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Isolation and characterization of microsatellite loci in the Charal de Xochimilco Chirostoma humboldtianum
Aislamiento y caracterización de loci de microsatelites en el charal de Xochimilco Chirostoma humboldtianum
Rosa María García-Martínez1, Francisco Javier García-De León2, Omar Mejía3
1 Labooratorio de Genética y Biología Molecular de la Planta Experimental de Producción Acuícola, Departamento de Hidrobiología, Universidad Autónoma Metropolitana-Iztapalapa. San Rafael Atlixco 186, Vicentina, 09340 México, D. F., Mexico
2 Laboratorio de Genética para la Conservación, Centro de Investigaciones Biológicas del Noroeste, S. C. Instituto Politécnico Nacional 195, Playa Palo de Santa Rita Sur, 23096 La Paz, Baja California Sur, Mexico
3 Laboratorio de Variación Biológica y Evolución, Departamento de Zoología, Escuela Nacional de Ciencias Biológicas, Instituto Politécnico Nacional. Prolongación de Carpio y Plan de Ayala S/N, Colonia Casco de Santo Tomás, 11340 México, D. F., Mexico
Irene de los Ángeles Barriga-Sosa
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    "textoCompleto" => "<span class="elsevierStyleSections"><p id="par0005" class="elsevierStylePara elsevierViewall">The Charal de Xochimilco <span class="elsevierStyleItalic">Chirostoma humboldtianum</span> &#40;Atherinopsidae&#58; Menidiinae&#41; is an endemic species from Central Mexico&#46; In the last 6 decades their populations have been reduced or extirpated from some of their natural habitats as a result of habitat alteration &#40;<a class="elsevierStyleCrossRef" href="#bib0010">Barbour&#44; 1973</a>&#41;&#46; However&#44; the population size of this species has not been determined&#44; and the population genetic structure remains unknown&#44; thus&#44; microsatellites markers can be useful addressing related questions&#46; We isolated and characterized 8 polymorphic microsatellite loci for this species in order to increase our knowledge on the population structure and to test hypothesis about diversification and speciation that aid in defining strategies for the conservation and maintenance of the species&#46;</p><p id="par0010" class="elsevierStylePara elsevierViewall">Enrichment procedures were utilized to develop microsatellite genetic markers &#40;<a class="elsevierStyleCrossRef" href="#bib0050">Glenn and Schable&#44; 2005</a>&#41;&#46; Out of 165- screened clones&#44; 29 possessed repetitive elements that allow designing primers for 12 loci according to MSATCOMMANDER &#40;<a class="elsevierStyleCrossRef" href="#bib0045">Faircloth&#44; 2008</a>&#41;&#44; the remaining 17 clones had flanking region that did not allow primer design&#46; PCR reactions were carried<a name="p1283"></a> out using15<span class="elsevierStyleHsp" style=""></span>&#956;l reactions containing 50 to 100<span class="elsevierStyleHsp" style=""></span>ng DNA&#44; 10 pmol each primer&#44; 0&#46;5<span class="elsevierStyleHsp" style=""></span>U <span class="elsevierStyleItalic">Taq</span> DNA polymerase &#40;Promega&#41;&#44; 200<span class="elsevierStyleHsp" style=""></span>&#956;M of each dNTP&#44; 2&#46;5<span class="elsevierStyleHsp" style=""></span>&#956;mM MgCl<span class="elsevierStyleInf">2</span> and 1X PCR buffer &#40;15<span class="elsevierStyleHsp" style=""></span>&#956;mM MgCl<span class="elsevierStyleInf">2</span>&#44; 200<span class="elsevierStyleHsp" style=""></span>&#956;mMTris-HCl&#44; pH 8&#46;5&#44;75<span class="elsevierStyleHsp" style=""></span>&#956;mM &#40;NH<span class="elsevierStyleInf">4</span>&#41;<span class="elsevierStyleInf">2</span>SO<span class="elsevierStyleInf">4</span>&#41;&#46; PCR amplifications were performed in a BIO-RAD thermocycler as follows&#58; 95<span class="elsevierStyleHsp" style=""></span>&#176; C for 5<span class="elsevierStyleHsp" style=""></span>min or 10<span class="elsevierStyleHsp" style=""></span>min&#44; followed by 30 cycles of 15<span class="elsevierStyleHsp" style=""></span>sec at 94<span class="elsevierStyleHsp" style=""></span>&#176; C&#44; 45<span class="elsevierStyleHsp" style=""></span>sec at 50<span class="elsevierStyleHsp" style=""></span>&#176; C to 60<span class="elsevierStyleHsp" style=""></span>&#176; C depending on the locus and 15<span class="elsevierStyleHsp" style=""></span>sec at 72<span class="elsevierStyleHsp" style=""></span>&#176; C&#44; with a final extension at 70<span class="elsevierStyleHsp" style=""></span>&#176; C for 5<span class="elsevierStyleHsp" style=""></span>min&#46; The PCR products were analyzed by capillary electrophoresis in the automatic sequencer ABI Prism 3100 Avant at Laboratorio Divisional de Biolog&#237;a Molecular &#40;LDBM&#41; of the UAM- Iztapalapa&#46; Allele sizes were determined using LIZ-500 as size standard &#40;Applied Biosystems&#41; and GeneMarker 2&#46;4&#46;0 software&#46;</p><p id="par0015" class="elsevierStylePara elsevierViewall">Polymorphism for each