DOI: 10.5061/dryad.x3ffbg810
title: Phylogenetic systematics of Juncaceae, alignments associated with phylogenetic trees

description: Juncaceae has needed a taxonomic revision for some time. Specifically, the
 genus Juncus s.l. is known to be paraphyletic, as five small southern
 hemisphere genera have been shown repeatedly to be nested within it. In
 2022, a new classification was proposed, based on phylogenies built from
 one nuclear and three chloroplast regions sequenced across much of
 Juncaceae, meant to resolve the paraphyletic nature of Juncus s.l. It
 created six new genera and was criticized on the basis that the genera
 proposed are not necessarily monophyletic, given the limited nature of the
 phylogenetic analyses available and the fact that the generic
 circumscriptions draw heavily on the morphology-based classifications
 found in the latest monograph of Juncaceae from 2002. We assembled a
 dataset consisting of hundreds of chloroplast and nuclear loci, using the
 Angiosperms 353 target capture probe set, to assess the monophyly of the
 newly proposed genera of Juncaceae. We found that the proposed genera
 mostly represent monophyletic groups, but that the proposed genus
 Juncinella is nested within the proposed genus Boreojuncus. We
 additionally found that Juncinella capitata was placed as sister to either
 Luzula or Oreojuncus and should be recognized as a monotypic genus.
 Finally, Australojuncus cyperoides and Verojuncus chlorocephalus were
 recovered outside of their morphologically assigned genera and require
 further investigation to be placed confidently. Here, we propose taxonomic
 revisions to rectify the issues stated above, but also find that further
 research is necessary, particularly to correctly place the South African
 annual taxa in Juncus s.l.
description: # Phylogenetic systematics of Juncaceae, alignments associated with
 phylogenetic trees Dataset DOI: 10.5061/dryad.x3ffbg810 Associated
 publication: Kenny, R. J., L. Z. Drábková, and D. Potter. 2026.
 Phylogenetic systematics of Juncaceae. American Journal of Botany 113:
 XXXX. #### Description of the data and file structure This dataset
 contains multiple sequence alignments used as input for phylogenetic
 analyses of the rush family Juncaceae (Poales). The study used the
 Angiosperms 353 target capture probe set combined with genome skimming to
 generate nuclear and plastid sequence data across the family, with the
 goal of assessing the monophyly of genera proposed by Brožová et al.
 (2022) and Proćków and Záveská Drábková (2023). Data were generated from
 81 species of Juncaceae plus outgroup taxa in Cyperaceae and Thurniaceae.
 DNA was extracted from herbarium specimens, silica-dried field
 collections, and sequences were downloaded from publicly available
 sequence read archives. Sequence reads were processed using HybPiper
 version 2.1.6 to extract Angiosperms 353 nuclear loci and plastid coding
 sequences. Alignments were generated using MAFFT version 7 (L-INS-i) and
 trimmed using trimAl version 1.4.rev15. Three separate datasets were
 analyzed: (1) Angiosperms 353 nuclear loci, (2) plastid coding sequences
 from off-target reads, and (3) an extended plastid dataset combining our
 plastid data with publicly available matK, rbcL, rpoC1, and trnA-psbA
 sequences downloaded from GenBank. Individual sequences within alignment
 files are labeled with species names, accession numbers or sample codes
 that can be cross referenced using file sample_information.csv. #### Files
 and variables **File: 353_gene_fastas.zip** Description: A zipped folder
 containing 199 individual aligned and trimmed FASTA files, one per locus,
 for the Angiosperms 353 nuclear gene analysis. Each file represents a
 single nuclear locus that passed filtering criteria (present in at least
 50% of taxa, no paralog warnings). Alignments were generated using MAFFT
 version 7 (L-INS-i) and trimmed using trimAl with the -gappyout parameter.
 Sequences within each file are labeled by sample code. These alignments
 were used as input for individual gene tree estimation with IQ-TREE2 and
 subsequently summarized using ASTRAL III to produce the multispecies
 coalescent tree (Figure 1 in the associated publication). The dataset
 includes 70 ingroup species and 37 outgroup taxa, with 14,981 individual
 sequences across all loci, 127,894 alignment sites, 81% variable sites,
 65% parsimony informative sites, and 6% missing data. **File:
 353_iqtree_genes_cat** Description: A single concatenated FASTA file
 containing all 199 Angiosperms 353 nuclear loci, produced by concatenating
 the individual trimmed gene alignments from 353_gene_fastas.zip. This file
 was used as input for the concatenated maximum likelihood analysis in
 IQ-TREE2 version 2.2.6. The matrix was initially partitioned by gene, and
 ModelFinder was used to merge partitions and select substitution models.
 Clade support was assessed using 1000 ultrafast bootstraps. Sequences are
 labeled by sequence code. **File: plast_genes.zip** Description: A zipped
 folder containing 59 individual aligned and trimmed FASTA files, one per
 locus, for the plastid analysis. Each file represents a single plastid
 coding sequence extracted from off-target reads using HybPiper with a
 target file composed of 87 coding sequences from the Juncus effusus
 chloroplast genome (GenBank accession MW366789.1). Loci present in fewer
 than 50% of taxa were excluded. Alignments were generated using MAFFT
 version 7 (L-INS-i) and trimmed using trimAl with the -gappyout parameter.
