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New disease-resistant, seedless grapes are developed using embryo rescue and molecular markers

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Abstract

Improving the operation of embryo rescue technology is a highly efficient way to breed new high-quality cultivars of seedless, disease-resistant grapes when using a stenospermocarpic Vitis vinifera L. (female parents) hybrid with Chinese wild Vitis (male parents). In this study, sampling time had a significant effect on embryo recovery. Four crosses were investigated to improve the embryo rescue efficiency by determining the best sampling time, which was found to be 40, 50, 55, and 57 days after pollination (DAP) for ‘Flame seedless’ × ‘Ruby seedless’, ‘Kunxiang seedless’ × ‘Flame seedless’, ‘Ruby seedless’ self-pollination and ‘Ruby seedless’ × ‘Flame seedless’, respectively. The highest percentage embryo germination ranged from 60.83 to 89.10% for four cross-combinations when the embryo was germinated on woody plant medium (WPM), with 1.0-μM thidiazuron (TDZ, a potent cytokinin). For 11 cross-combinations, the embryo recovery rate varied from 4.8 to 29.6% and the plant development rate varied from 17.1 to 78.9%. ‘Ruby seedless’ was the best female parent followed by ‘Flame seedless’. The Grape Seedless gene Probe 1 (GSLP1) and SCF27 molecular markers were used to assist the identification of the seedless traits of the 388 progeny. A total of 309 plantlets were amplified, with the specific bands at 569-bp and 2k-bp being preliminarily considered as seedless. The S382-615 marker was used to identify the downy mildew resistance of the ‘Ruby seedless’ × ‘Beichun’ cross. Two F1 progeny were amplified, with the specific band at 615-bp being identified as downy mildew resistance.

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Abbreviations

IAA:

Indole-3-acetic acid

6-BA:

6-Benzyladenine

MS:

Murashige and Skoog (1962) medium

WPM:

Woody plant medium

AC:

Activated charcoal

TDZ:

Thidiazuron

PGR:

Plant growth regulator

DAP:

Days after pollination

SCAR:

Sequence characterized amplified region

RAPD:

Random amplified polymorphic DNA

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Acknowledgements

This research at the State Key Laboratory of Crop Stress Biology in Arid Areas, Northwest A&F University, was supported by the earmarked fund for China Agriculture Research Systems for the grape industry (Grant No. CARS-29-yc-3). The authors specifically thank Dr Alexander (Sandy) Lang from RESCRIPT Co. (New Zealand) for useful language editing.

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Authors and Affiliations

  1. College of Horticulture, Northwest A&F University, Yangling, 712100, Shaanxi, People’s Republic of China

    Shasha Li, Zhiying Li, Yanan Zhao, Jiong Zhao & Yuejin Wang

  2. State Key Laboratory of Crop Stress Biology for Arid Areas, College of Horticulture, Northwest A&F University, Yangling, 712100, Shaanxi, People’s Republic of China

    Shasha Li, Zhiying Li, Yanan Zhao, Jiong Zhao & Yuejin Wang

  3. Key Laboratory of Horticultural Plant Biology and Germplasm Innovation in Northwest China, Ministry of Agriculture, College of Horticulture, Northwest A&F University, Yangling, 712100, Shaanxi, People’s Republic of China

    Shasha Li, Zhiying Li, Yanan Zhao, Jiong Zhao & Yuejin Wang

  4. Research Institute of Xinjiang Grape, Melon and Fruit, Shanshan, 838200, Xinjiang, People’s Republic of China

    Qiangwei Luo

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  1. Shasha Li
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13205_2019_1993_MOESM1_ESM.doc

Supplementary material 1: Supplementary Fig. S1. Analysis of amplification by the GSLP1 marker showing the linkage to seedlessness in parents. M. DNA ladder marker (Trans DNA 2K plus), (1) ‘Beichun’, (2) ‘Shuangyou’, (3) ‘Zixiang’, (4) ‘Centennial seedless’, (5) ‘Crimson seedless’, (6) ‘Flame seedless’, (7) ‘Kunxiang’, (8) ‘Ruby seedless’, (9) ‘Huozhouhongyu’. (+) the 569-bp specific band is present; (−) the 569-bp specific band is absent. Supplementary Fig. S2. Analysis of the amplification by the SCF27 marker showing the linkage to seedlessness in parents. M. DNA ladder marker (Trans DNA 2K plus), (1) ‘Beichun’, (2) ‘Shuangyou’, (3) ‘Zixiang’, (4) ‘Centennial seedless’, (5) ‘Crimson seedless’, (6) ‘Flame seedless’, (7) ‘Kunxiang’, (8) ‘Ruby seedless’, (9) ‘Huozhouhongyu’. (+) the 569-bp specific band is present; (−) the 569-bp specific band is absent. Supplementary Fig. S3. Amplification results for GSLP1 linked to the seedless gene in ‘Flame seedless’ × ‘Shuangyou’ hybrid progeny. M. DNA ladder marker (Trans DNA 2K plus), (1) ‘Flame seedless’, (2) ‘Shuangyou’, (3–25) the 23 hybrid progeny of ‘Flame seedless × Shuangyou’. (+) the 569-bp specific band is present, (−) the 569-bp specific band is absent. Supplementary Fig. S4. Amplification results for SCF27 linked to the seedless gene in ‘Kunxiang seedless’ × ‘Flame seedless’ hybrid progeny. M. DNA ladder marker (Trans DNA 2K plus), (1) ‘Kunxiang seedless’, (2) ‘Flame seedless’, (3–28) the 26 progeny of ‘Kunxiang seedless’ × ‘Flame seedless’. (+) the 2k-bp specific band is present; (−) the 2k-bp specific band is absent. Supplementary Fig. S5. Amplification results for SCF27 linked to the seedless gene in ‘Ruby seedless’ × ‘Flame seedless’ hybrid progeny. M. DNA ladder marker (Trans DNA 2K plus), (1) ‘Ruby seedless’, (2) ‘Flame seedless’, (3–341) the 339 progenies of the ‘Ruby seedless’ × ‘Flame seedless’. (+) the 2k-bp specific band is present; (–) the 2k-bp specific band is absent. Supplementary Fig. S6. Analysis of the amplification by the S382-615 marker showing linkage to disease-resistance in parents. M. DNA ladder marker (Trans DNA 2K plus), (1) ‘Beichun’, (2) ‘Shuangyou’, (3) ‘Zixiang’, (4) ‘Centennial seedless’, (5) ‘Crimson seedless’, (6) ‘Flame seedless’, (7) ‘Kunxiang’, (8) ‘Ruby seedless’, (9) ‘Huozhouhongyu’. (+) the 569-bp specific band is present; (−) the 569-bp specific band is absent. Supplementary Fig. S7. Amplification results for S382-615 linked to disease-resistance in ‘Ruby seedless’ × ‘Beichun’ hybrid progeny. M. DNA ladder marker (Trans DNA 2K plus), (1–34) The 34 progenies of the ‘Ruby seedless’ × ‘Beichun’. (+) the 2k-bp specific band is present; (−) the 2k-bp specific band is absent. (DOC 1978 kb)

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Li, S., Li, Z., Zhao, Y. et al. New disease-resistant, seedless grapes are developed using embryo rescue and molecular markers. 3 Biotech 10, 4 (2020). https://doi.org/10.1007/s13205-019-1993-0

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