microsatellite loci was characterized by screening a sample of 32 to 48 individuals of <span class="elsevierStyleItalic">C&#46; humboldtianum</span> from Tepuxtepec Dam&#44; Michoac&#225;n &#40;19&#176;59&#39;42&#8221; N&#44; 100&#176;13&#39;33&#8221; W&#41;&#46; The total number of alleles per locus&#44; observed &#40;H<span class="elsevierStyleInf"><span class="elsevierStyleItalic">0</span></span>&#41; and expected &#40;H<span class="elsevierStyleInf"><span class="elsevierStyleItalic">E</span></span>&#41; heterozygosities were calculated&#46; All loci were tested for Hardy-Weinberg equilibrium &#40;HWE&#41;&#44; and all pairwise combinations of loci were tested for linkage disequilibrium &#40;LD&#41;&#46; All these parameters and tests were computed in Arlequin version 3&#46;5&#46;1&#46;2 &#40;<a class="elsevierStyleCrossRef" href="#bib0040">Excoffier and Lischer&#44; 2010</a>&#41;&#46; Microchecker version 2&#46;2&#46;3 &#40;<a class="elsevierStyleCrossRef" href="#bib0065">Van Oosterhout et al&#46;&#44; 2004</a>&#41; was used to estimate the frequency of null alleles in the microsatellites markers&#46; The genotypes of the loci that showed null alleles were corrected using FreeNA &#40;<a class="elsevierStyleCrossRef" href="#bib0025">Chapui and Estoup&#44; 2007</a>&#41;&#46; Finally&#44; the <span class="elsevierStyleItalic">C&#46; humboldtianum</span> population was evaluated in order to detect the occurrence of genetic bottleneck using TPM &#40;two phases mutation&#41; model and the Wilcoxon rank test as implemented in Bottleneck ver&#46; 1&#46;2&#46;1 &#40;<a class="elsevierStyleCrossRef" href="#bib0030">Cornuet and Luikart 1996</a>&#41;&#46;</p><p id="par0020" class="elsevierStylePara elsevierViewall">From the 12 amplified microsatellite loci&#44; 4&#44; Chum422&#44; 443&#44; 490 and 363 were monomorphic in all the individuals analyzed&#46; The number of alleles per locus in the remaining 8 loci ranged from 3 &#40;Chum21&#41; to 10 &#40;Chum496&#41;&#46; Expected and observed heterozygosities ranged from 0&#46;44 to 0&#46;78 and 0&#46;34 to 0&#46;89&#44; respectively &#40;<a class="elsevierStyleCrossRef" href="#tbl0005">Table 1</a>&#41;&#46; No LD was detected between each pair of loci&#46; All loci exhibited HWE departure after the sequential Bonferroni correction &#40;<span class="elsevierStyleItalic">&#945;</span><span class="elsevierStyleHsp" style=""></span>&#61;<span class="elsevierStyleHsp" style=""></span>0&#46;05&#44; <span class="elsevierStyleItalic">k</span><span class="elsevierStyleHsp" style=""></span>&#61;<span class="elsevierStyleHsp" style=""></span>8&#41;&#46; Micro-Checker &#40;<a class="elsevierStyleCrossRef" href="#bib0065">Van Oosterhout et al&#46;&#44; 2004</a>&#41; suggested that this phenomenon might be due to the presence of null alleles that were present in 3 of the 8 loci &#40;Chum496&#44; Chum450 and Chum358&#41;&#46; However&#44; the HWE departures were still observed after the genotyping correction using FreeNA&#46; Deviation from HWE may be outcome of heterozygote deficiency found in some loci &#40;Chum387&#44; Chum411&#44; Chum450&#44; Chum358&#41;&#44; which in turn might be caused by endogamy as has been suggested in other species of the genus &#40;<a class="elsevierStyleCrossRef" href="#bib0015">Barriga-Sosa et al&#46;&#44; 2004</a>&#41;&#46; On the other hand&#44; an excess of heterozygotes were detected in the remaining 4 loci&#46; Heterozygosity excess has been<a name="p1284"></a> associated with population bottleneck events &#40;<a class="elsevierStyleCrossRef" href="#bib0030">Cornuet and Luikart&#44; 1996</a>&#41;&#44; and has also been a phenomenon suggested in other freshwater fish species inhabiting central Mexico &#40;<a class="elsevierStyleCrossRef" href="#bib0035">Dom&#237;nguez-Dom&#237;nguez et al&#46;&#44; 2008</a>&#41;&#46; The bottleneck event could be associated to disturbances and habitat loss caused by human activities&#44; which in turn might give result to small population sizes causing deviations from HWE&#46; The TPM under Wilcoxon test did not show significant recent bottleneck &#40;heterozygosity excess&#41; &#40;<span class="elsevierStyleItalic">p</span><span class="elsevierStyleHsp" style=""></span>&#61;<span class="elsevierStyleHsp" style=""></span>0&#46;187&#41;&#46; Likewise&#44; decrease in observed heterozygosity suggests nonrandom mating and genetic drift &#40;<a class="elsevierStyleCrossRef" href="#bib0055">Loew et al&#46;&#44; 2005</a>&#41;&#44; due that genetic drift causes random fixation and loss of alleles within population&#46; In contrast&#44; heterozygote excess appears to be due to the presence of hybrids within the sample &#40;<a class="elsevierStyleCrossRef" href="#bib0060">Pitchar et al&#46;&#44; 2007</a>&#41; as has been earlier suggested for species of the genus &#40;<a class="elsevierStyleCrossRef" href="#bib0020">Barriga-Sosa et al&#46;&#44; 2001</a>&#41;&#46; Translocations among