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<record><TEI><teiHeader><fileDesc><titleStmt><title xml:lang="en">Development of 23 polymorphic microsatellite loci in invasive silver wattle, <italic>Acacia dealbata</italic> (Fabaceae)<xref ref-type="author-notes" rid="fn1"><sup>1</sup></xref></title><author><name sortKey="Guillemaud, Thomas" sort="Guillemaud, Thomas" uniqKey="Guillemaud T" first="Thomas" last="Guillemaud">Thomas Guillemaud</name><affiliation><nlm:aff id="aff2">Institut national de la recherche agronomique (INRA), Université Nice Sophia Antipolis, Centre national de la recherche scientifique (CNRS), UMR 1355-7254 Institut Sophia Agrobiotech, 06900 Sophia Antipolis, France</nlm:aff></affiliation></author><author><name sortKey="Broadhurst, Linda" sort="Broadhurst, Linda" uniqKey="Broadhurst L" first="Linda" last="Broadhurst">Linda Broadhurst</name><affiliation><nlm:aff id="aff3">Centre for Australian National Biodiversity Research, National Collections Australia, Commonwealth Scientific and Industrial Research Organisation (CSIRO), GPO Box 1600, Canberra, Australian Capital Territory 2601, Australia</nlm:aff></affiliation></author><author><name sortKey="Legoff, Isabelle" sort="Legoff, Isabelle" uniqKey="Legoff I" first="Isabelle" last="Legoff">Isabelle Legoff</name><affiliation><nlm:aff id="aff2">Institut national de la recherche agronomique (INRA), Université Nice Sophia Antipolis, Centre national de la recherche scientifique (CNRS), UMR 1355-7254 Institut Sophia Agrobiotech, 06900 Sophia Antipolis, France</nlm:aff></affiliation></author><author><name sortKey="Henery, Martin" sort="Henery, Martin" uniqKey="Henery M" first="Martin" last="Henery">Martin Henery</name><affiliation><nlm:aff id="aff3">Centre for Australian National Biodiversity Research, National Collections Australia, Commonwealth Scientific and Industrial Research Organisation (CSIRO), GPO Box 1600, Canberra, Australian Capital Territory 2601, Australia</nlm:aff></affiliation></author><author><name sortKey="Blin, Aurelie" sort="Blin, Aurelie" uniqKey="Blin A" first="Aurélie" last="Blin">Aurélie Blin</name><affiliation><nlm:aff id="aff2">Institut national de la recherche agronomique (INRA), Université Nice Sophia Antipolis, Centre national de la recherche scientifique (CNRS), UMR 1355-7254 Institut Sophia Agrobiotech, 06900 Sophia Antipolis, France</nlm:aff></affiliation></author><author><name sortKey="Ducatillion, Catherine" sort="Ducatillion, Catherine" uniqKey="Ducatillion C" first="Catherine" last="Ducatillion">Catherine Ducatillion</name><affiliation><nlm:aff id="aff4">Unité Expérimentale Villa Thuret, INRA–Villa Thuret, 90 Chemin Raymond, 06160 Antibes–Juan-les-Pins, France</nlm:aff></affiliation></author><author><name sortKey="Ferrando, Nathalie" sort="Ferrando, Nathalie" uniqKey="Ferrando N" first="Nathalie" last="Ferrando">Nathalie Ferrando</name><affiliation><nlm:aff id="aff2">Institut national de la recherche agronomique (INRA), Université Nice Sophia Antipolis, Centre national de la recherche scientifique (CNRS), UMR 1355-7254 Institut Sophia Agrobiotech, 06900 Sophia Antipolis, France</nlm:aff></affiliation></author><author><name sortKey="Malausa, Thibaut" sort="Malausa, Thibaut" uniqKey="Malausa T" first="Thibaut" last="Malausa">Thibaut Malausa</name><affiliation><nlm:aff id="aff2">Institut national de la recherche agronomique (INRA), Université Nice Sophia Antipolis, Centre national de la recherche scientifique (CNRS), UMR 1355-7254 Institut Sophia Agrobiotech, 06900 Sophia Antipolis, France</nlm:aff></affiliation></author></titleStmt><publicationStmt><idno type="wicri:source">PMC</idno><idno type="pmid">25995979</idno><idno type="pmc">4435469</idno><idno type="url">http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4435469</idno><idno type="RBID">PMC:4435469</idno><idno type="doi">10.3732/apps.1500018</idno><date when="2015">2015</date><idno type="wicri:Area/Pmc/Corpus">000248</idno><idno type="wicri:explorRef" wicri:stream="Pmc" wicri:step="Corpus" wicri:corpus="PMC">000248</idno></publicationStmt><sourceDesc><biblStruct><analytic><title xml:lang="en" level="a" type="main">Development of 23 polymorphic microsatellite loci in invasive silver wattle, <italic>Acacia dealbata</italic> (Fabaceae)<xref ref-type="author-notes" rid="fn1"><sup>1</sup></xref></title><author><name sortKey="Guillemaud, Thomas" sort="Guillemaud, Thomas" uniqKey="Guillemaud T" first="Thomas" last="Guillemaud">Thomas Guillemaud</name><affiliation><nlm:aff id="aff2">Institut national de la recherche agronomique (INRA), Université Nice Sophia Antipolis, Centre national de la recherche scientifique (CNRS), UMR 1355-7254 Institut Sophia Agrobiotech, 06900 Sophia Antipolis, France</nlm:aff></affiliation></author><author><name sortKey="Broadhurst, Linda" sort="Broadhurst, Linda" uniqKey="Broadhurst L" first="Linda" last="Broadhurst">Linda Broadhurst</name><affiliation><nlm:aff id="aff3">Centre for Australian National Biodiversity Research, National Collections Australia, Commonwealth Scientific and Industrial Research Organisation (CSIRO), GPO Box 1600, Canberra, Australian Capital Territory 2601, Australia</nlm:aff></affiliation></author><author><name sortKey="Legoff, Isabelle" sort="Legoff, Isabelle" uniqKey="Legoff I" first="Isabelle" last="Legoff">Isabelle Legoff</name><affiliation><nlm:aff id="aff2">Institut national de la recherche agronomique (INRA), Université Nice Sophia Antipolis, Centre national de la recherche scientifique (CNRS), UMR 1355-7254 Institut Sophia Agrobiotech, 06900 Sophia Antipolis, France</nlm:aff></affiliation></author><author><name sortKey="Henery, Martin" sort="Henery, Martin" uniqKey="Henery M" first="Martin" last="Henery">Martin Henery</name><affiliation><nlm:aff id="aff3">Centre for Australian National Biodiversity Research, National Collections Australia, Commonwealth Scientific