DOI: 10.5061/dryad.m905qfv9w
title: Data from: Limitations of ad hoc genotyping in detecting ash dieback tolerance in <em>Fraxinus excelsior</em>

description: The data presented here was generated to study common ash tolerance to Ash
 dieback (ADB), caused by the invasive alien fungal pathogen Hymenoscyphus
 fraxineus. The data was generated from a field trial aiming at the
 identification of ADB tolerant genotypes while encompassing the genetic
 diversity of common ash in Austria. Over 35,000 progenies from 611
 putatively tolerant mother trees and a selection of 71 clones from two
 clonal seed orchards were assessed for ADB symptoms over a period of three
 years. One tolerant and one susceptible progeny from a subset of the
 mother tree range (570 trees) were genotyped using the 4TREE array, a
 genotyping tool developed specifically to screen common ash populations
 for their tolerance to ADB.
description: Please refer to this publication for a detailed description of
 the experimental design: Szukala, A., Unger, G.M.,
 Trujillo-Moya C., Geburek, T., Kirisits, T., Schueler, S., and Konrad, H.
 (2025). Limitations of <em>ad hoc</em> genotyping in detecting
 ash dieback tolerance in <em>Fraxinus excelsior</em>.
 <em>Tree genetics & Genomes</em>, doi:
 10.1007/s11295-025-01725-6 Summary:
 To establish a sufficiently large breeding base of resistant
 clones, we selected 611 putatively ADB tolerant and seed-bearing single
 trees (i.e., functional females) of <em>F. excelsior</em>
 distributed in forests across Austria. Forest stands were selected based
 on the rare presence of remarkably healthy or only slightly damaged trees
 despite these being surrounded by a high number of infected and heavily
 damaged or dead individuals, whereas sites in rural settings or in areas
 with low disease incidence were avoided because of insufficient infection
 pressure. Seeds were harvested in 2015 and 2017 from
 the selected ash trees (i.e., putatively ADB tolerant mothers, with the
 only exception of few mothers growing in the seed orchards). From the sown
 seeds 35,718 seedlings germinated and were reared in plastic
 multi-container blocks in the experimental nursery of the Austrian
 Research Centre for Forests (BFW) in Tulln an der Donau (Lower Austria).
 To systematically assess the susceptibility to ADB over a three-year
 period, the one-year-old offspring (on average about 50 seedlings per
 mother tree) were planted in a 1.5 ha field trial in a completely
 randomized block-design. To increase the level of natural infection
 likelihood and thus the selection pressure on the seedlings, ash leaf
 litter (i.e., pseudosclerotial ash leaf material collected in heavily
 infested ash stands) was placed between seedlings.
 <em>Fraxinus excelsior</em> is characterized by
 complex seed dormancy: seeds harvested in 2015 germinated during the two
 following years, in 2016 and 2017, while those harvested in 2017
 germinated in 2018 and 2019. Thus, the field trial included four sets of
 seedlings germinated during four subsequent years (2016 to 2019) and
 planted in four separate but adjacently arranged sub-trials, named
 hereafter R1 (established in 2017), R2 (established in 2018), R3
 (established in 2019) and R4 (established in 2020; not investigated in the
 present study). Progeny of sub-trials R1, R2, R3 and R4 were assessed for
 their susceptibility to ADB (intensity/degree of ADB) annually over a
 period of three years. Leaf samples were collected from
 all offspring seedlings during the years of trial establishment, dried on
 silica gel and stored at room temperature until DNA extraction. For the
 present study, offspring from 570 ash mother trees in the sub-trials R1,
 R2 and R3 were selected for genotyping. These included 503 mother trees
 from forests, 22 from the seed orchard in Feldkirchen (three assessed as
 tolerant, five as intermediate and 14 as susceptible) and 45 from the seed
 orchard in Bad Gleichenberg (7 assessed as tolerant, 9 as intermediate and
 29 as susceptible). Where possible, one completely healthy progeny (i.e.,
 DC = 1) and a dead one (i.e., DC = 6) from each of the 570 mother trees
 were sampled for genotyping in 2020, at the end of the third, second, and
 first assessment periods in the R1, R2 and R3 field trials, respectively.
 Where no healthy or dead individuals were available, individuals of
 intermediate damage classes were chosen instead. For the analyses the
 final scoring of ADB intensity (third year of documentation) was used for
 all genotyped samples. DNA of the selected progenies
 was extracted at the Austrian Institute of Technology (Tulln an der Donau,
 Lower Austria) using the Qiagen DNeasy 96 Plant Kit according to the
 manufacturer’s recommendation. DNA quality and quantity were measured
 using PicoGreen™ dsDNA Assay Kits (ThermoFisher). We conducted genotyping
 of a total of 1,107 samples, including 11 technical replicates of the same
 individual (ID = 32806) to control for potential plate-related technical
 biases. Thus, the total number of genotyped individual trees was 1,096.
