Development of unique Australian dried grape varieties Peter Clingeleffer CSIRO Plant Industry Project Number: DG01001

DG01001 This report is published by Horticulture Australia Ltd to pass on information concerning horticultural research and development undertaken for the dried grape industry. The research contained in this report was funded by Horticulture Australia Ltd with the financial support of the dried grape industry. All expressions of opinion are not to be regarded as expressing the opinion of Horticulture Australia Ltd or any authority of the Australian Government. The Company and the Australian Government accept no responsibility for any of the opinions or the accuracy of the information contained in this report and readers should rely upon their own enquiries in making decisions concerning their own interests.

ISBN 0 7341 1076 6 Published and distributed by: Horticultural Australia Ltd Level 1 50 Carrington Street Sydney NSW 2000 Telephone: (02) 8295 2300 Fax: (02) 8295 2399 E-Mail: [email protected] © Copyright 2005

Final report for HA Ltd Project Number DG 01001

Development of unique Australian dried grape varieties

P. R. Clingeleffer and C. R. Tarr CSIRO Plant Industry

HAL Project No. DG01001 Project Leader Peter R. Clingeleffer CSIRO Plant Industry PMB Merbein Vic. 3505 03 50513100 email [email protected] Key personnel Caroline Tarr, David Emanuelli, Dr. Steve Sykes, Dr. Rob Walker Statement of Purpose The project was designed to provide a direct benefit to the Australian dried vine fruits industry. The research aimed to develop improved drying varieties with potential to overcome problems and or deficiencies of existing standard varieties. It was anticipated that the research would lead to an expanded export market with new niche products and provide the industry with new dried grape varieties which spread the risk of crop loss due to characteristics such as early maturity, disease resistance and adaptation to more efficient management practices. A ‘Unique Dried Grape Varieties Steering Committee’ represented by all industry sectors and HA Ltd. was established to ensure that the CSIRO program was consistent with industry needs and to identify selections with potential and facilitate their commercial adoption. Acknowledgement of funding sources Horticulture Australia Ltd. $ 341,670 CSIRO $ 453,000

Report finalized 31st December 2004

Disclaimer Any recommendations contained in this publication do not necessarily represent current HAL Limited policy. No person should act on the basis of the contents of this publication, whether as to matters of fact or opinion or other content, without first obtaining specific, independent professional advice in respect of the matters set out in this publication.

Table of Contents Media summary

3

Technical summary

4

Introduction

6

Materials and methods

7

Results Section 1. Investigations of disease resistance Section 2. Breeding strategies and in-ovulo embryo rescue Section 3. Evaluation of single vines, multiplied selections and imported varieties Section 4. Low browning sultanas Section 5. Tissue-cultured Bruce’s Sport trial Section 6. Sunmuscat rootstock trial Section 7. Shirana

9 11 15 19 20 22 25

Discussion

28

Technology transfer

28

Recommendations

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2

Media Summary In recent years problems in dried fruit production have highlighted the deficiencies of existing drying varieties, especially Sultana. These include substantial production and quality losses associated with rain damage at harvest, the development of mouldy fruit and related problems with toxins, and low fruitfulness associated with weather conditions during fruit bud initiation in November-December. To address these problems CSIRO, with support of Horticulture Australia Ltd. has maintained a structured and targeted breeding program aimed at developing rain and disease tolerant grape varieties for dried fruit production which meet market requirements. As part of these studies, production and dried fruit quality have been assessed from grapevine material at all stages of development. These include individual seedlings from various breeding populations, promising selections multiplied into larger test plots, established rootstock trials, imported varieties and selections established on grower properties for evaluation under commercial conditions. Seven rain or disease resistant sultana types have been identified for planting on a larger scale under semicommercial conditions while two selections, the early ripening disease resistant currant (M48-42) and the rain tolerant sultana type (F 64-74) have been moved to the final stages of evaluation with larger plantings established on commercial sites by top-working. Results from the Sunmuscat rootstock trial indicate that 1103 Paulsen should be the preferred rootstock while performance on 140 Ruggeri and Ramsey was adequate.