and Industrial Research Organisation (CSIRO), GPO Box 1600, Canberra, Australian Capital Territory 2601, Australia</nlm:aff></affiliation></author><author><name sortKey="Blin, Aurelie" sort="Blin, Aurelie" uniqKey="Blin A" first="Aurélie" last="Blin">Aurélie Blin</name><affiliation><nlm:aff id="aff2">Institut national de la recherche agronomique (INRA), Université Nice Sophia Antipolis, Centre national de la recherche scientifique (CNRS), UMR 1355-7254 Institut Sophia Agrobiotech, 06900 Sophia Antipolis, France</nlm:aff></affiliation></author><author><name sortKey="Ducatillion, Catherine" sort="Ducatillion, Catherine" uniqKey="Ducatillion C" first="Catherine" last="Ducatillion">Catherine Ducatillion</name><affiliation><nlm:aff id="aff4">Unité Expérimentale Villa Thuret, INRA–Villa Thuret, 90 Chemin Raymond, 06160 Antibes–Juan-les-Pins, France</nlm:aff></affiliation></author><author><name sortKey="Ferrando, Nathalie" sort="Ferrando, Nathalie" uniqKey="Ferrando N" first="Nathalie" last="Ferrando">Nathalie Ferrando</name><affiliation><nlm:aff id="aff2">Institut national de la recherche agronomique (INRA), Université Nice Sophia Antipolis, Centre national de la recherche scientifique (CNRS), UMR 1355-7254 Institut Sophia Agrobiotech, 06900 Sophia Antipolis, France</nlm:aff></affiliation></author><author><name sortKey="Malausa, Thibaut" sort="Malausa, Thibaut" uniqKey="Malausa T" first="Thibaut" last="Malausa">Thibaut Malausa</name><affiliation><nlm:aff id="aff2">Institut national de la recherche agronomique (INRA), Université Nice Sophia Antipolis, Centre national de la recherche scientifique (CNRS), UMR 1355-7254 Institut Sophia Agrobiotech, 06900 Sophia Antipolis, France</nlm:aff></affiliation></author></analytic><series><title level="j">Applications in Plant Sciences</title><idno type="eISSN">2168-0450</idno><imprint><date when="2015">2015</date></imprint></series></biblStruct></sourceDesc></fileDesc><profileDesc><textClass></textClass></profileDesc></teiHeader><front><div type="abstract" xml:lang="en"><sec><title>Premise of the study:</title><p>Microsatellite markers were developed for silver wattle, <italic>Acacia dealbata</italic> (Fabaceae), which is both an ornamental and an invasive weed species. It is native to southeastern Australia and invasive in Europe, Africa, Asia, and the Americas.</p></sec><sec><title>Methods and Results:</title><p>The pyrosequencing of a microsatellite-enriched genomic DNA library of <italic>A. dealbata</italic> produced 33,290 sequences and allowed the isolation of 201 loci with a minimum of seven repeats of microsatellite motifs. Amplification tests led to the setup of two multiplex PCR mixes allowing the amplification of 21 loci. The polymorphism of these markers was evaluated on a sample of 32 individuals collected in southeastern Australia. The number of alleles and the expected heterozygosity varied between two and 11, and between 0.11 and 0.88, respectively.</p></sec><sec><title>Conclusions:</title><p>The level of polymorphism of this set of 23 microsatellites is large enough to provide valuable information on the genetic structure and the invasion history of <italic>A. dealbata</italic>.</p></sec></div></front><back><div1 type="bibliography"><listBibl><biblStruct><analytic><author><name sortKey="Aldrich, P R" uniqKey="Aldrich P">P. R. Aldrich</name></author><author><name sortKey="Hamrick, J L" uniqKey="Hamrick J">J. L. Hamrick</name></author><author><name sortKey="Chavarriaga, P" uniqKey="Chavarriaga P">P. Chavarriaga</name></author><author><name sortKey="Kochert, G" uniqKey="Kochert G">G. Kochert</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Benjamini, Y" uniqKey="Benjamini Y">Y. Benjamini</name></author><author><name sortKey="Hochberg, M" uniqKey="Hochberg M">M. Hochberg</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Broadhurst, L M" uniqKey="Broadhurst L">L. M. Broadhurst</name></author><author><name sortKey="Young, A G" uniqKey="Young A">A. G. Young</name></author></analytic></biblStruct><biblStruct></biblStruct><biblStruct><analytic><author><name sortKey="Estoup, A" uniqKey="Estoup A">A. Estoup</name></author><author><name sortKey="Guillemaud, T" uniqKey="Guillemaud T">T. Guillemaud</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Guichoux, E" uniqKey="Guichoux E">E. Guichoux</name></author><author><name sortKey="Lagache, L" uniqKey="Lagache L">L. Lagache</name></author><author><name sortKey="Wagner, S" uniqKey="Wagner S">S. Wagner</name></author><author><name sortKey="Chaumeil, P" uniqKey="Chaumeil P">P. Chaumeil</name></author><author><name sortKey="Leger, P" uniqKey="Leger P">P. Léger</name></author><author><name sortKey="Lepais, O" uniqKey="Lepais O">O. Lepais</name></author><author><name sortKey="Lepoittevin, C" uniqKey="Lepoittevin C">C. Lepoittevin</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Kull, C A" uniqKey="Kull C">C. A. Kull</name></author><author><name sortKey="Shackleton, C M" uniqKey="Shackleton C">C. M. Shackleton</name></author><author><name sortKey="Cunningham, P J" uniqKey="Cunningham P">P. J. Cunningham</name></author><author><name sortKey="Ducatillion, C" uniqKey="Ducatillion C">C. Ducatillion</name></author><author><name sortKey="Dufour Dror, J M" uniqKey="Dufour Dror J">J. M. Dufour-Dror</name></author><author><name sortKey="Esler, K J" uniqKey="Esler K">K. J. Esler</name></author><author><name sortKey="Friday, J B" uniqKey="Friday J">J. B. Friday</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Lorenzo, P" uniqKey="Lorenzo P">P. Lorenzo</name></author><author><name sortKey="Gonzalez, L" uniqKey="Gonzalez L">L. González</name></author><author><name sortKey="Reigosa, M J" uniqKey="Reigosa M">M. J. Reigosa</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Malausa, T" uniqKey="Malausa T">T. Malausa</name></author><author><name sortKey="Gilles, A" uniqKey="Gilles