Genetics and plant breeding in Australia

1. Genetics and plant breeding in Australia CSIRO Vitis breeding: Dr. Rob Walker Power of genomic approaches: Dr. Chris Ford

2. A u s t r a l i a ’ s G r o w i n g F u t u r e Grapevine Breeding Viticultural Seminar Argentina May 2007 Dr. Rob Walker, CSIRO Plant Industry

3. Table grapes Dried grapes Wine grapes Rootstocks Grapevine breeding in Australia CSIRO is working with Australia’s grape industries to develop new and improved grapevine varieties

4. Tablegrape breeding and evaluation Table grapes • National program • Breeding by CSIRO • Evaluation in WA, Qld and NT by state agencies • Aiming for seedlessness, excellent taste, ripe and uniform maturity, large and uniform berry size and good production and postharvest characteristics. M1 vines from irradiated buds Top-working new selections Range of colours, sizes, textures and flavours Red seedless Bright red early seedless White muscat seedless Crisp white seedless Late white seedless

5. Millennium Muscat™ Seeded Very early Muscat flavour Domestic market

6. Magic Seedless™ Black Early Seedless

7. Dried grapes • Emphasis is on rain tolerant, low browning sultana types which are disease resistant and show consistent fruitfulness • Involves: • In-ovulo embryo rescue • Top-working for accelerated evaluation • Evaluation under mechanised systems (world’s best practice) • Comparison with comparator varieties • Range of selections under evaluation

8. Heritability and expected genetic gain

9. Rootstocks Screening techniques, selection & inheritance Assessment of selections as grafted vines Regional semi-commercial evaluation Germplasm – Vitis spp. Crosses  Interspecific families Creation and recombination of genetic variability Ease of propagation Graft compatibility Nematode tolerance Mineral discrimination 1000s • Viticultural performance • Grafted vine vigour • Phylloxera tolerance • Yield • WUE & drought tolerance • Carbohydrate partitioning • Fruit quality • Wine quality 75 Commercial performance G x E 4 Industry adoption

10. Rootstocks for quality wine Current Focus Stock-scion compatibility • New rootstocksPBR recently released • to industry for evaluation • Selected for: • low to moderate vigour • ‘reduced’ potassium uptake • enhanced wine quality • tolerance of phylloxera and nematodes Merbein 5489 Merbein 5512 Merbein 6262 Salt tolerance and water use efficiency SALT SUSCEPTIBLE SALT TOLERANT

11. Winegrapes • Emphasis on varieties suited to Australian conditions, particularly warm-climate regions. • Selection for yield performance, improved grape juice composition and wine quality. • January 2000 releases: TyrianPBR, CiennaPBRandRubiennePBR Compared with (parent) Cabernet Sauvignon, all 3 have improved yields, higher juice titratable acidity, higher wine colour density and total anthocyanins and have higher colour hue (brighter wine).

12. Pre-breeding • Genes and traits (CSIRO Plant Industry, Adelaide) • Colour and tannins • Grape berry development and ripening • Fruit flavour and aroma • Fungal pathology • Carotenoids, hormones and flavour • Rapid flowering genotypes and breeding efficiency ... to make this wine... Growing these grapes... ... to respond to this consumer

13. Gene expression and vine performance Microarray mapping of gene expression (Mark Thomas and Chris Davies CSIRO Plant Industry Adelaide) Future – microarrays will enable :- Comparison of gene expression in: - high versus low quality - warm versus cool climate - full versus restricted irrigation International Grapevine Genome Program Multinational research initiative to use a genomics approach to discover and determine the function of all grapevine genes.