3

Technical Summary Problems in dried fruit production in recent years highlighted deficiencies of existing drying varieties, especially Sultana. To address these problems CSIRO, with support of Horticulture Australia Ltd. has maintained a structured and targeted breeding program to address key industry issues. These include development of rain tolerant selections to eliminate mould development and minimise Ochratoxin A levels; the need for new drying varieties for consistent production of high quality dried fruit products to meet buyer/ consumer specifications and disease resistant drying varieties to reduce chemical use, enhance marketing opportunities and meet consumer demands for ‘chemical free’ products. Enhanced knowledge of the inheritance of key production and quality characteristics has been incorporated into development of breeding strategies. Significant scientific contributions have been made to determine the causes of ‘seedlessness’, improve techniques for in-ovulo embryo rescue and in the development and application of laboratory based techniques for disease resistance screening. In-ovulo embryo rescue techniques are used to develop new hybrids from seedless x seedless grape crosses. Since 1998, 170 seedless x seedless combinations have been tried, of which 40 were unsuccessful and produced no progeny. In some cases this may be attributed to the fact that seedlessness can be due to pathenocarpy, where berries develop without fertilization and development of an embryo. Detailed investigations were undertaken with the aim to improve the efficiency of the embryo rescue. They included effects of genotype; the culture medium, including supplements of various minerals, plant growth regulators and other organic substances; and the effect of ovule age at removal on ovule elongation, embryo recovery, embryo growth and plantlet formation. As a result of this investigation, both the number of crosses undertaken in a season and survival rates at all stages of culture have increased, giving results comparable to overseas programs. Studies on the seedless character highlighted the need to understand the mechanism leading to the seedless phenotype in grapes in order to maximise breeding and selection strategies to develop new seedless varieties. Studies conducted with Carina showed that it is not only male sterile but also parthenocarpic and stenospermocarpic depending on the availability of viable pollen at flowering. Consequently, it can be used as a female parent in conjunction with in-ovulo embryo rescue techniques to transmit these characteristics. Development of disease resistant varieties has targeted powdery mildew (Uncinula necator) and downy mildew (Plasmopara viticola). A reliable method of using leafed single node cuttings to screen for downy mildew resistance has been developed and was used to study resistance in breeding lines. Depending on family, segregation of resistant to susceptible phenotypes fitted 1:15, 1:3 or 1:1 ratios. Similarly, screening for powdery mildew resistance using a protocol based on leaf disc assays and rooted single node cuttings showed segregation of resistant to susceptible phenotypes fitted 1:3 or 1:15 ratios. The information on disease resistance is used in the development of breeding strategies to develop new disease resistant dried grape varieties. Productivity and dried fruit quality was assessed for grapevine material at all stages of development (ie. seedlings in breeding populations, multiplied selections, established trials eg. the Sunmuscat and Bruce’s Sport rootstock trials), and imported varieties and selections were established in larger commercial sites (ie. Shirana and M48-042). A number of seedling selections have been identified for larger scale evaluation in multiplied plots, including grafting on rootstocks and top-working on existing vines to facilitate more rapid evaluation. Seven rain or disease resistant sultana types have been established in a larger scale, semi-commercial grower site. The early ripening disease resistant currant (M48-42) and the rain tolerant sultana type (F 64-74) have been moved to the final stages of evaluation with larger plantings established on commercial sites. Results from the Sunmuscat 4

rootstock trial showed that 1103 Paulsen produced the highest yields followed by 140 Ruggeri and Ramsey.

