Diagnostic KASP markers for Flower sex and stenospermocarpic seedlessness in diverse vitis, muscadinia, and wide hybrid populations

Authors: VAUGHN, ISABELLA - University Arkansas For Medical Sciences (UAMS); ZOU, CHENG - Cornell University; Cadle Davidson, Lance; JOHNS, CARMEN - University Of Arkansas; NELSON, LACY - University Arkansas For Medical Sciences (UAMS); COUSINS, PETER - E & J Gallo Winery; HEINITZ, CLAIRE - US Department Of Agriculture (USDA); BLOODWORTH, JEFF - Gardens Alive!; SUN, QI - Cornell University; WORTHINGTON, MARGARET - University Arkansas For Medical Sciences (UAMS)

Submitted to: HortScience
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 1/24/2026
Publication Date: 3/6/2026
Citation: Vaughn, I., Zou, C., Cadle Davidson, L.E., Johns, C., Nelson, L., Cousins, P., Heinitz, C., Bloodworth, J., Sun, Q., Worthington, M. 2026. Diagnostic KASP markers for Flower sex and stenospermocarpic seedlessness in diverse vitis, muscadinia, and wide hybrid populations . HortScience. Volume 151: Issue 3 Pages 147-160. https://journals.ashs.org/view/journals/jashs/151/3/article-p147.xml.
DOI: https://doi.org/10.21273/JASHS05580-25

Interpretive Summary: Grapes grown in the United States come from two related groups: Vitis, which includes most table, juice, and wine grapes, and Muscadinia, which includes muscadine grapes. Both groups are important for fresh eating and processing, and breeders are working to improve traits that consumers and growers want. Two traits that are especially valuable are seedlessness and perfect flowers (flowers that pollinate themselves and reliably make fruit). Seedless grapes usually come from a natural process where seeds start to form but stop developing early. This trait has recently been brought from common grapes into muscadines through traditional cross-pollination, even though the two groups have different chromosome numbers. However, breeders have not yet had dependable DNA tests to identify seedless muscadine seedlings at a young age. Wild grapes in both groups also normally have separate male and female plants, but forms with perfect flowers were selected long ago because they make more consistent fruit. A DNA test for flower type has also not been available for muscadines and hybrids. In this study, we developed and tested two low-cost DNA markers that can predict whether a young grapevine will be seedless or have perfect flowers, long before the plant produces fruit. We tested these markers on more than 1,100 vines, including muscadines, wine and table grapes, wild species, and hybrids from the University of Arkansas breeding program. The seedless and flower-type markers predicted these traits correctly in 99.7% and 100% of the vines tested, respectively. These new DNA tools will help grape breeders select the best seedlings early in the breeding process, saving time, money, and field space, and speeding up the development of improved seedless and reliable-fruiting grape varieties for growers and consumers.

Technical Abstract: The genus Vitis is composed of two subgenera, Vitis (2n = 38) and Muscadinia (2n = 40), which are both cultivated for fresh-market, juice, and wine industries. Stenospermocarpic seedlessness and perfect-flowered vines are highly desired traits in both Vitis vinifera and muscadine (Muscadinia rotundifolia) breeding programs. Stenospermocarpy has recently been introgressed from V. vinifera to muscadines through conventional breeding despite their differing chromosome number, but no molecular markers for this trait have been validated in muscadine germplasm. Wild vines in both subgenera are dioecious, but perfect-flowered forms were independently selected during domestication to enhance reproductive efficiency and fruit production. This study reports the development and validation of two Kompetitive Allele Specific PCR (KASP) markers targeting causal polymorphisms within candidate genes for male sterility (VviINP1) and stenospermocarpy (VviAGL11). Sequence alignments with published M. rotundifolia genomes suggested that the seedless_Arg197Leu_site56.fas and female_INP_indel_site56.fas KASP markers might be broadly effective across diverse species within Vitis and Muscadinia. Marker performance was evaluated using a validation panel including 918 Vitis × Muscadinia hybrid seedlings from the University of Arkansas Fruit Breeding Program and a diverse set of cultivated and wild accessions (209 accessions evaluated with the seedless marker and 315 accessions with the flower sex marker). After excluding incomplete phenotype and genotype data, the stenospermocarpic marker (seedless_Arg197Leu_site56.fas) accurately predicted seedlessness in 921 of 924 (99.7%) entries. Additionally, 148 out of 203 seedlings that failed to produce fruit across both growing seasons were predicted to be stenospermocarpic with the seedless KASP marker, suggesting that seedless Vitis × Muscadinia hybrids may have partial sterility or lack cold hardiness. The flower sex marker (female_INP_indel_site56.fas) correctly predicted flower sex in all 1,138 (100%) entries. Together, these KASP markers provide highly accurate, cost-effective tools for early selection of seedless and perfect-flowered genotypes across Vitis, Muscadinia, and hybrid breeding programs.