Abstract
High-throughput DNA and RNA sequencing technologies have resulted in the successful identification of Single nucleotide polymorphisms (SNPs). In order to develop a large SNP set for wide application in apricot (Prunus armeniaca L.), we carried out RNA high-throughput sequencing (RNA-Seq) in two apricot genotypes, “Rojo Pasión” and “Z506-7.” After trimming and cleaning, 70 % of RNA-Seq reads were aligned to the reference peach genome. Sequences uniquely mapped on the peach genome allowed for the discovery of 300 k SNP/INDEL variations, with a density of one SNP per 850 bp. Some 95 SNPs of the 99 tested were analyzed in a set of 37 apricot accessions using SNPlex™ genoty** technology. The results provide accurate values for nucleotide diversity in coding sequences in apricot. The combination of a highly efficient RNA-Seq approach and SNPlex™ high-throughput genoty** technology thus provide a powerful tool for apricot genetic analysis. SNP markers produced a total of 267 allelic combinations in the 37 apricot accessions assayed with a mean of 2.8 combinations per locus, an observed heterozygosity per marker ranging from 0.06 to 0.65, and a power of discrimination ranging from 0.12 to 0.66. In addition, SNP markers confirmed parentage and also determined relationships between the accessions in a manner consistent with their pedigree relationships.
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Acknowledgments
This study was supported by the projects “Gene expression analysis of the resistance to Plum pox virus, PPV (sharka) in apricot by transcriptome deep-sequencing (RNA-Seq)” (AGL2010-16335) and “Apricot breeding” (AGL2010-21903) of the Spanish Ministry of Science. The authors would also like to thank the Fondazione Cassa di Risparmio in Bologna (Italy) for supporting the Sequenom analysis in the Centre for Applied Biomedical Research (CRBA) of Bologna as well as Manuel Castro de Moura of aScidea Computational Biology Solutions, S.L. in Barcelona (Spain) for his help in the bioinformatic analysis of the RNA-Seq data.
Data archiving statement
The plant material studied is registered in the Plant Variety Database (PLUTO; http://www.upov.int/pluto/en) belonging to the International Union for the Protection of New Varieties of Plants (UPOV) (http://www.upov.int). Apricot cultivars and new selections not included in PLUTO belong to the germplasm collection and breeding programs of CEBAS-CSIC and the Università degli Studi di Bologna, which includes some breeding research material in the Genome Database for Rosaceae (GDR, http://www.rosaceae.org). Part of the RNA-Seq reads from the sequencing of the Rojo Pasión transcriptome were included in the DNA Data Bank of Japan (DDBJ) (http://trace.ddbj.nig.ac.jp) and the European Molecular Biology Laboratory-European Bioinformatics Institute (EMBL-EBI) (http://www.ebi.ac.uk/) Sequence Read Archive (SRA). Submission SRA = 48787; DDBJ Reference SRP009939; EMBL-EBI Reference SRS283323. The rest of the reads not included in this file are available at the Department of Plant Breeding of CEBAS-CSIC.
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Communicated by A. G. Abbott
Juan Alfonso Salazar and Manuel Rubio contributed equally to this work.
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Table S1
Summary of the quality control and the Paired-end RNA-Seq reads conserved after preprocesing in the apricot transcriptomes of ‘Rojo Pasión’ (3 libraries: ceb1, ceb3, ceb4) and ‘Z506-7’ (2 libraries: ceb5, ceb6) sequenced. (DOCX 776 kb)
Table S2
Total variants (including SNPs and INDELs) and INDELs identified in the different scaffolds by RNA-Seq in the two apricot transcriptomes assayed of ‘Rojo Pasión’ and ‘Z506-7’. (XLSX 43518 kb)
Table S3
SNP markers identified in the apricot transcriptomes of ‘Rojo Pasión’ (3 libraries: ceb1, ceb3, ceb4) and ‘Z506-7’ (2 libraries: ceb5, ceb6) and related sequences. (XLSX 37 kb)
Table S4
Polymorphisms obtained in the application of 96 polymorphic SNP markers by Sequenom technology in the 37 apricot genotypes. (XLSX 29 kb)
Table S5
Number of genotypic classes, heterozygosity and power of discrimination of the SNP markers detected after Sequenom assay in the 37 apricot genotypes. (XLSX 25 kb)
Table S6
Pairwise genetic distances of the 37 apricot genotypes studied obtained via the application of the 96 validated SNP markers assayed by Sequenom technology and the MCL method. (XLSX 21 kb)
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Salazar, J.A., Rubio, M., Ruiz, D. et al. SNP development for genetic diversity analysis in apricot. Tree Genetics & Genomes 11, 15 (2015). https://doi.org/10.1007/s11295-015-0845-2
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DOI: https://doi.org/10.1007/s11295-015-0845-2