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Introduction Problems in dried fruit production highlight the deficiencies of existing drying varieties, especially Sultana. These include substantial production and quality losses associated with rain damage at harvest, the development of mouldy fruit and related problems with toxins, and low fruitfulness associated with weather conditions during fruit bud initiation in November-December. CSIRO Plant Industry maintains a vine improvement program to provide the viticultural industries with material suited to Australian conditions and industry needs. As part of this program new seedless drying varieties have been developed by hybridisation or have been imported and maintained in the CSIRO germplasm collection. Varieties released for dried fruit production from the breeding program include Carina (1975), Merbein Seedless (1981), Marroo Seedless (1988) and Sunmuscat (1997). Previous studies evaluated the range of seedless varieties in the germplasm collection (Newman and Clingeleffer 1987) and identified some large berried types as potential alternatives to seedless raisin varieties or specialty lines. The best of the large berried types were included in CSH17 (‘Integration of alternative drying varieties with low input, production systems of management’). DFRDC provided on going support in 1992-1998 for trial establishment and evaluation of small and large berried sultana types, new currant types, specialty lines, new raisin selections (included from DAV76 project) and disease resistant or breeding lines from in-ovulo embryo rescue (CSH24 and 54). In the 1998-2001 triennium the DFRDC project (CSH 60D, ‘Evaluation of new seedless drying grapes, selected for disease resistance and market requirements’) completed the evaluation of larger, semi-commercial plantings of currants (including muscat types), large berried selections and new seedless raisin types. Larger plantings of seedless selections from disease resistant and in-ovulo embryo breeding lines, recent imports (Black Emerald, C88-89) and low browning transgenic Sultanas were initiated. The evaluation of Sunmuscat grafted on a range of rootstocks also commenced. The import of two new USDA varieties, ie. the early ripening DoVine and Summer Muscat was facilitated through quarantine. Two new initiatives were implemented in the project with the aim to enable smarter approaches to breeding new drying varieties. These were the evaluation of disease resistant breeding lines using laboratory based and non-sprayed field assays and the conduct of inheritance studies with selected populations of CSIRO breeding lines for important dried fruit quality characteristics, ie. seedlessness, yield, berry size and colour, sugar level, etc. To ensure that the dried fruit breeding strategies are consistent with industry requirements, CSIRO has maintained close consultation with industry. Key forums were held in June 1992 and September 1997 with researchers and representatives from all industry sectors, i.e. production, processing and marketing sectors. These forums identified the limitations of existing varieties and set specifications for desirable characteristics for sultana types, currant types, raisins and other types suited to high value products and niche marketing. A ‘Unique Dried Grape Varieties Steering Committee’ has been formed to ensure that the CSIRO program is consistent with industry needs, to identify selections with most potential and to facilitate their commercial adoption. Members of this committee include representatives from all industry sectors, HA Ltd and researchers. A sub-committee was formed to specifically deal with the final evaluation and commercialisation of the CSIRO selection S67 (now Shirana) with PBR protection, which at the time of project commencement was being grown by 4 growers. S67 is a small-berried sultana type that produces a uniform amber dried product. It has proved to be very rain tolerant and has given consistent production. The Riverlink/DFRDC workshop (Breeding and Selection Strategy for the Dried Grape Industry) held in September 1997 recognised the need to implement a procedure for testing the disease 6

resistance/tolerance of selections and breeding lines to the major fungal diseases, i.e. powdery and downy mildew. The workshop also recognised that breeding strategies could be improved if inheritance of the key production and quality characteristics was understood. Knowledge of the genetic control of important characteristics obtained from inheritance studies has provided immense benefit in breeding field crops, annual and perennial horticulture crops (eg. inheritance of seedlessness in citrus and disease resistance and fruit quality traits such as sugar level in melons). The workshop outcomes and the strategy developed formed the basis for developing the previous 1998-2001 project and formed the foundation for determining key objectives for this project. Initially, the disease resistance testing has concentrated on powdery mildew assessment. Laboratory and glasshouse screening of advanced selections with potential for disease resistance has shown that 13 out of 15 selections with drying potential were tolerant of powdery mildew. Laboratory based screening of 2 seedling populations commenced in spring 2000. Preliminary results suggest that powdery mildew resistance mechanisms are under polygenic control. With respect to inheritance of key dried fruit characteristics, analysis of data from more than 5000 seedlings with a diverse range of parents was undertaken. The results indicate that berry weight is strongly inherited while seedlessness, sugar level and acidity have moderate levels of inheritance. Bunch weight, vigour and yield had low levels of inheritance. Key trials established previously which were assessed for yield and quality in this project included:• • • • • •

