Identification, utilisation and mapping of novel transcriptome-based markers from blackcurrant (Ribes nigrum)

<p>Abstract</p> <p>Background</p> <p>Deep-level second generation sequencing (2GS) technologies are now being applied to non-model species as a viable and favourable alternative to Sanger sequencing. Large-scale SNP discovery was undertaken in blackcurrant (<it>Ribes nigrum </it>L.) using transcriptome-based 2GS 454 sequencing on the parental genotypes of a reference mapping population, to generate large numbers of novel markers for the construction of a high-density linkage map.</p> <p>Results</p> <p>Over 700,000 reads were produced, from which a total of 7,000 SNPs were found. A subset of polymorphic SNPs was selected to develop a 384-SNP OPA assay using the Illumina BeadXpress platform. Additionally, the data enabled identification of 3,000 novel EST-SSRs. The selected SNPs and SSRs were validated across diverse <it>Ribes </it>germplasm, including mapping populations and other selected <it>Ribes </it>species.</p> <p>SNP-based maps were developed from two blackcurrant mapping populations, incorporating 48% and 27% of assayed SNPs respectively. A relatively high proportion of visually monomorphic SNPs were investigated further by quantitative trait mapping of theta score outputs from BeadStudio analysis, and this enabled additional SNPs to be placed on the two maps.</p> <p>Conclusions</p> <p>The use of 2GS technology for the development of markers is superior to previously described methods, in both numbers of markers and biological informativeness of those markers. Whilst the numbers of reads and assembled contigs were comparable to similar sized studies of other non-model species, here a high proportion of novel genes were discovered across a wide range of putative function and localisation. The potential utility of markers developed using the 2GS approach in downstream breeding applications is discussed.</p

Construction of a SNP and SSR linkage map in autotetraploid blueberry using genotyping by sequencing

The construction of the first genetic map in autotetraploid blueberry has been made possible by the development of new SNP markers developed using genotyping by sequencing in a mapping population created from a cross between two key highbush blueberry cultivars, Draper × Jewel (Vaccinium corymbosum). The novel SNP markers were supplemented with existing SSR markers to enable the alignment of parental maps. In total, 1794 single nucleotide polymorphic (SNP) markers and 233 simple sequence repeat (SSR) markers exhibited segregation patterns consistent with a random chromosomal segregation model for meiosis in an autotetraploid. Of these, 700 SNPs and 85 SSRs were utilized for construction of the ‘Draper’ genetic map, and 450 SNPs and 86 SSRs for the ‘Jewel’ map. The ‘Draper’ map comprises 12 linkage groups (LG), associated with the haploid chromosome number for blueberry, and totals 1621 cM while the ‘Jewel’ map comprises 20 linkage groups totalling 1610 cM. Tentative alignments of the two parental maps have been made on the basis of shared SSR alleles and linkages to double-simplex markers segregating in both parents. Tentative alignments of the two parental maps have been made on the basis of shared SSR alleles and linkages to double-simplex markers segregating in both parents.This is the publisher’s final pdf. The published article is copyrighted by Springer and can be found at: http://link.springer.com/journal/11032Keywords: Vaccinium corymbosum, Tetraploid Map, Genotyping by sequencing, Autotetraploi

Candidate genes associated with bud dormancy release in blackcurrant (<it>Ribes nigrum </it>L.)

Abstract Background The detrimental effects of mild winter temperatures on the consistency of cropping of blackcurrant (Ribes nigrum L.) in parts of Europe have led to increasing interest in the genetic control of dormancy release in this species. This study examined patterns of gene expression in leaf buds of blackcurrant to identify key differential changes in these profiles around the time of budbreak. Results Using leaf bud tissue of blackcurrant, a cDNA library was generated as a source of blackcurrant ESTs for construction of a custom microarray, which was used to identify differential gene expression during dormancy release. Gene activity was lowest in early stages of dormancy, increasing to reach a maximum around the time of budbreak. Genes with significantly changing expression profiles were clustered and evidence is provided for the transient activity of genes previously associated with dormancy processes in other species. Expression profiling identified candidate genes which were mapped onto a blackcurrant genetic linkage map containing budbreak-related QTL. Three genes, which putatively encode calmodulin-binding protein, beta tubulin and acetyl CoA carboxylase respectively, were found to co-localise with budbreak QTL. Conclusions This study provides insight into the genetic control of dormancy transition in blackcurrant, identifying key changes in gene expression around budbreak. Genetic mapping of ESTs enabled the identification of genes which co-localise with previously-characterised blackcurrant QTL, and it is concluded that these genes have probable roles in release of dormancy and can therefore provide a basis for the development of genetic markers for future breeding deployment.</p

