A medium-density genotyping platform for cultivated strawberry using DArTag technology

Authors: Hardigan, Michael; FELDMANN, MITCHELL - University Of California; CARLING, JASON - Diversity Arrays Technology; ZHU, ANYU - Diversity Arrays Technology; KILIAN, ANDRZEJ - Diversity Arrays Technology; FAMULA, RANDI - University Of California; COLE, GLENN - University Of California; KNAPP, STEVEN - University Of California

Submitted to: The Plant Genome
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 9/22/2023
Publication Date: 11/8/2023
Citation: Hardigan, M.A., Feldmann, M.J., Carling, J., Zhu, A., Kilian, A., Famula, R.F., Cole, G.S., Knapp, S.J. 2023. A medium-density genotyping platform for cultivated strawberry using DArTag technology. The Plant Genome. 16(4). Article e20399. https://doi.org/10.1002/tpg2.20399.
DOI: https://doi.org/10.1002/tpg2.20399

Interpretive Summary: Genomic prediction should be a part of the strawberry molecular breeding toolbox. However, molecular breeding of small fruit crops such as strawberry (F. × ananassa) has been hampered by a combination of complex polyploid genomes and the lack of economy of scale for marker genotyping services that helps drive down costs for crops with greater sample volumes. Genotyping populations containing hundreds or thousands of individuals remains prohibitively expensive for most research programs. We have addressed this shortfall by developing 3K and 5K marker genotyping panels for use with Diversity Arrays Technology (DArT)'s 'DArTag' genotyping platform. These marker panels were designed to target subgenome-specific loci for disentangling the four subgenomes found within octoploid strawberry for better ease of use and interpretation by researchers, and with a cost of roughly $10/sample, offers a more scalable genotyping solution for genomic prediction than current "SNP chip" platforms and a more reproducible solution than genotyping-by-sequencing (GBS) due to its use of amplicons for targeting conserved regions of the genome. The performance of the marker panels has been validated and will assist public and private research programs in the transition from conventional to marker-assisted breeding.

Technical Abstract: Genomic prediction in breeding populations containing hundreds to thousands of parents and seedlings is prohibitively expensive with current high-density genetic marker platforms designed for strawberry. We developed mid-density pan- els of molecular inversion probes (MIPs) to be deployed with the ‘DArTag’ marker platform to provide a low-cost, high-throughput genotyping solution for strawberry genomic prediction. In total, 7,742 target SNP regions were used to generate MIP assays that were tested with a screening panel of 376 octoploid Fragaria accessions. We evaluated the performance of DArTag assays based on genotype segregation, amplicon coverage, and their ability to produce subgenome-specific amplicon alignments to the FaRR1 assembly and subsequent alignment-based variant calls with strong concordance to DArT’s alignment-free, count-based genotype reports. We used a combination of marker performance metrics and physical distribution in the FaRR1 assembly to select 3K and 5K production panels for genotyping of large strawberry populations. We show that the 3K and 5K DArTag panels are able to target and amplify homologous alleles within subgenomic sequences with low amplification bias between reference and alternate alleles, supporting accurate genotype calling while producing marker genotypes that can be treated as functionally diploid for quantitative genetic analysis. The 3K and 5K target SNPs show high levels of polymorphism in diverse F. × ananassa germplasm and UC Davis cultivars, with mean pairwise diversity (p) estimates of 0.40 and 0.32, and mean heterozygous genotype frequencies of 0.35 and 0.33, respectively.