larger plantings of seedless selections with high potential derived from disease resistant and inovulo embryo breeding lines (40 selections on Shaw and T-trellis) Bruces Sport (tissue-cultured to eliminate leaf roll virus) on own roots and Ramsey potentially low browning transgenic Sultanas with down-regulated PPO activity plantings of the USDA selections, Black Emerald and C88-89 (a muscat seedless) a non-sprayed site with disease resistant selections to test field tolerance/ resistance by observation of leaves and fruit. Black spot, downy mildew and powdery mildew occurred in this site in spring, 2000 a rootstock trial with Sunmuscat on the property of Mr. A. Martin at Merbein which produced its first crop in 1999

New plantings identified for establishment in this project will include:• • • •

plantings of DoVine and Summer Muscat on completion of their release from quarantine larger commercial plantings of the most promising early ripening and disease tolerant lines as the final step to commercialisation, selected with input from the steering committee replicated trials of selections identified for commercialisation for Plant Breeders Rights (PBR) purposes extension of the non-sprayed site with promising disease resistant selections identified from seedling populations as part of the laboratory based testing and inheritance studies.

7

Materials and Methods Drying and fruit quality assessments Routine assessments of the range of material available to this project included:• • • • •

pre-harvest sampling for berry weight, sugar, acid and pH determination. measurement of yield, and if appropriate bunch number drying as naturals and/or after treatment with drying emulsion on racks, by trellis drying or dehydration. processing in the small-scale, CSIRO packing plant. product evaluation, including colour and its uniformity, moisture, berry size, flavour etc., processing and storage characteristics. Samples of the best selections were submitted to the ‘Unique Dried Grape Varieties Steering Committee’ and where appropriate to processors and marketers for preliminary market testing.

Trial sites from which fruit was evaluated • larger plantings of most promising selections established in the previous project, CSH60, (2nd stage) including seedless selections from disease resistant and in-ovulo embryo breeding lines. These sites include management on Shaw swing-arm trellis to assess mechanisation potential of selections. • seedless single vine selections from early ripening and disease resistant lines and consolidated plantings of in-ovulo embryo plantings • Sunmuscat rootstock trial established on the property of Mr. A. Martin which produced some fruit in 1999 and a substantial crop in season 2000 (Clingeleffer 2000). The trial included a range of rootstocks aiming to reduce the partial incompatibility found with Ramsey in the previous project (CSH 54). • Bruce’s Sport (tissue-cultured to eliminate leaf roll virus) rootstock/trellis trial which produced the first crop in 2001. Bruce’s Sport is being assessed because of its low browning potential. The original material was severely affected by leaf roll viruses. • low browning transgenic Sultanas with down-regulated PPO activity. The first bunches from some plants were harvested and dried in season 2000/2001. • plantings of recent imports established in spring 2000 (ie. Black Emerald and C88-89) • disease resistant selections planted at the CSIRO Koorlong site that are not sprayed with fungicides. Field tolerance/ resistance of these selections to the major fungal pathogens will be assessed by observation of leaves and fruit. • commercial grower plantings of S67 (Shirana), including establishment on various trellises and rootstocks. Establishment of new trial sites • plantings of DoVine and Summer Muscat, and potentially, Diamond Muscat and Princess (previously called Melissa) from the USDA upon release from quarantine. • second stage evaluation plantings (8 vine plots) of the most promising selections identified among current seedling populations • larger commercial grower plantings of the most promising early ripening and disease tolerant lines as the final step to commercialisation. • replicated trials of selections identified for commercialisation to enable data collection for Plant Breeders Rights (PBR) purposes.