14. Key people and inputs • Table grapes (Peter Clingeleffer and Steve Sykes) • Dried grapes (Peter Clingeleffer, Steve Sykes and Steve Swain) • Rootstocks (Peter Clingeleffer, Steve Sykes, Rob Walker and Tim Jones)

15. Genetics and plant breeding in Australia CSIRO Vitis breeding: Dr. Rob Walker Power of genomic approaches: Dr. Chris Ford

16. Genomics and post-genomics approaches to understanding grape berry composition Christopher M Ford School of Agriculture Food and Wine The University of Adelaide

17. Grapevine biotechnology • Classical era – ‘genetic modification’ • Technically difficult with grapevines • Widespread opposition to GM crops • Post-genomics – a ‘systems approach’ • All genes identified ( + physical mapping) • Control of gene expression • All proteins and metabolites identified • varietal; developmental; stress-response • Control of metabolic pathways - composition

18. Grape berry acidity • 2 major acids – tartaric, malic • Malic acid – central metabolite • Tartaric acid – unique, unusual and important • Use of acids in winemaking • $$s costs • Future considerations – global warming… • Context – physiology of TA and acid metabolism

19. Pathways to tartaric acid O COOH 1COOH 1 CO OH 2 OH COOH COOH COOH A HO HO 2 3 HO O O OH OH 4 COOH CHO HO HO HO HO 3 L-tartaric acid + OH C4/C5 cleavage OH OH 4 5CHO HO HO HO 5 CH2OH CH2OH CH2OH CH2OH 6 6CH2OH L-Ascorbic acid 2-keto-L-gulonic acid L-idonic acid 5-keto-D-gluconic acid Glycoaldehyde C2/C3 cleavage B 3COOH OH 1COOH 4 HO + 5 2 COOH CH2OH 6 Oxalic acid L-threonic acid

20. The search for TA synthesis components • Classical approaches – • biochemical • homologous genes • grapevines as experimental systems… • Molecular resources – • analysis of transcripts – identity and abundance • predicted enzymatic activity of encoded sequences

21. Identification of candidate sequences 1. cDNA libraries are clustered based upon similarity of EST expression pattern Leaf Flower Flower Bud, root & shoot Root & leaf veraison, pre- veraison Veraison Ripe berry Petiole, stem Pre-veraison Pre-veraison Post-veraison leaf berry petiole, flower, bud, root 2. Unigenes, grouped on similarity of expression across all 55 cDNA libraries

22. Cross referencing candidate genes for in-depth sequence analysis Identification of differentially expressed cDNAs: 565 candidate genes Limiting EST number to no less than 6 per TC narrowed the number of candidate genes:87 candidate genes Domain and motif screening for oxidoreductase enzymes:8 candidate genes

23. Berries were sampled from 25 species of grapevine. Organic acid levels were compared for each species. Tartaric acid Oxalic acid

24. Correlating biosynthesis with gene expression A. One species identified in this study makesNO tartaric acid B B B. Gene expression studies showed that one candidate gene correlated with tartaric acid biosynthesis; other candidates were expressed in both TA and non TA accumulating grapevines

25. Combined metabolic and transcriptional profiling to identify candidate tartrate biosynthetic genes Prov. Patent 503479 Overexpression in E. coli of the protein encoded by candidate gene assays and tag-facilitated purification allowed determination of its catalytic activity Enzymology revealed substrate specificity and catalytic activity for a key TA synthesis intermediate

26. Outcomes, and future directions • Continuing to investigate the pathway and regulation of TA synthesis • Integrating malate and tartaric acid metabolism • Understanding the metabolism of ascorbic acid during grape berry development

27. The people… • University of Adelaide: • Seth DeBolt (PhD candidate 2003-2006) • Vanessa Melino (PhD candidate 2005- ) • Steve Tyerman, Matt Hayes • Flinders University of South Australia: • Crystal Sweetman (PhD candidate 2006- ) • Crista Burbidge (PhD candidate 2007- ) • Kathy Soole • University of California (Davis) • Doug Cook

28. The funding • This project was supported by the Australian Government’s Cooperative Research Centre Program and conducted by the Cooperative Research Centre for Viticulture. • This work was supported by Australia’s grape growers and winemakers through their investment body, the Grape and Wine Research and Development Corporation, with matching funds from the Australian Government.