A">A. Gilles</name></author><author><name sortKey="Meglecz, E" uniqKey="Meglecz E">E. Meglecz</name></author><author><name sortKey="Blanquart, H" uniqKey="Blanquart H">H. Blanquart</name></author><author><name sortKey="Duthoy, S" uniqKey="Duthoy S">S. Duthoy</name></author><author><name sortKey="Costedoat, C" uniqKey="Costedoat C">C. Costedoat</name></author><author><name sortKey="Dubut, V" uniqKey="Dubut V">V. Dubut</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Meglecz, E" uniqKey="Meglecz E">E. Meglecz</name></author><author><name sortKey="Costedoat, C" uniqKey="Costedoat C">C. Costedoat</name></author><author><name sortKey="Dubut, V" uniqKey="Dubut V">V. Dubut</name></author><author><name sortKey="Gilles, A" uniqKey="Gilles A">A. Gilles</name></author><author><name sortKey="Malausa, T" uniqKey="Malausa T">T. Malausa</name></author><author><name sortKey="Pech, N" uniqKey="Pech N">N. Pech</name></author><author><name sortKey="Martin, J F" uniqKey="Martin J">J.-F. Martin</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Nei, M" uniqKey="Nei M">M. Nei</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Rousset, F" uniqKey="Rousset F">F. Rousset</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Sokal, R R" uniqKey="Sokal R">R. R. Sokal</name></author><author><name sortKey="Rolf, F J" uniqKey="Rolf F">F. J. Rolf</name></author></analytic></biblStruct></listBibl></div1></back></TEI><pmc article-type="research-article"><pmc-dir>properties open_access</pmc-dir>
<front><journal-meta><journal-id journal-id-type="nlm-ta">Appl Plant Sci</journal-id><journal-id journal-id-type="iso-abbrev">Appl Plant Sci</journal-id><journal-id journal-id-type="publisher-id">apps</journal-id><journal-title-group><journal-title>Applications in Plant Sciences</journal-title></journal-title-group><issn pub-type="epub">2168-0450</issn><publisher><publisher-name>Botanical Society of America</publisher-name></publisher></journal-meta><article-meta><article-id pub-id-type="pmid">25995979</article-id><article-id pub-id-type="pmc">4435469</article-id><article-id pub-id-type="publisher-id">apps1500018</article-id><article-id pub-id-type="doi">10.3732/apps.1500018</article-id><article-categories><subj-group subj-group-type="heading"><subject>Primer Note</subject></subj-group></article-categories><title-group><article-title>Development of 23 polymorphic microsatellite loci in invasive silver wattle, <italic>Acacia dealbata</italic> (Fabaceae)<xref ref-type="author-notes" rid="fn1"><sup>1</sup></xref></article-title></title-group><contrib-group><contrib contrib-type="author"><name><surname>Guillemaud</surname><given-names>Thomas</given-names></name><xref ref-type="aff" rid="aff2"><sup>2</sup></xref><xref ref-type="corresp" rid="cor1"><sup>5</sup></xref></contrib><contrib contrib-type="author"><name><surname>Broadhurst</surname><given-names>Linda</given-names></name><xref ref-type="aff" rid="aff3"><sup>3</sup></xref></contrib><contrib contrib-type="author"><name><surname>Legoff</surname><given-names>Isabelle</given-names></name><xref ref-type="aff" rid="aff2"><sup>2</sup></xref></contrib><contrib contrib-type="author"><name><surname>Henery</surname><given-names>Martin</given-names></name><xref ref-type="aff" rid="aff3"><sup>3</sup></xref></contrib><contrib contrib-type="author"><name><surname>Blin</surname><given-names>Aurélie</given-names></name><xref ref-type="aff" rid="aff2"><sup>2</sup></xref></contrib><contrib contrib-type="author"><name><surname>Ducatillion</surname><given-names>Catherine</given-names></name><xref ref-type="aff" rid="aff4"><sup>4</sup></xref></contrib><contrib contrib-type="author"><name><surname>Ferrando</surname><given-names>Nathalie</given-names></name><xref ref-type="aff" rid="aff2"><sup>2</sup></xref></contrib><contrib contrib-type="author"><name><surname>Malausa</surname><given-names>Thibaut</given-names></name><xref ref-type="aff" rid="aff2"><sup>2</sup></xref></contrib><aff id="aff2"><label>2</label>Institut national de la recherche agronomique (INRA), Université Nice Sophia Antipolis, Centre national de la recherche scientifique (CNRS), UMR 1355-7254 Institut Sophia Agrobiotech, 06900 Sophia Antipolis, France</aff><aff id="aff3"><label>3</label>Centre for Australian National Biodiversity Research, National Collections Australia, Commonwealth Scientific and Industrial Research Organisation (CSIRO), GPO Box 1600, Canberra, Australian Capital Territory 2601, Australia</aff><aff id="aff4"><label>4</label>Unité Expérimentale Villa Thuret, INRA–Villa Thuret, 90 Chemin Raymond, 06160 Antibes–Juan-les-Pins, France</aff></contrib-group><author-notes><fn id="fn1"><label>1</label><p>The authors thank the Institut national de la recherche agronomique (INRA) for providing the AIP BioRessources ‘EcoMicro’ grant to T.M.</p></fn><corresp id="cor1"><label>5</label>Author for correspondence: <email>guillem@sophia.inra.fr</email></corresp></author-notes><pub-date pub-type="collection"><month>5</month><year>2015</year></pub-date><pub-date pub-type="epub"><day>11</day><month>5</month><year>2015</year></pub-date><volume>3</volume><issue>5</issue><elocation-id>apps.1500018</elocation-id><history><date date-type="received"><day>20</day><month>2</month><year>2015</year></date><date date-type="accepted"><day>25</day><month>3</month><year>2015</year></date></history><permissions><copyright-statement>© 2015 Guillemaud et al. Published by the Botanical Society of America</copyright-statement><copyright-year>2015</copyright-year><license license-type="creative-commons" xlink:href="http://creativecommons.org/licenses/by-nc/4.0/"><license-p>This work is licensed under a Creative Commons Attribution License (CC-BY-NC-SA).</license-p></license></permissions><abstract><sec><title>Premise of the study:</title><p>Microsatellite markers were developed for silver wattle, <italic>Acacia dealbata</italic> (Fabaceae), which is both an ornamental and an invasive weed species. It is native to southeastern Australia and invasive in Europe, Africa, Asia, and the Americas.