEBIO COFUND project PCI2020-111994 'BestBrood'. IGC is funded by a predoctoral fellowship from INIA.

Subjects
Phenomena and Processes::Genetic Phenomena::Phenotype [Medical Subject Headings], Acuicultura, Anatomy::Animal Structures::Animal Fins [Medical Subject Headings], Body height, Peduncle (anatomy), Veterinary medicine, growth, Productos pesqueros, Production cycle, Growth, Biology, Breeding, Variación genética, Cruzamiento, shape, Crecimiento, Article, Forma de la célula, genetic estimates, 03 medical and health sciences, Animal science, Flatfish, Technology and Food and Beverages::Technology, Industry, and Agriculture::Agriculture::Aquaculture [Medical Subject Headings], SF600-1100, Organisms::Eukaryota::Animals [Medical Subject Headings], Phenomena and Processes::Physiological Phenomena::Physiological Processes::Growth and Development [Medical Subject Headings], Solea senegalensis, 030304 developmental biology, Phenomena and Processes::Biological Phenomena [Medical Subject Headings], 0303 health sciences, General Veterinary, Organisms::Eukaryota::Animals::Chordata::Vertebrates::Fishes::Flatfishes [Medical Subject Headings], Fish fin, Shape, 04 agricultural and veterinary sciences, Genetic estimates, biology.organism_classification, QL1-991, breeding, Animales, 040102 fisheries, 0401 agriculture, forestry, and fisheries, %22">Fish , Animal Science and Zoology, Phenomena and Processes::Genetic Phenomena::Genetic Variation [Medical Subject Headings], Zoology, Senegalese sole, Peces planos
Abstract

Shape quality is very important in flatfish aquaculture due to the impact on commercialization. The Senegalese sole (Solea senegalensis) is a valuable flatfish with a highly elliptic body that slightly changes with age and size, and it is prone to accumulating malformations during the production cycle. The present study aims to investigate the genetic parameters of two growth traits (weight and standard length) and six shape quality predictors (ellipticity, three body heights (body height at the pectoral fin base [BHP], body maximum height [BMH] and caudal peduncle height [CPH]) and two ratios (BMH/BHP and BMH/CPH)). These traits were measured before the on-growing stage (age ~400 days (d)) and at harvest (~800 d). Phenotypic data, heritabilities and genetic and phenotypic correlations between the traits are presented and discussed. High or very high heritabilities (0.433–0.774) were found for growth traits, body heights and ellipticity and they were higher at 400 than 800 d. In contrast, the ratios of BMH/BHP and BMH/CPH were less heritable (0.144–0.306). Positive and very high (&gt, 0.95) correlations between growth traits and the three heights were found and decreased with age. In contrast, ellipticity had negative and medium-high genetic correlations with growth traits and heights, indicating fish selected for bigger size would also become rounder. The ratio of BMH/CPH showed low genetic correlations with all traits and provided complementary information to ellipticity for a better fitting to the expected lanceolate body morphology of sole. The genetic correlations for all traits at both ages were very high, indicating that selection before entering the growth-out stage in recirculation aquaculture systems is recommended to accelerate genetic gains.

Published
2021
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