8

Inheritance studies Data collected in the previous project (CSH60) for important dried fruit quality characteristics, eg. seedlessness, yield, berry size and colour, maturity time and sugar level across all seedling progeny of selected populations, were augmented by assessment of new populations planted in recent years. A key focus will be to complete the inheritance studies for disease resistance using laboratory based assays for powdery mildew and downy mildew. Laboratory based screening techniques for disease resistance Protocols for laboratory techniques to screen selections for resistance to powdery mildew were developed, with input from Dr. I. Dry at the Adelaide laboratory, in the previous project (CSH60). The technique was compared to glasshouse screening results for the most promising disease resistant selections and was applied across seedling populations as part of the inheritance of disease resistance studies. A laboratory-based assay that used leaves sampled from the field was used to assess downy mildew resistance of advanced selections and breeding lines. Further breeding Breeding strategies to meet industry requirements have been implemented by CSIRO based on enhanced knowledge of the inheritance of key dried fruit quality characteristics and disease resistance using in-ovulo embryo rescue techniques for seedless x seedless crosses. Milestones established at commencement of the project • evaluation of CSIRO disease resistant advanced selections for both dried fruit quality and disease resistance using laboratory-based and non-sprayed field assays will be completed by September 2002 • inheritance studies of disease resistance and dried fruit quality characteristics with selected populations of CSIRO breeding lines will completed by June 2004 • breeding strategies, based on enhanced knowledge of the inheritance of key quality characteristics and disease resistance using in-ovulo embryo rescue techniques for seedless x seedless crosses will be implemented at flowering each season • evaluation of plantings established in the previous project will be completed each season by June 30th, ie. selections from disease resistant, in-ovulo embryo breeding lines and imports (eg. Black Emerald, C88-89) • evaluation of low browning transgenic Sultanas will be completed each season by June 30th • plantings will be established of DoVine (spring 2001) and Summer Muscat (spring 2002) for evaluation as drying grapes once release from quarantine is completed • evaluation of the newly established rootstock trial with tissue-cultured Bruces Sport will commence in season 2001and recommendations will be finalised by June 30th, 2004. • evaluation of the Sunmuscat rootstock trial trained to Shaw Swing-arm trellis on a grower property will be undertaken each season and recommendations finalised by June 30th, 2004. • larger commercial plantings of the most promising early ripening and disease tolerant lines as the final step to commercialisation will be developed based on annual recommendations of the ‘unique dried grape steering committee’ • Plant Breeders Rights (PBR) for S67 will be completed by September 2002 • continuing and annual progress reports will be completed each year by December 15 and July 30, respectively