</p></sec><sec><title>Methods and Results:</title><p>The pyrosequencing of a microsatellite-enriched genomic DNA library of <italic>A. dealbata</italic> produced 33,290 sequences and allowed the isolation of 201 loci with a minimum of seven repeats of microsatellite motifs. Amplification tests led to the setup of two multiplex PCR mixes allowing the amplification of 21 loci. The polymorphism of these markers was evaluated on a sample of 32 individuals collected in southeastern Australia. The number of alleles and the expected heterozygosity varied between two and 11, and between 0.11 and 0.88, respectively.</p></sec><sec><title>Conclusions:</title><p>The level of polymorphism of this set of 23 microsatellites is large enough to provide valuable information on the genetic structure and the invasion history of <italic>A. dealbata</italic>.</p></sec></abstract><kwd-group><kwd><italic>Acacia dealbata</italic></kwd><kwd>Fabaceae</kwd><kwd>invasion</kwd><kwd>microsatellites</kwd><kwd>pyrosequencing</kwd><kwd>silver wattle</kwd></kwd-group></article-meta></front><body><p>Silver wattle, <italic>Acacia dealbata</italic> Link (Fabaceae), is a tree species native to southeastern Australia, where it is widespread and common. It has been introduced since the 18th century in various parts of the world for ornamental or forestry purposes (<xref rid="bib7" ref-type="bibr">Kull et al., 2011</xref>). It is now considered an invasive species in southern Europe, South Africa, India, Madagascar, New Zealand, Chile, and California (<xref rid="bib7" ref-type="bibr">Kull et al., 2011</xref>). In those regions, <italic>A. dealbata</italic> invades woodland and disturbed environments where it outcompetes native species. <italic>Acacia dealbata</italic> has a long and complex history of worldwide introductions from Australia, at least since its introduction in the early 18th century in Europe (<xref rid="bib7" ref-type="bibr">Kull et al., 2011</xref>). Silver wattle is the focus of research both because it is a model for the impact of landscape fragmentation in its native area (<xref rid="bib3" ref-type="bibr">Broadhurst and Young, 2006</xref>) and because it is a famous invader (<xref rid="bib8" ref-type="bibr">Lorenzo et al., 2010</xref>), classified on the Delivering Alien Invasive Species Inventories for Europe (DAISIE) list as one of the 100 European invaders with the most significant ecological impacts (<xref rid="bib4" ref-type="bibr">DAISIE European Invasive Alien Species Gateway, 2006</xref>). In both cases, the use of highly polymorphic genetic markers could help answer questions related to the spatial genetic structure of <italic>A. dealbata</italic> populations.</p><p>Variable genetic markers such as microsatellite markers are a common tool used to infer invasion routes of invading species and specifically to identify the sources of invasive populations (<xref rid="bib5" ref-type="bibr">Estoup and Guillemaud, 2010</xref>). They are also commonly used in conservation biology studies to decipher the impact of environmental disturbance on the genetic structure of tree species (e.g., <xref rid="bib1" ref-type="bibr">Aldrich et al., 1998</xref>). Recently, next-generation sequencing technologies have been used to obtain very large numbers of shotgun sequences of genomic DNA from which microsatellites could be isolated (<xref rid="bib6" ref-type="bibr">Guichoux et al., 2011</xref>). This method requires a large effort of sequencing to obtain a sufficient number of sequences containing microsatellites. Microsatellite enrichment of the genomic DNA prior to sequencing allows a drastic cost reduction (<xref rid="bib9" ref-type="bibr">Malausa et al., 2011</xref>). Here we present the isolation and properties of 23 polymorphic microsatellite loci of <italic>A. dealbata</italic> using 454 GS FLX Titanium (Roche Applied Science, Penzberg, Germany) pyrosequencing of a microsatellite-enriched genomic DNA library.</p><sec sec-type="methods"><title>METHODS AND RESULTS</title><p>Total genomic DNA was extracted from leaf material of a plant collected in southeastern Australia (34°30′51.498″S, 148°49′53.3892″E) using the DNeasy Plant Kit (QIAGEN, Hilden, Germany). We followed the procedure of <xref rid="bib9" ref-type="bibr">Malausa et al. (2011)</xref> for enrichment and sequencing. Genomic DNA was submitted to sonication, ligation to standard adapters, and purification on a NucleoFast PCR plate (Macherey-Nagel, Düren, Germany). DNA was denatured and then hybridized for 20 min at 56°C to eight biotin-labeled oligonucleotides, the sequences of which are microsatellite motifs [(AG)<sub>10</sub>, (AC)<sub>10</sub>, (AAC)<sub>8</sub>, (AGG)<sub>8</sub>, (ACG)<sub>8</sub>, (AAG)<sub>8</sub>, (ACAT)<sub>6</sub>, and (ATCT)<sub>6</sub>].</p><p>We then used primers corresponding to the adapters (5′-GTTTAAGGCCTAGCTAGCAGAATC-3′ and 5′-GATTCTGCTAGCTAGGCCTT-3′) to amplify the microsatellite-enriched DNA. Preparation of the enriched library for emulsion PCR, sequencing, and analytical processing using the 454 GS FLX Titanium followed the manufacturer’s protocols. We then sorted the obtained sequences, removed the sequence of the adapters, and selected sequences with desirable properties using QDD version 1 (<xref rid="bib10" ref-type="bibr">Meglecz et al., 2010</xref>).</p><p>Among the 33,290 sequences obtained, a total of 201 sequences longer than 80 bp, containing microsatellite motifs with at least seven uninterrupted repeats and flanking regions free of tandem repetitions, were used to design PCR primer pairs. We tested the 201 primer pairs for amplification on seven individuals, and then tested the successful primer pairs for fluorescent PCR on 32 individuals collected in southeastern Australia, between latitudes 34°37′43.1″S and 34°29′08.9″S and between longitudes 148°50′28.3″E and 148°45′23.9″E (see <xref ref-type="app" rid="app1">Appendix 1</xref> for individual coordinates). A representative voucher specimen (CANB77329, deposited in the Australian National Herbarium [CANB], Canberra, Australia) was previously collected in the same region as the samples studied here.