populations and hybridization have been reported for this and others species of the genus &#40;<a class="elsevierStyleCrossRef" href="#bib0005">Alaye&#44; 1996</a>&#59; <a class="elsevierStyleCrossRef" href="#bib0020">Barriga-Sosa et al&#46;&#44; 2001</a>&#41;&#44; however&#44; this issues needs to be punctually addressed in further studies&#46; These 8 microsatellite loci are the first developed for <span class="elsevierStyleItalic">C&#46; humboldtianum</span> and will be useful in investigating population genetics&#44; conservation&#44; and management of this and closely related species&#46;</p><elsevierMultimedia ident="tbl0005"></elsevierMultimedia><p id="par0025" class="elsevierStylePara elsevierViewall">This study is part of the doctoral research of Rosa Mar&#237;a Garc&#237;a Mart&#237;nez at Doctorado en Ciencias Biol&#243;gicas y de la Salud at Universidad Aut&#243;noma Metropolitana- Iztapalapa&#44; Mexico City&#59; with fellowship Conacyt-224707&#46; We thank PhD Rub&#233;n Valles and PhD Miguel Correa for laboratory assistance at CIBNOR and Biol&#46; Ram&#243;n Cisneros for collecting specimens&#46; Funding for this research was provided by Conacyt-CB-2009-01-130220 and UAM&#46;147&#46;07&#46;03&#47;147&#46;09&#46;01 granted to IDLABS&#46; Mexican government kindly issued permit number DG0PA&#46;07343&#46;310810&#46;4128 to conduct this research&#46;</p></span>"
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          "palabras" => array:4 [
            0 => "Atherinopsidae"
            1 => "Hardy-Weinberg"
            2 => "alelos nulos"
            3 => "enriquecimiento"
          ]
        ]
      ]
    ]
    "tieneResumen" => true
    "resumen" => array:2 [
      "en" => array:2 [
        "titulo" => "Abstract"
        "resumen" => "<span id="abst0005" class="elsevierStyleSection elsevierViewall"><p id="spar0015" class="elsevierStyleSimplePara elsevierViewall">Microsatellite loci were isolated and characterized for the endemic fish <span class="elsevierStyleItalic">Chirostoma humboldtianum</span> using an enrichment procedure&#46; Eight polymorphic microsatellites were genotyped for 32 - 48 individuals from Tepuxtepec Dam&#44; Michoac&#225;n&#46; The number of alleles per locus ranged from 3 to 11 and the average observed and expected heterozygosities were 0&#46;61 and 0&#46;63&#44; respectively&#46; All loci deviated significantly from Hardy-Weinberg expectations&#44; which might be related to small population sizes associated to human disturbances and habitat loss&#46; These are the first loci described for the species and the genus and could be useful in studies of population genetics&#44; conservation and management of the species&#46;</p></span>"
      ]
      "es" => array:2 [
        "titulo" => "Resumen"
        "resumen" => "<span id="abst0010" class="elsevierStyleSection elsevierViewall"><p id="spar0020" class="elsevierStyleSimplePara elsevierViewall">Se aislaron y caracterizaron los loci microsat&#233;lites para el pez end&#233;mico <span class="elsevierStyleItalic">Chirostoma humboldtianum</span> a partir de una genoteca enriquecida&#46; Se genotipificaron 8 microsat&#233;lites polim&#243;rficos para 32 a 48 individuos de la presa Tepuxtec&#44; Michoac&#225;n&#46; El n&#250;mero de alelos por locus vari&#243; de 3 a 11 y la heterocigosidad observada y esperada promedio fue de 0&#46;61 y 0&#46;63&#44; respectivamente&#46; Todos los loci presentaron desviaciones significativas del equilibrio de Hardy-Weinberg&#44; lo cual puede deberse al tama&#241;o poblacional peque&#241;o asociado a disturbios humanos y p&#233;rdida del h&#225;bitat&#46; &#201;stos son los primeros loci descritos para la especie y el g&#233;nero y podr&#237;an ser &#250;tiles en estudios de gen&#233;tica de poblaciones&#44; conservaci&#243;n y manejo de la especie&#46;</p></span>"
      ]
    ]
    "multimedia" => array:1 [
      0 => array:7 [
        "identificador" => "tbl0005"
        "etiqueta" => "Table 1"
        "tipo" => "MULTIMEDIATABLA"
        "mostrarFloat" => true
        "mostrarDisplay" => false
        "tabla" => array:2 [
          "leyenda" => "<p id="spar0010" class="elsevierStyleSimplePara elsevierViewall">p<span class="elsevierStyleHsp" style=""></span>&#60;<span class="elsevierStyleHsp" style=""></span>0&#46;05 after sequential Bonferroni corrections</p>"
          "tablatextoimagen" => array:1 [
            0 => array:2 [
              "tabla" => array:1 [
                0 => """
                  <table border="0" frame="\n
                  \t\t\t\t\tvoid\n
