Sandra Gordon

L-Ascorbic acid accumulation in fruit of Ribes nigrum occurs by in situ biosynthesis via the L-galactose pathway

Blackcurrant (Ribes nigrum L.) is a widely grown commercial crop valued for its high vitamin C (l-ascorbic acid, AsA) content. In the present study, a systematic analysis of the mechanism of fruit AsA accumulation was undertaken. AsA accumulation occurred during fruit expansion and was associated with high in situ biosynthetic capacity via the l-galactose pathway and low rates of turnover. Cessation of AsA accumulation was associated with reduced biosynthesis and increased turnover. Translocation of AsA from photosynthetic or vegetative tissues contributed little to fruit AsA accumulation. Manipulation of substrate availability by defoliation had no effect on fruit AsA concentration but significantly reduced fruit yields. Supply of the AsA precursor l-galactono-1,4-lactone to intact, attached fruit transiently increased fruit AsA concentration which rapidly returned to control levels after removal of the compound. These data suggest strong developmental, metabolic and genetic control of AsA accumulation in blackcurrant fruit and indicate the potential for breeding high AsA cultivars.

Candidate genes associated with bud dormancy release in blackcurrant (Ribes nigrum L.)

BackgroundThe 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.ResultsUsing 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.ConclusionsThis 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.

The use of genotyping by sequencing in blackcurrant ( Ribes nigrum ): developing high-resolution linkage maps in species without reference genome sequences

A genotyping by sequencing (GbS) approach is reported in blackcurrant (Ribes nigrum L.) using a de novo read assembly method developed because of the current absence of a reference genome sequence for this species. A new approach to single nucleotide polymorphism (SNP) genotype calling is described, where individual genotypes for a large number of SNPs were characterised from the GbS counts using a novel method based on a functional regression of major and minor allele read counts. The high-quality GbS SNPs were combined with SNPs and simple sequence repeats generated from other technologies to develop a linkage map with increased marker density and improved genome coverage, containing up to 204 SNPs on each linkage group. SNPs of lower quality were then located on the map using quantitative trait locus (QTL) interval mapping of the proportion of the major allele. Two QTL each for 100-berry weight and Brix scores, measured over three years, were identified using the map. The use of this approach to identify and map a significant number of novel SNPs in a woody species with hitherto limited genomic resources may have generic application to other under-resourced and minor species in the development of cost-effective and efficient high-density genetic maps.

The development of a genetic linkage map of blackcurrant (Ribes nigrum L.) and the identification of regions associated with key fruit quality and agronomic traits

The first genetic linkage map of blackcurrant (Ribes nigrum L.) was constructed using AFLP, SSR (genomic and EST-derived) and SNP markers, in a mapping population derived from two diverse breeding clones of blackcurrant from the SCRI breeding programme. Cluster analysis of the population revealed that the individuals within the population formed two distinct sub-populations, with segregation ratios consistent with one sub-population having the two intended parents, and the other being selfed segregants. The latter sub-population improves the map by providing a more informative estimate of recombination frequency than the crossed sub-population for some marker configurations, and also revealed the presence of two unlinked loci affecting viability. Several important phenological, agronomic and fruit quality traits were evaluated in the mapping population, and QTLs affecting these are located on the linkage map. This provides a framework for the development of marker-assisted breeding strategies for blackcurrant, to improve breeding efficiency and time to cultivar.

The development of a PCR-based marker linked to resistance to the blackcurrant gall mite (Cecidophyopsis ribis Acari: Eriophyidae)

Gall mite (Cecidophyopsis ribis) is the most serious pest of blackcurrant (Ribes nigrum L.), causing the damaging condition known as ‘big bud’ and also transmitting blackcurrant reversion virus (BRV) within and between plantations. The identification of resistant germplasm is at present a time-consuming and expensive process, dependent on field infestation plots. Resistance based on gene Ce introgressed from gooseberry has been used in UK breeding programmes for blackcurrant. Using a bulked segregant analysis, 90 AFLP primer combinations were screened and a linkage map constructed around the resistance locus controlled by Ce. Sixteen of the primer combinations produced a fragment in the resistant bulked progeny and the gall mite-resistant parent, but not in the susceptible bulked progeny and parent; subsequent testing on individual progeny identified an AFLP fragment closely linked to gall mite resistance. This fragment, designated E41M88-280, was converted to a PCR-based marker based on sequence-specific primers, amplifying only in resistant individuals. Validation of this marker across a range of susceptible and resistant blackcurrant germplasm with different genetic backgrounds confirmed its reliability in the identification of mite-resistant germplasm containing gene Ce. The conversion of an AFLP fragment to a sequence-based PCR marker simplifies its application and therefore increases its utility for selection of mite-resistant germplasm in high-throughput breeding programmes for blackcurrant.

Chilling requirement of ribes cultivars

It is usually thought that adequate winter chill is required for the full flowering of many temperate woody species. This paper investigates the sensitivity of blackcurrant bud burst and flowering to natural weather fluctuations in a temperate maritime climate, and compares a range of chill models that have been proposed for assessing the accumulation of winter chill. Bud break for four contrasting cultivars are compared in an exceptionally cold and in a mild winter in Eastern Scotland. The results confirm the importance of chilling at temperatures lower than 0°C and demonstrate that no single chilling function applies equally to all blackcurrant cultivars. There is a pressing need for further model development to take into account the relationship between chilling temperatures and warming temperatures occurring both during and after the chill accumulation period.

A Transcript and Metabolite Atlas of Blackcurrant Fruit Development Highlights Hormonal Regulation and Reveals the Role of Key Transcription Factors

Blackcurrant fruit collected at six stages of development were assessed for changes in gene expression using custom whole transcriptome microarrays and for variation in metabolite content using a combination of liquid chromatography-mass spectrometry and gas chromatography-mass spectrometry. Principal components analysis demonstrated that fruit development could be clearly defined according to their transcript or metabolite profiles. During early developmental stages, metabolite profiles were dominated by amino acids and tannins, whilst transcript profiles were enriched in functions associated with cell division, anatomical structure morphogenesis and cell wall metabolism. During mid fruit development, fatty acids accumulated and transcript profiles were consistent with seed and embryo development. At the later stages, sugars and anthocyanins accumulated consistent with transcript profiles that were associated with secondary metabolism. Transcript data also indicated active signaling during later stages of fruit development. A targeted analysis of signaling networks revealed a dynamic activation and repression of almost 60 different transcripts encoding transcription factors across the course of fruit development, many of which have been demonstrated as pivotal to controlling such processes in other species. Transcripts associated with cytokinin and gibberellin were highly abundant at early fruit development, whilst those associated with ABA and ethylene tended to be more abundant at later stages. The data presented here provides an insight into fruit development in blackcurrant and provides a foundation for further work in the elucidation of the genetic basis of fruit quality.

Developing Tools for Long-Term Breeding of Blueberry Germplasm for UK Production

Fruit consumption in the UK, particularly of berry fruits, is expanding rapidly and consumer demand for blueberries (Vaccinium spp.) is at record levels with UK growers unable to meet current demand. Consumers are aware that eating fruit can be pleasurable and with appropriate packaging, convenient, but blueberry remains unfamiliar to a wide cross section of UK consumers. There is great potential for UK growers to supply high quality fresh blueberries across a wide season so that consumption of this healthy fruit can be integrated into the British diet. A genetic framework for future crop improvement is required to develop a thriving and sustainable industry. The genetic component of this project builds on the statistical developments derived from the software program, TetraploidMap, to identify fruit quality, health, and agronomic-related quantitative trait loci in tetraploid blueberry for marker assisted breeding.