</p><p>These 32 individuals were used to test the amplification of these loci according to the following procedure: PCR amplifications were performed in a 10-μL volume containing 1× QIAGEN Multiplex Master Mix, 0.2 μM of each primer, and ca. 20 ng of genomic DNA extracted from individual leaves using the DNeasy Plant Kit (QIAGEN). The amplification reactions were performed in an Eppendorf (Hamburg, Germany) Mastercycler thermocycler and included a 15-min denaturation step at 95°C; followed by 35 cycles of 30 s at 94°C, 1.5 min at 56°C, and 1 min at 72°C; followed by a final extension step at 60°C for 30 min. Fifty primer pairs gave positive PCR amplification of the predicted size for the seven individuals. They were then used in fluorescent PCR to amplify the 32 individuals as described above but with forward primers labeled with a fluorescent dye on the 5′ end, and with 25 cycles of PCR instead of 35. Twenty-three polymorphic microsatellites could be unambiguously scored using an ABI 3700 sequencer (Applied Biosystems, Waltham, Massachusetts, USA) with the GeneScan 500 LIZ Size Standard (Applied Biosystems) and GeneMarker software (version 1.75; SoftGenetics LLC, State College, Pennsylvania, USA) (<xref ref-type="table" rid="tbl1">Table 1</xref>). The sequences containing the 23 microsatellites led to no significant similarity when blasted against National Center for Biotechnology Information (NCBI) nonredundant (NR) and expressed sequence tags (dbEST) databases.</p><table-wrap id="tbl1" position="float"><label>Table 1.</label><caption><p>Characteristics and variability of 23 microsatellite loci of <italic>Acacia dealbata</italic> based on 32 individuals sampled in southeastern Australia.</p></caption><table frame="hsides" rules="groups"><thead><tr><td rowspan="1" colspan="1">Locus</td><td align="center" rowspan="1" colspan="1">Primer sequences (5′–3′)</td><td align="center" rowspan="1" colspan="1">Repeat motif<xref ref-type="table-fn" rid="tblfn1"><sup>a</sup></xref></td><td align="center" rowspan="1" colspan="1">Allele size range (bp)<xref ref-type="table-fn" rid="tblfn2"><sup>b</sup></xref></td><td align="center" rowspan="1" colspan="1">5′ end-labeled dye</td><td align="center" rowspan="1" colspan="1">Multiplex marker set<xref ref-type="table-fn" rid="tblfn3"><sup>c</sup></xref></td><td align="center" rowspan="1" colspan="1">GenBank accession no.</td></tr></thead><tbody><tr><td rowspan="1" colspan="1">Ad-33</td><td rowspan="1" colspan="1">F: GAGAAGAGAAAGGGGATGGG</td><td rowspan="1" colspan="1">(GA)<sub>8</sub></td><td align="center" rowspan="1" colspan="1">159–169</td><td rowspan="1" colspan="1">FAM</td><td align="center" rowspan="1" colspan="1">2</td><td rowspan="1" colspan="1">KP702738</td></tr><tr><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1">R: CCATCATTATTGTTTGTCCCG</td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td></tr><tr><td rowspan="1" colspan="1">Ad-41</td><td rowspan="1" colspan="1">F: TGAGATTATAAGCTTCTGGTAAAAAG</td><td rowspan="1" colspan="1">(GA)<sub>8</sub></td><td align="center" rowspan="1" colspan="1">102–108</td><td rowspan="1" colspan="1">PET</td><td align="center" rowspan="1" colspan="1">2</td><td rowspan="1" colspan="1">KP702739</td></tr><tr><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1">R: AAAAGCCACGCCCTTTAGTT</td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td></tr><tr><td rowspan="1" colspan="1">Ad-45</td><td rowspan="1" colspan="1">F: TCTAATAAAAGAACAGCAACGCA</td><td rowspan="1" colspan="1">(AC)<sub>8</sub></td><td align="center" rowspan="1" colspan="1">74–87</td><td rowspan="1" colspan="1">NED</td><td align="center" rowspan="1" colspan="1">2</td><td rowspan="1" colspan="1">KP702740</td></tr><tr><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1">R: AATTGGTTTGCATGTGATGG</td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td></tr><tr><td rowspan="1" colspan="1">Ad-47</td><td rowspan="1" colspan="1">F: CCCACATAAAGAAGAAACTAGTGAA</td><td rowspan="1" colspan="1">(AG)<sub>8</sub></td><td align="center" rowspan="1" colspan="1">89–95</td><td rowspan="1" colspan="1">PET</td><td align="center" rowspan="1" colspan="1">1</td><td rowspan="1" colspan="1">KP702741</td></tr><tr><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1">R: CGTGGGAAAGAAACATGGAT</td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td></tr><tr><td rowspan="1" colspan="1">Ad-48</td><td rowspan="1" colspan="1">F: TAGCTTTCCTCGGATCCCTT</td><td rowspan="1" colspan="1">(AC)<sub>9</sub></td><td align="center" rowspan="1" colspan="1">196–214</td><td rowspan="1" colspan="1">VIC</td><td align="center" rowspan="1" colspan="1">2</td><td rowspan="1" colspan="1">KP702742</td></tr><tr><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1">R: ACACATCAGAACACTGGGCA</td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td></tr><tr><td rowspan="1" colspan="1">Ad-49</td><td rowspan="1" colspan="1">F: CTTCACAAGCAAAAGGGCAC</td><td rowspan="1" colspan="1">(AG)<sub>8</sub></td><td align="center" rowspan="1" colspan="1">168–184</td><td rowspan="1" colspan="1">FAM</td><td align="center" rowspan="1" colspan="1">1</td><td rowspan="1" colspan="1">KP702743</td></tr><tr><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1">R: CAGCCATGTTCATGGTTAAAA</td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td></tr><tr><td rowspan="1" colspan="1">Ad-54</td><td rowspan="1" colspan="1">F: TGCAGCCAGAAAGTGAAATG</td><td rowspan="1" colspan="1">(TTC)<sub>8</sub></td><td