                  \t\t\t\t" class=""><thead title="thead"><tr title="table-row"><th class="td" title="table-head  " align="left" valign="middle" scope="col" style="border-bottom: 2px solid black"><span class="elsevierStyleItalic">Locus and GenBank accession</span>&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</th><th class="td" title="table-head  " align="center" valign="middle" scope="col" style="border-bottom: 2px solid black"><span class="elsevierStyleItalic">Primer sequences &#40;5&#39;-3&#39;&#41;</span>&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</th><th class="td" title="table-head  " align="center" valign="middle" scope="col" style="border-bottom: 2px solid black"><span class="elsevierStyleItalic">Repeat motif</span>&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</th><th class="td" title="table-head  " align="center" valign="middle" scope="col" style="border-bottom: 2px solid black"><span class="elsevierStyleItalic">Ta &#40;&#176;C&#41;</span>&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</th><th class="td" title="table-head  " align="center" valign="middle" scope="col" style="border-bottom: 2px solid black"><span class="elsevierStyleItalic">Allele size &#40;pb&#41;</span>&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</th><th class="td" title="table-head  " align="center" valign="middle" scope="col" style="border-bottom: 2px solid black"><span class="elsevierStyleItalic">n</span>&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</th><th class="td" title="table-head  " align="center" valign="middle" scope="col" style="border-bottom: 2px solid black"><span class="elsevierStyleItalic">N</span><span class="elsevierStyleInf"><span class="elsevierStyleItalic">A</span></span>&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</th><th class="td" title="table-head  " align="center" valign="middle" scope="col" style="border-bottom: 2px solid black"><span class="elsevierStyleItalic">H</span><span class="elsevierStyleInf"><span class="elsevierStyleItalic">O</span></span>&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</th><th class="td" title="table-head  " align="center" valign="middle" scope="col" style="border-bottom: 2px solid black"><span class="elsevierStyleItalic">H</span><span class="elsevierStyleInf"><span class="elsevierStyleItalic">E</span></span>&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</th><th class="td" title="table-head  " align="center" valign="middle" scope="col" style="border-bottom: 2px solid black"><span class="elsevierStyleItalic">P</span><span class="elsevierStyleInf"><span class="elsevierStyleItalic">HW</span></span>&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</th><th class="td" title="table-head  " align="center" valign="middle" scope="col" style="border-bottom: 2px solid black"><span class="elsevierStyleItalic">Null</span>&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</th></tr></thead><tbody title="tbody"><tr title="table-row"><td class="td" title="table-entry  " align="left" valign="middle">Chum48&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">F&#58; GCTAGCAGAAGCCATCTAAGTC&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">&#40;CA&#41;<span class="elsevierStyleInf">9</span>&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">50&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">132-170&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">46&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">4&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">0&#46;522&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">0&#46;484&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">0&#46;018&#42;&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">&#8722;0&#46;0289&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td></tr><tr title="table-row"><td class="td" title="table-entry  " align="left" valign="middle">KF016982&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">R&#58;TCTACAGGCTCATGAAAGCAG&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td></tr><tr title="table-row"><td class="td" title="table-entry  " align="left" valign="middle">Chum78&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">F&#58;TCCTCTCGCTGCTTCTCAC&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">&#40;CT&#41;<span class="elsevierStyleInf">34</span>&#46;&#40;A&#41;<span class="elsevierStyleInf">8</span>&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">52&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">220&#8211;252&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">40&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">7&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">0&#46;800&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">0&#46;731&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">0&#46;000&#42;&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">&#8722;0&#46;0452&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td></tr><tr title="table-row"><td class="td" title="table-entry  " align="left" valign="middle">KF016983&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">R&#58;CCTTCTGTGGCCTTGCTTG&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td></tr><tr title="table-row"><td class="td" title="table-entry  " align="left" valign="middle">Chum387&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">F&#58;AAGCTGCTGGACATTCTGC&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">&#40;T&#41;<span class="elsevierStyleInf">8</span>&#46;&#40;AC&#41;<span class="elsevierStyleInf">12</span>&#46;&#40;CT&#41;<span class="elsevierStyleInf">23</span>&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">55&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">245&#8211;310&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">43&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">7&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">0&#46;535&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">0&#46;719&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">0&#46;000&#42;&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">&#8722;0&#46;0106&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td></tr><tr title="table-row"><td class="td" title="table-entry  " align="left" valign="middle">KF016984&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">R&#58; TTGCACACAGCTTAAGCACATG&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td></tr><tr title="table-row"><td class="td" title="table-entry  " align="left" valign="middle">Chum496&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">F&#58;CTTTGGCTGTGGGAGGAAG&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">&#40;AG&#41;<span class="elsevierStyleInf">4</span>&#46;&#40;T&#41;<span class="elsevierStyleInf">17</span>&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">57&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">134&#8211;169&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">42&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">10&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">0&#46;786&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">0&#46;778&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">0&#46;000&#42;&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">0&#46;2658&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td></tr><tr title="table-row"><td class="td" title="table-entry  " align="left" valign="middle">KF016985&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">R&#58;AGCAGCTGAGTGTGCGAAAC&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td></tr><tr title="table-row"><td class="td" title="table-entry  " align="left" valign="middle">Chum21&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">F&#58;CCATGGCTCAGGGTGGGC&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">&#40;GA&#41;<span class="elsevierStyleInf">12</span>GGGC&#40;GA&#41;<span class="elsevierStyleInf">10</span>&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">58&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">78&#8211;106&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">46&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">3&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">0&#46;891&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">0&#46;625&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">0&#46;003&#42;&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">&#8722;0&#46;1686&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td></tr><tr title="table-row"><td class="td" title="table-entry  " align="left" valign="middle">KF016986&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">R&#58;GTCGTGTGTGCTTTCTCGTCC&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td></tr><tr title="table-row"><td class="td" title="table-entry  " align="left" valign="middle">Chum411&nbsp;\t\t\t\t\t\t\n
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                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">&nbsp;\t\t\t\t\t\t\n
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                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">F&#58;TCTCCGCTCATGTGTGTTTG&nbsp;\t\t\t\t\t\t\n
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                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="table-entry  " align="center" valign="middle">&nbsp;\t\t\t\t\t\t\n
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                  """
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          "en" => "<p id="spar0005" class="elsevierStyleSimplePara elsevierViewall">Primer sequences and microsatellite repeat motifs for 8 microsatellite loci on <span class="elsevierStyleItalic">Chirostoma humboldtianum&#46;</span> Annealing temperature &#40;Ta&#41;&#44; number of individuals &#40;n&#41;&#44; number of alleles &#40;N<span class="elsevierStyleInf">A</span>&#41;&#44; observed and expected heterozygosities &#40;Ho and He&#44; respectively&#41;&#44; p value for test of Hardy-Weinberg equilibrium &#40;P<span class="elsevierStyleInf">HW</span>&#41;&#44; expected null allele frequency &#40;Null&#41; and GenBank accession numbers</p>"
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Article information
ISSN: 18703453
Original language: English
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