McCallumConstructionSNPandSSRSupplementaryMaterial3.pdf

The construction of the first genetic map in autotetraploid blueberry has been made possible by the development of new SNP markers developed using genotyping by sequencing in a mapping population created from a cross between two key highbush blueberry cultivars, Draper × Jewel (Vaccinium corymbosum). The novel SNP markers were supplemented with existing SSR markers to enable the alignment of parental maps. In total, 1794 single nucleotide polymorphic (SNP) markers and 233 simple sequence repeat (SSR) markers exhibited segregation patterns consistent with a random chromosomal segregation model for meiosis in an autotetraploid. Of these, 700 SNPs and 85 SSRs were utilized for construction of the ‘Draper’ genetic map, and 450 SNPs and 86 SSRs for the ‘Jewel’ map. The ‘Draper’ map comprises 12 linkage groups (LG), associated with the haploid chromosome number for blueberry, and totals 1621 cM while the ‘Jewel’ map comprises 20 linkage groups totalling 1610 cM. Tentative alignments of the two parental maps have been made on the basis of shared SSR alleles and linkages to double-simplex markers segregating in both parents. Tentative alignments of the two parental maps have been made on the basis of shared SSR alleles and linkages to double-simplex markers segregating in both parents.Keywords: Vaccinium corymbosum, Tetraploid Map, Autotetraploid, Genotyping by sequencin

McCallumConstructionSNPandSSRSupplementaryMaterial2.xlsx

The construction of the first genetic map in autotetraploid blueberry has been made possible by the development of new SNP markers developed using genotyping by sequencing in a mapping population created from a cross between two key highbush blueberry cultivars, Draper × Jewel (Vaccinium corymbosum). The novel SNP markers were supplemented with existing SSR markers to enable the alignment of parental maps. In total, 1794 single nucleotide polymorphic (SNP) markers and 233 simple sequence repeat (SSR) markers exhibited segregation patterns consistent with a random chromosomal segregation model for meiosis in an autotetraploid. Of these, 700 SNPs and 85 SSRs were utilized for construction of the ‘Draper’ genetic map, and 450 SNPs and 86 SSRs for the ‘Jewel’ map. The ‘Draper’ map comprises 12 linkage groups (LG), associated with the haploid chromosome number for blueberry, and totals 1621 cM while the ‘Jewel’ map comprises 20 linkage groups totalling 1610 cM. Tentative alignments of the two parental maps have been made on the basis of shared SSR alleles and linkages to double-simplex markers segregating in both parents. Tentative alignments of the two parental maps have been made on the basis of shared SSR alleles and linkages to double-simplex markers segregating in both parents.Keywords: Genotyping by sequencing, Autotetraploid, Vaccinium corymbosum, Tetraploid Ma

McCallumConstructionSNPandSSR.pdf

The construction of the first genetic map in autotetraploid blueberry has been made possible by the development of new SNP markers developed using genotyping by sequencing in a mapping population created from a cross between two key highbush blueberry cultivars, Draper × Jewel (Vaccinium corymbosum). The novel SNP markers were supplemented with existing SSR markers to enable the alignment of parental maps. In total, 1794 single nucleotide polymorphic (SNP) markers and 233 simple sequence repeat (SSR) markers exhibited segregation patterns consistent with a random chromosomal segregation model for meiosis in an autotetraploid. Of these, 700 SNPs and 85 SSRs were utilized for construction of the ‘Draper’ genetic map, and 450 SNPs and 86 SSRs for the ‘Jewel’ map. The ‘Draper’ map comprises 12 linkage groups (LG), associated with the haploid chromosome number for blueberry, and totals 1621 cM while the ‘Jewel’ map comprises 20 linkage groups totalling 1610 cM. Tentative alignments of the two parental maps have been made on the basis of shared SSR alleles and linkages to double-simplex markers segregating in both parents. Tentative alignments of the two parental maps have been made on the basis of shared SSR alleles and linkages to double-simplex markers segregating in both parents.Keywords: Autotetraploid, Genotyping by sequencing, Vaccinium corymbosum, Tetraploid Ma