DOI: 10.5061/dryad.0gb5mkm73
title: Data from: Molecular variation and phylogeography within European Isothecium alopecuroides and Pseudisothecium myosuroides (Bryophyta, Lembophyllaceae)

description: Earlier studies on Isothecium s.l. suggested potential interchange of
 genetic material between species that are nowadays known to belong to
 separate genera. In this study, we analyze the molecular variation in
 three molecular markers (nuclear gdp; mitochondrial nad5; and plastid
 trnG) in a wide selection of Isothecium alopecuroides and Pseudisothecium
 myosuroides specimens to infer whether this exchange of genetic material
 is frequent. In addition, we explore the phylogeographic structure of
 these two species in Europe. Our results show repeated genetic interchange
 for the nuclear and mitochondrial markers in both species, with
 intergeneric hybridization as the most probable explanation. The genetic
 lineages found indicate a complex biogeographical history for I.
 alopecuroides, including both postglacial immigration from different
 glacial refugia and developing haplotypes potentially adapted to cold
 climates. As for P. myosuroides, genetic variants could either indicate a
 postglacial immigration history or adaptation to oceanic conditions.
 Further studies including a larger set of more variable molecular markers
 could help to reach final conclusions on the results here presented.
description: We included 94 <em>Isothecium alopecuroides</em> and
 40 <em>Pseudisothecium myosuroides</em> specimens, and 13
 additional members of the Lembophyllaceae lineage to which these two
 species belong: <em>Isotheciastrum subdiversiforme</em> (1),
 <em>Isothecium algarvicum</em> (2), <em>Pseudisothecium
 cristatum</em> (1), <em>P. holtii</em> (3), <em>P.
 interludens</em> (3), <em>P. prolixum</em> (2), and
 <em>P. stoloniferum</em> (1). Most sequences were newly
 generated for the present study, and some came from an earlier study. We
 studied a portion of the nuclear glyceraldehyde 3-phosphate dehydrogenase
 (<em>gdp</em>), the mitochondrial <em>nad</em>5 G1
 intron (<em>nad</em>5), and the plastid
 trnG<sub>UCC</sub> G2 intron
 (<em>trn</em>G). Total DNA was extracted
 using the DNeasy® Plant Mini Kit for DNA isolation from plant tissue
 (Qiagen) according to the manufacturer’s instructions. PCR was performed
 using Ready-To-Go<sup>TM</sup> PCR Beads (Amersham Pharmacia
 Biotech) in a 25 ml reaction volume, also following the manufacturer’s
 protocol. PCR protocol for nad5: 1x 95º 5 min; 1x TC 94º 40 sec, 60‑54º 40
 sec, 72º 1 min 30 sec; 34x 94º 30 sec, 54º 30 sec, 72º 1 min 30 sec; 1x
 72º 10 min, with the primers nad5-F4, nad5-R3 (Cox et al. 2004),
 nad5-Bryo-F160 (ATCCTCAGAGACTGTACGTG), nad5-Bryo-R645
 (GATGCTGCGTCCCTAACAACTA). PCR protocol for gdp: 1x 95º 5 min; 1x TC 94º 40
 sec, 60‑54º 40 sec, 72º 1 min 30 sec; 34x 94º 30 sec, 54º 30 sec, 72º 1
 min 30 sec; 1x 72º 10 min, with the primers: gdp1790F, gdp3050Rr (Wall
 2002), or 1x 95º 5 min; 1x TC 94º 40 sec, 60‑54º 40 sec, 72º 1 min 30 sec;
 40x 94º 30 sec, 54º 30 sec, 72º 1 min 30 sec; 1x 72º 10 min, with the
 primers gdp-Iso-254F (GCTCGCAAAGGTATCATATC), gdp-Iso-784R
 (CACCTTGCCCACAGCCTTG). PCR protocol for trnG: 1x 95º 5 min; 38x 95º 30
 sec, 52º 40 sec, 72º 1 min 30 sec; 1x 72º 8 min, with the primers trnGf,
 trnGr (Werner et al. 2009). Amplified fragments were
 purified using one portion of Exonuclease I (EXO I) (20 u
 ml<sup>-1</sup>) plus four portions of
 FastAP<sup>TM</sup> Thermosensitive Alkaline Phosphatase (1 u
 ml<sup>-1</sup>) (Fermenta LIFE SCIENCE). For 20 ml PCR
 product, 1 ml EXO I + 4 ml FastAP<sup>TM</sup> were used.
 Sequencing was performed using the ABI BigDye Terminator Kit (Applied
 Biosystems) according to the manufacturer’ instructions (BDT ver. 3.1).
 Sequencing products were cleaned using the DyeEx<sup>®</sup>
 96 Kit (QUIAGEN) with the same primers used for the initial PCRs.
 Sequencing products were resolved on an ABI3130xl automated sequencer.
 Double-stranded sequencing was performed in all cases.
 Nucleotide sequence fragments were edited and assembled for each
 DNA region using PhyDE® 0.9971 (http://www.phyde.de/index.html; accessed 22 November 2018). The assembled sequences were aligned manually in PhyDE®. Regions of partially incomplete data at the beginning and end of the sequences were identified and were excluded from subsequent analyses. Gaps were coded using the simple indel coding of Simmons and Ochoterena (2000) in SeqState (Müller 2005).
description: # Data from: Molecular variation and phylogeography within European
 Isothecium alopecuroides and Pseudisothecium myosuroides (Bryophyta,
 Lembophyllaceae) ##
 Description of the data and file structure The data includes the following
 four files: Isothecium alopecuroides_gpd.txt; Isothecium
 alopecuroides_nad5.txt; Isothecium alopecuroides_trnG.txt; Isothecium
 alopecuroides_TOTAL.txt The files include sequence alignments in FASTA
 format Usage notes: The positions in the files are as follows: Isothecium
 alopecuroides_gpd.txt: base positions: 1-510 indel coding positions:
 512-515 Isothecium alopecuroides_nad5.txt: base positions: 1-954 indel
 coding positions: 956-963 Isothecium alopecuroides_trnG.txt: base
 positions: 1-557 indel coding positions: 559-567 Isothecium
 alopecuroides_TOTAL.txt [only samples with all three markers are included
 in this alignment]: base positions: 1-510 (gpd); 517-1466 (nad5);
 1476-2032 (trnG) indel coding positions: 512-515 (gpd); 1468-1474 (nad5);
 2034-2042 (trnG)