Scion genetics and improvement: desirable consumer traits
Abstract
The use of genetic markers for important consumer-preferred sensory traits will streamline the breeding process making it more rapid and less expensive. Initial studies have identified quantitative trait loci that explain differences in the volatile composition of wines from grapevine mapping populations that are related to ‘fruity’, ‘green’ and ‘floral’ sensory characters. Genome-wide association studies using a variety collection have identified quantitative trait loci and markers that predict the abundance of wine volatiles. Other markers have been designed to genes already shown to be important for wine phenotypes (methoxypyrazines, rotundone, anthocyanin stability, red flesh) to aid the selection of new scions for specific wine styles.
Summary
The winegrape varieties most extensively used throughout Australia are hundreds of years old and lack resistance to the common fungal pathogens powdery and downy mildew. Breeding programs around the world are using resistance genes from wild Vitis species introgressed into vinifera to produce new fungal disease resistant varieties, and this is true for Australia in the form of Wine Australia Project CSA 1701-1.1. The uptake of these new varieties will be improved if the advances in agronomic performance can be matched by these cultivars producing wines with high consumer acceptance. This project was designed to streamline the selection of such varieties by developing genetic markers for consumer-preferred sensory traits so that these plants can be identified without the need for small scale winemaking and sensory analysis. Added benefits from the development of such markers include a greater understanding of how grape composition influences wine style and the development of tools to predict how environment and management will affect grape and wine composition.
The project methodology consisted of four main parts. 1) Use of existing biparental crosses between the microvine and either Riesling or Cabernet Sauvignon to generate micro-scale wines and genetic linkage maps from the progeny. Volatile compound profiling from the wines would then allow the mapping of quantitative trait loci (QTLs) associated with the concentration of the wine compounds. Candidate genes in the genome regions underlying the QTLs would be identified, characterised and markers designed if found to be causing the phenotypic changes. 2) Sample leaves and berries from the varietal collection held by SARDI at the Nuriootpa Research Centre and use these to generate genome single nucleotide polymorphism (SNP) data using DArTseq and volatile compound profiles from micro-scale wines. Genome-wide association studies (GWAS) would then be used to identify SNPs associated with the concentration of wine volatile compounds and candidate genes characterised and/or markers then developed if appropriate. 3) Generate new germplasm or identify existing populations that are useful for mapping consumer-preferred traits. Sample leaves and berries and produce matching SNP and wine volatile compound datasets for mapping using linkage maps or GWAS. 4) Develop easy-to-use markers for laboratory detection of SNPs found to be responsible for consumer-preferred traits according to research published in peer-reviewed journals by other research teams.
The biparental crosses were successfully used to map genome regions associated with the following wine compounds: (Z)-3-hexenol, associated with ‘leafy green’ sensory characters; acetate esters, which cause ‘fruity’ and ‘floral’ type aromas; linalool, which contributes to ‘citrus’ and ‘floral’ sensory attributes and; β-damascenone, an enhancer of ‘fruity’ flavours and aromas. In the case of (Z)-3-hexenol, causal genes were identified in the genome region that catalyse the isomerisation of (Z)-3-hexenol to (E)-3-hexenal and markers were generated to predict this phenotype in breeding programs. For the acetate esters, SNPs in several candidate genes in the single QTL region identified, were tested against the data from the SARDI variety collection, but none were significantly associated with the acetate ester concentrations.
Nevertheless, the mapping of this phenotype to the grape genome continues to provide evidence that grape composition can change acetate ester production, by yeast, during fermentation. Moderate linalool production, which imparts ‘citrus’ and ‘floral’ characters in varieties like Riesling and Viognier, appeared to be inhibited by a gene linked to the dwarf locus in the microvine. Potential causal genes were characterised, and these NUDX hydrolases were shown to be able to dephosphorylate the precursors of monoterpenes and sesquiterpenes. However, genetic differences in these genes between the parents of the progeny in the biparental cross were not found and so the genotype:phenotype link was not proven. Finally, β-damascenone concentration in wines made from the microvine × Cabernet Sauvignon progeny mapped to a region on chromosome 2 that co-localised with the flower sex locus. It was found that the plants with female flowers had higher concentrations of β-damascenone in the wines, probably driven by a higher skin:flesh ratio in the very small berries produced from such flowers.
Valuable datasets were developed during the preparation for the GWAS which can be used in future projects. Small-scale wines were produced from 177 grape varieties and these were analysed using gas chromatography-mass spectrometry to obtain volatile compound profiles. To compliment the phenotypic analysis, DNA was prepared from each variety and thousands of markers generated using the DArTseq platform of an external provider. The GWAS using these datasets identified QTLs for 17 wine volatile compounds and, in many cases, candidate genes were identified in the vicinity of the significant markers. Importantly, the GWAS confirmed the location of the QTL for (Z)-3-hexenol concentration in wine previously identified in the biparental crosses. The results of this work will inform future studies into the genetics of consumer-preferred traits that could feed into new germplasm improvement initiatives.
One new population was generated for genetic studies comprising F2 progeny derived from a self- pollinated F1 plant originally generated from the microvine × Riesling cross which produced Riesling- like concentrations of linalool. A dense genetic map was generated from this population and the fruit from the progeny was analysed for linalool concentrations using two extraction techniques. Subsequent QTL mapping found that the production of linalool in this population was multi-genic with seven QTLs identified. However, the future utility of this cross was demonstrated through the identification of a single major QTL responsible for the production of the monoterpene α- phellandrene with credible candidate genes located in the associated genome region. An existing population of 1st generation mildew resistant vines, located at the SARDI Nuriootpa Research Centre, was also used for GWAS to identify genome regions associated with wine volatile compounds. While some significant relationships were observed, a deeper examination of the data is required to determine if another mapping strategy would be more successful.
Markers for five traits were developed from previously published research and the work conducted in this project. The markers were designed as cleaved amplified polymorphic sequences (CAPS) markers for ease-of-use in molecular biology laboratories. The traits targeted were: acylated (stable) anthocyanins; methoxypyrazine production in berries; α-guaiene production (rotundone precursor); red-fleshed berries; (Z)-3-hexenol concentration in wine.
In addition to the identification of QTLs for many wine volatile compounds and the development of markers for mapping consumer-preferred traits, this project has generated materials, data and analytical pipelines that can be used to deliver superior germplasm to the Australian wine sector by identifying targets for DNA-free gene editing technology or conventional breeding.