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Results Section 1. Investigations of Disease Resistance A long-term objective of the dried grape breeding program is the development of disease resistant varieties with the aim to minimise the use of agri-chemicals and hence, lower production costs, reduce environmental impacts and meet consumer demands for chemical-free products. The main diseases targeted are powdery mildew (Uncinula necator) and downy mildew (Plasmopara viticola), which originate from the American continent. A diverse range of disease resistant genotypes have been and are being used as parents. In some cases, crosses over a number of generations have produced complex pedigrees in the breeding lines. Parents used have included V. rotundifolia, V. cinerea, V. caribaea, V. longii, V. aestivalis and V. labrusca as well as hybrids with pedigrees incorporating many species. Progeny from ‘disease resistant’ crosses bred involving use of these complex species hybrids as parents have been developed and assessed for tolerance of key fungal diseases under controlled laboratory conditions. The information on disease resistance is used in the development of breeding strategies to develop new disease resistant dried grape varieties. Downy mildew A method involving use of leafed single node cuttings to screen for downy mildew resistance has been developed. This quantifies the intensity of leaf chlorosis and sporulation, and records the necrotic response of infected leaves under laboratory conditions (Liu et al. 2003b). In optimising this technique, effects of leaf maturity and sporangia suspension concentrations on infection and symptom expression were examined. The improved reliability of the method relative to a leaf disc method was demonstrated by assaying a range of commercial varieties and a number of hybrids from CSIRO’s breeding program. Resistance to downy mildew between and within seven full-sib grapevine families was investigated under controlled culture conditions. The families were generated by crossing resistant x susceptible and susceptible x susceptible genotypes. Family size ranged between 19 and 89 vines, which were aged 9-to-10 years-old. Leaf infection following downy mildew inoculation was assessed using the cultured leafed single node cuttings method (Liu et. al. 2003b). The severity of disease infection 7 days after inoculation was based on ranking leaf chlorosis and sporulation using a 1-to-9 scale where 1 represented no visible sign of infection and 9 where > 80% of the leaf was infected. A measure of the hypersensitive response (HR) was also used to rank vine reaction to inoculation. Hybrids rated as ≤ 3 in the severity ratings were classified as downy mildew resistant. The proportion of vines within families scored as resistant and hence no leaf chlorosis ranged from 4.6% to 22.5% and for sporulation from 4.6% to 47.4%. There was a strong relationship between leaf chlorosis and sporulation expression within each cross. Depending on family, segregation of resistant to susceptible phenotypes fitted 1:15, 1:3 or 1:1 ratios. In families where the hypersensitive reaction was observed, segregation for resistant to susceptible phenotypes fitted 1:1 or 1:3 ratios. Where vines within families displayed the hypersensitive reaction, lower mean ratings for leaf chlorosis and sporulation were observed. These results are being prepared for publication. Powdery mildew Five hybrid families were screened for powdery mildew resistance using a laboratory based protocol based on leaf disc assays and rooted single node cuttings. The same vines were also screened for 10

disease incidence/infection in an unsprayed vineyard using an established field based scoring method. The families were generated by crossing susceptible x susceptible genotypes and family size ranged between 19 and 81 vines. The severity of disease infection was based on the rate of the coverage of mycelium and sporulation (10 to 12 days after inoculation for laboratory tests) using a 1-to-9 scale where 1 represented no visible sign of infection and 9 where > 80% of the leaf surface or disc was infected. Within families, the percentage of hybrids having a disease severity score ≤ 3.0 ranged from 0% to 25%. Individual hybrids within families were grouped as either resistant (disease severity rating ≤ 3.0) or susceptible (disease severity rating > 3.0). Segregation of hybrids based on this grouping scheme fitted either a 1 i.e. resistant (R) to 3 i.e. susceptible (S) or a 1R:15S ratio within families. As for the downy mildew results, these data are being prepared for publication and will be used in developing future breeding strategies. Conclusion The disease screening results show that development of varieties, which are tolerant or resistant to both powdery and downy mildew is achievable. Among current hybrid populations screened so far there are no lines completely resistant to powdery mildew but some that have complete resistance to downy mildew. While resistance to both mildews appears to be governed by 4 genes (unpublished data), correlations between powdery and downy mildew resistance were weak.