align="center" rowspan="1" colspan="1">131–143</td><td rowspan="1" colspan="1">NED</td><td align="center" rowspan="1" colspan="1">1</td><td rowspan="1" colspan="1">KP702744</td></tr><tr><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1">R: GAGAAACTAAAGATGAAGACGGAGA</td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td></tr><tr><td rowspan="1" colspan="1">Ad-59</td><td rowspan="1" colspan="1">F: CCCAGGTCAAACGCAGTAAT</td><td rowspan="1" colspan="1">(TC)<sub>10</sub></td><td align="center" rowspan="1" colspan="1">167–175</td><td rowspan="1" colspan="1">NED</td><td align="center" rowspan="1" colspan="1">1</td><td rowspan="1" colspan="1">KP702745</td></tr><tr><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1">R: CTGTCAAACACCAGAAAGGTAGA</td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td></tr><tr><td rowspan="1" colspan="1">Ad-63</td><td rowspan="1" colspan="1">F: CCTTTTCCCCACATTCTCCT</td><td rowspan="1" colspan="1">(TC)<sub>9</sub></td><td align="center" rowspan="1" colspan="1">120–133</td><td rowspan="1" colspan="1">VIC</td><td align="center" rowspan="1" colspan="1">NA</td><td rowspan="1" colspan="1">KP702746</td></tr><tr><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1">R: CGTCCCGCAATTTCTTATGT</td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td></tr><tr><td rowspan="1" colspan="1">Ad-66</td><td rowspan="1" colspan="1">F: TGTGAACGAAGCACAAGACC</td><td rowspan="1" colspan="1">(CA)<sub>8</sub></td><td align="center" rowspan="1" colspan="1">161–165</td><td rowspan="1" colspan="1">VIC</td><td align="center" rowspan="1" colspan="1">1</td><td rowspan="1" colspan="1">KP702747</td></tr><tr><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1">R: TGTGAGCAACGACATTGACA</td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td></tr><tr><td rowspan="1" colspan="1">Ad-70</td><td rowspan="1" colspan="1">F: CTCCCATCTCAGATCTTGGC</td><td rowspan="1" colspan="1">(GT)<sub>8</sub></td><td align="center" rowspan="1" colspan="1">131–132</td><td rowspan="1" colspan="1">PET</td><td align="center" rowspan="1" colspan="1">1</td><td rowspan="1" colspan="1">KP702748</td></tr><tr><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1">R: CAAGCAATGTTCCACTCCCT</td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td></tr><tr><td rowspan="1" colspan="1">Ad-86</td><td rowspan="1" colspan="1">F: TTTGAAAGCATGCTCCATTTT</td><td rowspan="1" colspan="1">(AC)<sub>8</sub></td><td align="center" rowspan="1" colspan="1">106–122</td><td rowspan="1" colspan="1">NED</td><td align="center" rowspan="1" colspan="1">1</td><td rowspan="1" colspan="1">KP702749</td></tr><tr><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1">R: GCCTTCCCTCCGTTAGAATC</td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td></tr><tr><td rowspan="1" colspan="1">Ad-89</td><td rowspan="1" colspan="1">F: TCATTCTGCACGTCATTTCC</td><td rowspan="1" colspan="1">(CT)<sub>11</sub></td><td align="center" rowspan="1" colspan="1">80–96</td><td rowspan="1" colspan="1">NED</td><td align="center" rowspan="1" colspan="1">1</td><td rowspan="1" colspan="1">KP702750</td></tr><tr><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1">R: CACAGGGCTTTGTATTTGTATCTTT</td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td></tr><tr><td rowspan="1" colspan="1">Ad-97</td><td rowspan="1" colspan="1">F: GCAGTTAAAACCTCGAAGGC</td><td rowspan="1" colspan="1">(GT)<sub>8</sub></td><td align="center" rowspan="1" colspan="1">92–96</td><td rowspan="1" colspan="1">FAM</td><td align="center" rowspan="1" colspan="1">1</td><td rowspan="1" colspan="1">KP702751</td></tr><tr><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1">R: CTGAACTTGATCACTTCATGGT</td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td></tr><tr><td rowspan="1" colspan="1">Ad-116</td><td rowspan="1" colspan="1">F: TTGGGTTGAATTCCTTTCTCA</td><td rowspan="1" colspan="1">(AC)<sub>9</sub></td><td align="center" rowspan="1" colspan="1">127–135</td><td rowspan="1" colspan="1">VIC</td><td align="center" rowspan="1" colspan="1">NA</td><td rowspan="1" colspan="1">KP702752</td></tr><tr><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1">R: GTATGCCTCGGAGATGGTGT</td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td></tr><tr><td rowspan="1" colspan="1">Ad-126</td><td rowspan="1" colspan="1">F: TCCACCCAAGAACTGATGAA</td><td rowspan="1" colspan="1">(AC)<sub>11</sub></td><td align="center" rowspan="1" colspan="1">111–121</td><td rowspan="1" colspan="1">NED</td><td align="center" rowspan="1" colspan="1">2</td><td rowspan="1" colspan="1">KP702753</td></tr><tr><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1">R: CAGGCGTCCCATAAACTTGT</td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td></tr><tr><td rowspan="1" colspan="1">Ad-127</td><td rowspan="1" colspan="1">F: GATGGTTATGGTGCCTCAAG</td><td rowspan="1" colspan="1">(GT)<sub>9</sub></td><td align="center" rowspan="1" colspan="1">106–136</td><td rowspan="1" colspan="1">VIC</td><td align="center" rowspan="1" colspan="1">1</td><td rowspan="1" colspan="1">KP702754</td></tr><tr><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1">R: GGCCTAAACCACAAAGGTGA</td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td></tr><tr><td rowspan="1" colspan="1">Ad-137</td><td rowspan="1" colspan="1">F: ACCCTCAACCCTGACTTCCT</td><td rowspan="1" colspan="1">(AC)<sub>9</sub></td><td align="center" rowspan="1" colspan="1">114–128</td><td rowspan="1" colspan="1">VIC</td><td align="center" rowspan="1" colspan="1">2</td><td rowspan="1" colspan="1">KP702755</td></tr><tr><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1">R: AACCCTCAATCCTGCAAGTG</td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td></tr><tr><td rowspan="1" colspan="1">Ad-145</td><td rowspan="1" colspan="1">F: CGTGAAAATGGCCTGTAGGT</td><td rowspan="1" colspan="1">(AG)<sub>10</sub></td><td align="center" rowspan="1" colspan="1">234–284</td><td rowspan="1" colspan="1">NED</td><td