 Genotyping was done at Thermo Fisher Scientific Inc. using the 4TREE Axiom
 SNP array (Guilbaud et al. 2020; Archambeau et al. 2023), which
 encompasses a set of 13,407 SNPs. Highest priority SNPs include 51
 associated with low susceptibility to emerald ash borer (<em>Agrilus
 planipennis</em>) (Kelly et al. 2020), and 1,633 associated with
 tolerance to ADB (Stocks et al. 2019). Additionally, 200 SNPs are included
 in the array that distinguish <em>F. angustifolia</em> from
 <em>F. excelsior</em> based on analyses that discriminated
 pure individuals of <em>F. excelsior</em> and <em>F.
 angustifolia</em> from each other.
description: # Data from: Limitations of ad hoc genotyping in detecting ash dieback
 tolerance in *Fraxinus excelsior* Dataset DOI:
 [10.5061/dryad.m905qfv9w](https://doi.org/10.5061/dryad.m905qfv9w) ##
 *Fraxinus* *excelsior* 4TREE SNP array data ###### Principle Investigator
 Contact Information ``` Name: Aglaia Szukala Institution: Austrian
 Research Center for Forests (BFW) Email: aglaia.szukala@bfw.gv.at ```
 ###### Alternate Contact Information ``` Name: Heino Konrad Institution:
 Austrian Research Center for Forests (BFW) Email: heino.konrad@bfw.gv.at
 ``` ## Dataset Overview This archive contains the raw data required to
 replicate analyses in Szukala, A., Unger, G.M., Trujillo-Moya C., Geburek,
 T., Kirisits, T., Schueler, S., and Konrad, H. (2025). Limitations of *ad
 hoc* genotyping in detecting ash dieback tolerance in *Fraxinus
 excelsior*. *Tree genetics & Genomes*, [doi:
 10.1007/s11295-025-01725-6](https://doi.org/10.1007/s11295-025-01725-6)
 Filtered data and code can be retrived under this gitHub directory:
 [https://github.com/aglaszuk/AshDiebackGenomics](https://github.com/aglaszuk/AshDiebackGenomics) ### Data Scope Data was collected to investigate the genetic basis of tolerance to ash dieback (ADB) in *Fraxinus* *excelsior* ### Recommended Citation Szukala, A., Unger, G.M., Trujillo-Moya C., Geburek, T., Kirisits, T., Schueler, S., and Konrad, H. (2025). Limitations of *ad hoc* genotyping in detecting ash dieback tolerance in *Fraxinus excelsior*. *Tree genetics & Genomes*, [doi: 10.1007/s11295-025-01725-6](https://doi.org/10.1007/s11295-025-01725-6) ## Description of the data and file structure ### Sample Tracking All samples were processed using a multi-channel pipette to minimize tracking errors. All samples were tracked during assay execution using a LIMS system. ### Files and Folders ##### raw_data.zip The files contained in this zip archive were generated by Thermo Fisher Scientific. ### Files and subfolders contained in the raw_data.zip ##### AxiomGT1.calls.txt Standard Axiom genotypes in flat text file format, with one row per probeset_id and one column per *.CEL. This file contains header columns (e.g.,#[comment]…) followed by the matrix of data. The call translations should be described in the header comments. * The probeset_id (with prefix “AX-”) is a unique identifier for a probe set that is used to interrogate a marker. The column headers contain the path and name of the CEL used in Axiom GT1 analysis. * The [array barcode].CEL part of the column header can be matched to the sample ID in Axiom Analysis Suite. ##### AxiomGT1.confidences.txt Standard Axiom confidences in flat text file format, with one row per probeset_id and one column per *.CEL. The Axiom GT1 confidences matrix file is similar to the Axiom GT1 calls matrix file, except confidence values are given in place of the calls. The confidence score = 1- Pr(x), where X is the called genotype. 0 is the highest confidence and 1 is the lowest confidence. Confidence values are similar to p-values, but are not p-values. ##### AxiomGT1.report.txt Summary values (call rate, het rate) per sample, for Axiom Analysis Suite. ##### AxiomGT1.snp-posteriors.txt Standard Axiom posteriors (cluster boundaries, including means and variance) in flat text file format, with one row per probe set. The posteriors are defined by experimental data, combining information from priors. Output format is described in the header. The information in this file can be used to help with SNP filtering. ##### AxiomGT1.summary.txt Table with one row per ProbeSet+Allele (A or B) and one column per .CEL file. Each entry in the table is the summarized intensity for the ProbeSet+Allele+Sample. The summarized intensities are produced by quantile normalizing the raw intensity of each feature in the *.CEL file to a constant or dynamic sketch. The normalized intensities of replicated features for the same probe are summarized. ##### cluster_summary.txt report file that summarizes the sample and SNP QC results of the genotyping cluster. ##### QC Subfolder containing QC data for Axiom Analysis Suite. * AxiomSS1.calls.txt: calls from signature SNP genotyping; will not be present if signature SNP genotyping was not done. * AxiomSS1.confidences.txt: confidences from signature SNP genotyping; will not be present if signature SNP genotyping was not done. * AxiomSS1.report.txt: report from signature SNP genotyping; will not be present if signature SNP genotyping was not done. * Geno-qcResults.txt: table with dQC information per sample * GenotypingQC.report.txt: table with QC CR information per sample * PlateQCDetails.txt: file with QC details per plate. * Plate_QCSummary.txt: table with QC metrics per plate * Sample_QCFilteredCR.txt: table with filtered CR and het rate information per sample * Sample_QCSummary.txt: file summary information. * sample_QC_cel_files.txt: CEL files passing dQC cutoff, used for QC CR genotyping. ## Access information Other publicly accessible locations of the data: * [https://github.com/aglaszuk/AshDiebackGenomics](https://github.com/aglaszuk/AshDiebackGenomics) Data was derived from the following sources: * Axiom Genotyping Services Projects