 The dataset includes 81 ingroup species and 32 outgroup taxa. On average,
 locus alignments contained sequences from 86 individuals and 2,729 sites,
 with 25% parsimony informative sites. Sequences are labeled by taxon name.
 **File: plast_iq-concat** Description: A single concatenated FASTA file
 containing all 59 plastid loci, produced by concatenating the individual
 trimmed gene alignments from plast_genes.zip. This file was used as input
 for the concatenated maximum likelihood analysis in IQ-TREE2 version
 2.2.6. One partition was used and ModelFinder selected the best-fit
 substitution model. Clade support was assessed using 1000 ultrafast
 bootstraps (Figure 2 in the associated publication). The alignment
 contains 174,096 sites, 44% variable sites, 25% parsimony informative
 sites, and 22% missing data. Sequences are labeled by sample code. **File:
 plast_maxtips_aln_trim.zip** Description: A zipped folder containing 59
 individual aligned and trimmed FASTA files for the extended plastid
 analysis. This dataset combines our plastid sequences with publicly
 available matK, rbcL, rpoC1, and trnA-psbA sequences downloaded from
 GenBank using the R package phylotaR version 1.3.0. The dataset includes
 730 samples representing 226 ingroup taxa (51% of species in Juncaceae),
 plus outgroup taxa. Alignments were generated using MAFFT version 7
 (L-INS-i) and trimmed using trimAl with the -automated1 parameter.
 Recovery varied across loci: rbcL (689 sequences), matK (263), rpoC1
 (189), and trnA-psbA (149). The alignment has 87% missing data because
 most publicly downloaded samples are represented only by the four commonly
 sequenced markers. Sample names and GenBank accession numbers for
 downloaded sequences are listed in Appendix S1 of the associated
 publication. Sequences are labeled by sample code. **File:
 plast_maxtips_aln_trim_concat** Description: A single concatenated FASTA
 file containing all 59 loci from the extended plastid analysis, produced
 by concatenating the individual trimmed gene alignments from
 plast_maxtips_aln_trim.zip. This file was used as input for the
 concatenated maximum likelihood analysis in IQ-TREE2 version 2.2.6. One
 partition was used and ModelFinder selected the best-fit substitution
 model. Clade support was assessed using 1000 ultrafast bootstraps (Figure
 3 in the associated publication). The alignment contains 53,441 sites, 66%
 variable sites, 39% parsimony informative sites, and 87% missing data.
 Sequences are labeled by sample code. **File: sample_information.csv**
 Description: A csv file containing the sample codes (tube_lab), species,
 accession number, and herbarium voucher information that can be used for
 cross reference purposes. Variables: * Herbarium: Herbaruim code where the
 voucher specimen is stored, if applicable * Sci_name: Scientific name of
 species sampled * Voucher: Herbarium voucher information * Source:
 Herbarium sample, silica dried tissue, or sequence download * Accession:
 SRA accession number if applicable * GenesWithSeqs_353: Number of
 sequences recovered for the Angionsperms 353 regions (NA- missing data) *
 GenesWithSeqs_plast: Number of sequences recovered for the plastid regions
 * Tube_lab: Sample code used for DNA extraction and analysis scripts ####
 Code/software Documentation for reproducing analyses using this data can
 be found at
 [https://github.com/reedjohnkenny/Juncaceae_353_analysis/tree/master/](https://github.com/reedjohnkenny/Juncaceae_353_analysis/tree/master/) Key software used in generating these data files: * HybPiper version 2.1.6 (Johnson et al., 2016): extraction and assembly of target loci from sequencing reads * MAFFT version 7 (Katoh and Standley, 2013): multiple sequence alignment (L-INS-i method) * trimAl version 1.4.rev15 (Capella-Gutiérrez et al., 2009): alignment trimming (-gappyout for 353 and plastid datasets; -automated1 for extended plastid dataset) * IQ-TREE2 version 2.2.6 (Minh et al., 2020): maximum likelihood tree inference and ModelFinder model selection * ASTRAL III (Zhang et al., 2018): multispecies coalescent species tree estimation from gene trees * TreeShrink version 1.3.7 (Mai and Mirarab, 2018): detection and removal of outlier long branches in gene trees * SPAdes (Bankevich et al., 2012): de novo assembly of loci * Trimmomatic (Bolger et al., 2014): read trimming and adapter removal * phylotaR version 1.3.0 (Bennett et al., 2018): retrieval of orthologous sequences from GenBank #### Access information Other publicly accessible locations of the data: Raw demultiplexed and trimmed sequence reads can be found at GenBank Project PRJNA1196610. Voucher information and accession numbers are provided in Appendix 1 of the associated publication. Additional publicly available sequences used in the extended plastid analysis were downloaded from GenBank and the European Nucleotide Archive; accession numbers are listed in Appendix S1 of the associated publication.