align="center" rowspan="1" colspan="1">1</td><td rowspan="1" colspan="1">KP702756</td></tr><tr><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1">R: CCTTCAACTCATTCACCATCTC</td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td></tr><tr><td rowspan="1" colspan="1">Ad-173</td><td rowspan="1" colspan="1">F: TCTCAACCTCAAACTGCGAA</td><td rowspan="1" colspan="1">(TG)<sub>8</sub></td><td align="center" rowspan="1" colspan="1">150–160</td><td rowspan="1" colspan="1">NED</td><td align="center" rowspan="1" colspan="1">2</td><td rowspan="1" colspan="1">KP702757</td></tr><tr><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1">R: GAGGTGAAGGCCAAATGAAG</td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td></tr><tr><td rowspan="1" colspan="1">Ad-176</td><td rowspan="1" colspan="1">F: TGACTGTGCGGAATTCATCT</td><td rowspan="1" colspan="1">(AC)<sub>9</sub></td><td align="center" rowspan="1" colspan="1">114–130</td><td rowspan="1" colspan="1">FAM</td><td align="center" rowspan="1" colspan="1">1</td><td rowspan="1" colspan="1">KP702758</td></tr><tr><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1">R: TCCTTATAAACAATGCGACATAGA</td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td></tr><tr><td rowspan="1" colspan="1">Ad-177</td><td rowspan="1" colspan="1">F: GCCAGCAAAACAAACACAGA</td><td rowspan="1" colspan="1">(CA)<sub>8</sub></td><td align="center" rowspan="1" colspan="1">149–151</td><td rowspan="1" colspan="1">PET</td><td align="center" rowspan="1" colspan="1">1</td><td rowspan="1" colspan="1">KP702759</td></tr><tr><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1">R: CTGCAGTCCTAAATGTCAGTGC</td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td></tr><tr><td rowspan="1" colspan="1">Ad-201</td><td rowspan="1" colspan="1">F: GACAAGTCTTGCCATGATGTTT</td><td rowspan="1" colspan="1">(TG)<sub>8</sub></td><td align="center" rowspan="1" colspan="1">197–210</td><td rowspan="1" colspan="1">VIC</td><td align="center" rowspan="1" colspan="1">1</td><td rowspan="1" colspan="1">KP702760</td></tr><tr><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1">R: CCATCCACTTTTTGCCTGAT</td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td><td rowspan="1" colspan="1"></td></tr></tbody></table><table-wrap-foot><fn id="tblfn1"><label>a</label><p>Repeat motifs are those of the pyrosequenced alleles.</p></fn><fn id="tblfn2"><label>b</label><p>Size ranges are based on the allele scoring performed on capillary electrophoresis data.</p></fn><fn id="tblfn3"><label>c</label><p>Two sets of possible multiplex reactions are listed. NA indicates that the primer pair produces unspecific products in multiplex.</p></fn></table-wrap-foot></table-wrap><p>The number of alleles measured on 32 individuals sampled from a single Australian population varied from two to 11, and Nei’s expected heterozygosity (<xref rid="bib11" ref-type="bibr">Nei, 1987</xref>) varied from 0.11 to 0.88 and was larger than 0.5 for 14 loci (<xref ref-type="table" rid="tbl2">Table 2</xref>). Deviations from Hardy–Weinberg equilibrium (HWE) and linkage equilibrium were tested using GENEPOP 4.0 (<xref rid="bib12" ref-type="bibr">Rousset, 2008</xref>). We took into account multiple testing (in the case of HWE tests) and nonindependence between tests (in the case of linkage tests) by using the false discovery rate (<xref rid="bib2" ref-type="bibr">Benjamini and Hochberg, 1995</xref>) and the sequential Bonferroni (<xref rid="bib13" ref-type="bibr">Sokal and Rolf, 1995</xref>: p. 236) adjustments, respectively. Five loci (Ad-145, Ad-41, Ad-59, Ad-116, and Ad-45) significantly departed from HWE after false discovery rate adjustment and displayed significant heterozygote deficiency (<xref ref-type="table" rid="tbl1">Table 1</xref>). Linkage disequilibrium was detected for three pairs of loci (between Ad-116 and Ad-137, Ad-116 and Ad-97, and Ad-49 and Ad-86). Twenty-one microsatellite loci could be amplified in two sets of PCR multiplex mixes, one with 14 and the other with seven primer pairs (<xref ref-type="table" rid="tbl1">Table 1</xref>). The two other primer pairs produced unspecific amplifications when used in multiplex PCR and thus were not included in any multiplex mix.</p><table-wrap id="tbl2" position="float"><label>Table 2.</label><caption><p>Variability of the 23 microsatellite loci developed for <italic>Acacia dealbata</italic> based on 32 individuals sampled in southeastern Australia.</p></caption><table frame="hsides" rules="groups"><thead><tr><td rowspan="1" colspan="1">Locus</td><td align="center" rowspan="1" colspan="1"><italic>A</italic></td><td align="center" rowspan="1" colspan="1"><italic>H</italic><sub>o</sub></td><td align="center" rowspan="1" colspan="1"><italic>H</italic><sub>e</sub></td><td align="center" rowspan="1" colspan="1">HW<xref ref-type="table-fn" rid="tblfn4"><sup>a</sup></xref></td></tr></thead><tbody><tr><td rowspan="1" colspan="1">Ad-33</td><td align="char" char="." rowspan="1" colspan="1">6</td><td align="char" char="." rowspan="1" colspan="1">0.677</td><td align="char" char="." rowspan="1" colspan="1">0.651</td><td rowspan="1" colspan="1"></td></tr><tr><td rowspan="1" colspan="1">Ad-41</td><td align="char" char="." rowspan="1" colspan="1">4</td><td align="char" char="." rowspan="1" colspan="1">0.344</td><td align="char" char="." rowspan="1" colspan="1">0.579</td><td align="center" rowspan="1" colspan="1">*, FDR</td></tr><tr><td rowspan="1" colspan="1">Ad-45</td><td align="char" char="." rowspan="1" colspan="1">4</td><td align="char" char="." rowspan="1" colspan="1">0.438</td><td align="char" char="." rowspan="1" colspan="1">0.588</td><td align="center" rowspan="1" colspan="1">*, FDR</td></tr><tr><td rowspan="1" colspan="1">Ad-47</td><td align="char" char="." rowspan="1" colspan="1">4</td><td align="char" char="." rowspan="1" colspan="1">0.656</td><td align="char" char="." rowspan="1" colspan="1">0.613</td><td