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Section 2. Breeding strategies and in-ovulo embryo rescue Outputs from studies to understand the inheritance of key dried fruit characters were reported in the previous report (CSH 60D). This information has been used in the development of breeding strategies for dried fruit types taking into account characteristics, which meet grower, processing and marketing requirements and are documented in specification manuals determined for the range of dried fruit product types. Significant contributions to improvements in in-ovulo embryo rescue techniques and to our understanding of ‘seedlessness’ in Carina currant and how this knowledge may be applied in development of breeding strategies have been made in this project. While the emphasis has been placed on in-ovulo embryo rescue with crosses between seedless types, conventional approaches using one seeded parent have also been incorporated in the strategies. Improvements in embryo rescue Breeding strategies for development of new drying varieties have been integrated with similar approaches for the development of seedless table grapes. In order to improve the efficiency of breeding seedless varieties, in-ovulo embryo rescue techniques are used to develop new hybrids from seedless x seedless grape crosses (Clingeleffer et. al. 2003). Under in-vitro conditions (ie. in tissue culture), embryos which would normally abort, continue to develop and can be established as normal plants in the research vineyard. Since 1998, 170 seedless x seedless combinations have been tried, of which 40 were unsuccessful and produced no progeny. In some cases this may be attributed to the fact that seedlessness can be due to pathenocarpy, where berry development occurs without fertilization and development of an embryo. Over the past 4 years, detailed investigations have been undertaken with the aim to improve the efficiency of the embryo rescue technique using the germplasm base at Merbein. These studies reported in detail by Liu et al. (2003) have included effects of specific genotypes (ie. the parents selected); the culture medium, including supplements of various minerals, plant growth regulators and other organic substances; and the effect of ovule age at removal on ovule elongation, embryo recovery, embryo growth and plantlet formation. As a result of this investigation, both the number of crosses undertaken in a season and survival rates at all stages of culture (ie. ovule culture, embryo rescue and plant acclimatisation) have increased, giving results comparable to overseas programs. For example in season 2002/2003, 89 seedless x seedless crosses were carried out in the field. From the berries that developed, 13,400 ovules were cultured and 3200 embryos were dissected out (ie. a recovery rate of 24%) of which 3125 have survived (ie. a 97% embryo survival rate). About 1400 plants have been established from the recovered embryos with an expected conversion rate of greater than 50% once propagation is completed. It should be noted that none of the progeny from recent embryo rescue produced a crop during this project.

12

Conventional breeding and embryo rescue methodology include: Emasculation of bunch primordia in the field Bagging bunches to exclude pollen other than desired parent pollen Pollination of emasculated bunches

Berry collection at veraison (berry softening) and removal of embryos

Then invitro plant growth

Planted seedlings in glasshouse and shadehouse.

13

Studies of ‘Seedlessness’ in Carina Enhanced understanding of factors contributing to seedless grape types is essential in developing breeding strategies for this character, particularly those based on in-ovulo embryo rescue from seedless x seedless crosses. Generally, berry set in hermaphrodite grape varieties is the result of ovule fertilization. However with seedless varieties, a number of mechanisms may be involved. Parthenocarpic varieties, such as Black Corinth, produce seedless berries following pollination, which stimulates fruit set, but without ovule fertilization. Stenospermocarpic types set seedless berries after ovule fertilisation, but ovules remain soft and fail to develop for reasons such as embryo abortion, early degradation of the nucellus in the embryo sac and inherent ovule abnormalities (Liu et al., submitted) Consequently a study was conducted to investigate pollen viability, berry set and seed status of Carina. It included assessing effects of self- and cross-pollination on in-ovulo embryo rescue and relationships between berry size and ovule development (Liu et al., submitted). Carina currant is a significant dried grape variety grown in Australia. Its yield and fruit quality, however, are restricted by pollination and can be improved by judicious use of hormone-based setting sprays. It produces abundant seedless berries, but berry size is small when unpollinated, and even when pollen is available berry size can be variable. Release of the calyptra, the term for the fused corolla shed at blooming in vines at anthesis is poor for Carina, especially during wet seasons. It was unclear whether Carina is parthenocarpic, stenospermocarpic or both, and also if it is male fertile and its pollen self-functional. Pollen fertility in Carina had not been documented and thus its value as a male parent for breeding seedless varieties was unknown. Results Carina pollen failed to germinate in-vitro and gave poor berry set when used to pollinate Hunisa and Kishmishi. Percentage berry set in Carina was unaffected by pollination treatments, but crosspollination increased berry size (Table 2.1). Larger seed traces (>1.2 mm in length) were present in most cross-pollinated Carina berries and their size and number were correlated positively with berry size. Table 2.1. Effect of self-, cross- and open-pollination on the percentages of large (L), medium (M) and small (S) berries in Carina inflorescences (adapted from Lui et al.) Total Berry diameter (mm)A (%) No ≥ 11.0 (L) ≥ 8.0