rowspan="1" colspan="1"></td></tr><tr><td rowspan="1" colspan="1">Ad-48</td><td align="char" char="." rowspan="1" colspan="1">6</td><td align="char" char="." rowspan="1" colspan="1">0.5</td><td align="char" char="." rowspan="1" colspan="1">0.569</td><td rowspan="1" colspan="1"></td></tr><tr><td rowspan="1" colspan="1">Ad-49</td><td align="char" char="." rowspan="1" colspan="1">9</td><td align="char" char="." rowspan="1" colspan="1">0.813</td><td align="char" char="." rowspan="1" colspan="1">0.858</td><td rowspan="1" colspan="1"></td></tr><tr><td rowspan="1" colspan="1">Ad-54</td><td align="char" char="." rowspan="1" colspan="1">5</td><td align="char" char="." rowspan="1" colspan="1">0.438</td><td align="char" char="." rowspan="1" colspan="1">0.449</td><td rowspan="1" colspan="1"></td></tr><tr><td rowspan="1" colspan="1">Ad-59</td><td align="char" char="." rowspan="1" colspan="1">5</td><td align="char" char="." rowspan="1" colspan="1">0.29</td><td align="char" char="." rowspan="1" colspan="1">0.457</td><td align="center" rowspan="1" colspan="1">*, FDR</td></tr><tr><td rowspan="1" colspan="1">Ad-63</td><td align="char" char="." rowspan="1" colspan="1">5</td><td align="char" char="." rowspan="1" colspan="1">0.406</td><td align="char" char="." rowspan="1" colspan="1">0.458</td><td rowspan="1" colspan="1"></td></tr><tr><td rowspan="1" colspan="1">Ad-66</td><td align="char" char="." rowspan="1" colspan="1">3</td><td align="char" char="." rowspan="1" colspan="1">0.531</td><td align="char" char="." rowspan="1" colspan="1">0.478</td><td rowspan="1" colspan="1"></td></tr><tr><td rowspan="1" colspan="1">Ad-70</td><td align="char" char="." rowspan="1" colspan="1">2</td><td align="char" char="." rowspan="1" colspan="1">0.125</td><td align="char" char="." rowspan="1" colspan="1">0.119</td><td rowspan="1" colspan="1"></td></tr><tr><td rowspan="1" colspan="1">Ad-86</td><td align="char" char="." rowspan="1" colspan="1">7</td><td align="char" char="." rowspan="1" colspan="1">0.531</td><td align="char" char="." rowspan="1" colspan="1">0.472</td><td rowspan="1" colspan="1"></td></tr><tr><td rowspan="1" colspan="1">Ad-89</td><td align="char" char="." rowspan="1" colspan="1">6</td><td align="char" char="." rowspan="1" colspan="1">0.4</td><td align="char" char="." rowspan="1" colspan="1">0.525</td><td rowspan="1" colspan="1"></td></tr><tr><td rowspan="1" colspan="1">Ad-97</td><td align="char" char="." rowspan="1" colspan="1">3</td><td align="char" char="." rowspan="1" colspan="1">0.29</td><td align="char" char="." rowspan="1" colspan="1">0.263</td><td rowspan="1" colspan="1"></td></tr><tr><td rowspan="1" colspan="1">Ad-116</td><td align="char" char="." rowspan="1" colspan="1">4</td><td align="char" char="." rowspan="1" colspan="1">0.625</td><td align="char" char="." rowspan="1" colspan="1">0.668</td><td align="center" rowspan="1" colspan="1">*, FDR</td></tr><tr><td rowspan="1" colspan="1">Ad-126</td><td align="char" char="." rowspan="1" colspan="1">6</td><td align="char" char="." rowspan="1" colspan="1">0.625</td><td align="char" char="." rowspan="1" colspan="1">0.58</td><td rowspan="1" colspan="1"></td></tr><tr><td rowspan="1" colspan="1">Ad-127</td><td align="char" char="." rowspan="1" colspan="1">8</td><td align="char" char="." rowspan="1" colspan="1">0.633</td><td align="char" char="." rowspan="1" colspan="1">0.791</td><td rowspan="1" colspan="1"></td></tr><tr><td rowspan="1" colspan="1">Ad-137</td><td align="char" char="." rowspan="1" colspan="1">7</td><td align="char" char="." rowspan="1" colspan="1">0.563</td><td align="char" char="." rowspan="1" colspan="1">0.606</td><td rowspan="1" colspan="1"></td></tr><tr><td rowspan="1" colspan="1">Ad-145</td><td align="char" char="." rowspan="1" colspan="1">11</td><td align="char" char="." rowspan="1" colspan="1">0.296</td><td align="char" char="." rowspan="1" colspan="1">0.876</td><td align="center" rowspan="1" colspan="1">*, FDR</td></tr><tr><td rowspan="1" colspan="1">Ad-173</td><td align="char" char="." rowspan="1" colspan="1">6</td><td align="char" char="." rowspan="1" colspan="1">0.594</td><td align="char" char="." rowspan="1" colspan="1">0.555</td><td rowspan="1" colspan="1"></td></tr><tr><td rowspan="1" colspan="1">Ad-176</td><td align="char" char="." rowspan="1" colspan="1">6</td><td align="char" char="." rowspan="1" colspan="1">0.313</td><td align="char" char="." rowspan="1" colspan="1">0.427</td><td align="center" rowspan="1" colspan="1">*</td></tr><tr><td rowspan="1" colspan="1">Ad-177</td><td align="char" char="." rowspan="1" colspan="1">2</td><td align="char" char="." rowspan="1" colspan="1">0.438</td><td align="char" char="." rowspan="1" colspan="1">0.437</td><td rowspan="1" colspan="1"></td></tr><tr><td rowspan="1" colspan="1">Ad-201</td><td align="char" char="." rowspan="1" colspan="1">5</td><td align="char" char="." rowspan="1" colspan="1">0.531</td><td align="char" char="." rowspan="1" colspan="1">0.58</td><td rowspan="1" colspan="1"></td></tr></tbody></table><table-wrap-foot><fn><p><italic>Note</italic>: <italic>A</italic> = number of alleles; <italic>H</italic><sub>e</sub> = expected heterozygosity of <xref rid="bib11" ref-type="bibr">Nei (1987)</xref>; <italic>H</italic><sub>o</sub> = observed heterozygosity; HW = first type error of the probability test of Hardy–Weinberg equilibrium.</p></fn><fn id="tblfn4"><label>a</label><p>An asterisk (*) indicates a significant test at the 5% level, and “FDR” indicates that the test is significant after the false discovery rate adjustment for multiple testing.</p></fn></table-wrap-foot></table-wrap></sec><sec><title>CONCLUSIONS</title><p>We chose to present all 23 primer pairs although some of them may display Hardy–Weinberg or linkage disequilibrium because those disequilibria may be specific to the population sampled for the current study. 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