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Giving IDs to turtles: SNP markers for assignment of individuals to lineages of the geographically structured Phrynops geoffroanus (Chelidae: Testudines)

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Abstract

We developed forensic nuclear SNPs for the side-necked turtle, Phrynops geoffroanus, using next-generation sequencing-based genotyping. We created a reduced-representation library for 23 individuals representing four previously identified lineages of P. geoffroanus using the ddRAD methodology for sequencing on the IonTorrent PGM. We analyzed the sequence data using the Stacks pipeline. We found 811 loci containing 1380 private SNPs. Of these, 473 were fixed in lineages 1, 2 and 3, or with frequency >0.91 in lineage 4. Seventy-three SNPs could be useful for forensics purpose (assignment of individuals to lineage of origin) using traditional PCR-based genotyping methods.

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Acknowledgments

This work was supported by the SISBIOTA/Conselho Nacional de Desenvolvimento Científico e Tecnológico/Fundação de Amparo à Pesquisa do Amazonas (CNPq/FAPEAM/SISBIOTA-BioPHAM) Grant No. 563348/2010 to IPF; CNPq Ciencia sem Fronteiras Project No. 400771/2013-6 to RCV; and Mohamed bin Zayed Species Conservation Fund Project No. 1025921 to RCV and VTC. We thank the Instituto Chico Mendes de Conservação da Biodiversidade for collection permits (ICMBio, processes 22885-1 and 43175-2). All specimens used were deposited in the Collection of Amphibians and Reptiles of the Instituto Nacional de Pesquisas da Amazonia. We are grateful to Enedina Nogueira of the Laboratório de Tecnologias de DNA for technical support during sequencing. VC was supported by Bolsa de Pesquisa scholarship from FAPEAM during his Doctoral studies in the Biodiversidade e Biotecnologia/Rede BIONORTE Program in the UFAM. IPF and TH were supported by a Bolsa de Pesquisa scholarship from CNPq during this study. JGM was supported by postdoctoral scholarship from CAPES in the MBT program of UEA.

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Authors and Affiliations

  1. Laboratório de Evolução e Genética Animal, Departamento de Genética, Universidade Federal do Amazonas, Av. Rodrigo Octávio Jordão Ramos, 3000, Manaus, Amazonas, 69077-000, Brazil

    Vinícius Tadeus de Carvalho, José Gregorio Martínez, Sandra M. Hernández-Rangel, Izeni Pires Farias & Tomas Hrbek

  2. Programa de Pós-graduação em Biodiversidade e Biotecnologia da Amazônia Legal—Rede Bionorte, Universidade Federal do Amazonas, Av. Rodrigo Octávio Jordão Ramos, 3000, Manaus, Amazonas, 69077-000, Brazil

    Vinícius Tadeus de Carvalho

  3. Laboratorio de Ecología Molecular de Vertebrados Acuáticos, Facultad de Ciencias, Departamento de Ciencias Biológicas, Universidad de los Andes, Cra 1 No. 18A-12, Código Postal 4–72, Bogotá, Colombia

    José Gregorio Martínez

  4. Laboratório de Proteômica e Genômica, Programa de Pós-graduação Mestrado em Biotecnologia e Recursos Naturais (MBT), Grupo de Pesquisa em Genética Molecular e Citogenética, Universidade do Estado do Amazonas, Manaus, Amazonas, 69065-001, Brazil

    José Gregorio Martínez

  5. Centro de Apoio Muldisciplinar, Laboratório de Tecnologias de DNA, Universidade Federal do Amazonas, Av. Rodrigo Octávio Jordão Ramos, 3000, Manaus, Amazonas, 69077-000, Brazil

    Spartaco Astolfi-Filho

  6. Coleção de Anfíbios e Répteis, Instituto Nacional de Pesquisas da Amazônia, Coordenação em Biodiversidade, Av. André Araújo, 2936, Aleixo, Manaus, Amazonas, 690600-001, Brazil

    Vinícius Tadeus de Carvalho & Richard C. Vogt

Authors
  1. Vinícius Tadeus de Carvalho
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  2. José Gregorio Martínez
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  3. Sandra M. Hernández-Rangel
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  4. Spartaco Astolfi-Filho
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  5. Richard C. Vogt
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  6. Izeni Pires Farias
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  7. Tomas Hrbek
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Corresponding author

Correspondence to Tomas Hrbek.

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The authors declare that they have no conflict of interest.

Electronic supplementary material

Below is the link to the electronic supplementary material.

12686_2016_626_MOESM1_ESM.txt

Supplementary material 1. Online Resource 1 Command line script file used for running Stacks pipeline programs components (process_radtags.pl, denovo_map.pl and populations.pl) for de novo SNPs genotyping across all individuals (Lineages 1, 2, 3 and 4) of the side-necked turtle Phrynops geoffroanus (TXT 2 KB)

12686_2016_626_MOESM2_ESM.txt

Supplementary material 2. Online Resource 2 SUMSTATS (summary statistics) file from Stacks pipeline (populations.pl) showing the loci, SNP position in the tag loci, population source, most frequent allele, alternative allele, frequency of the most frequent allele, number of individuals from which the SNP was recovered, heterozygosis for each one of the final 73 recovered SNPs from the Lineages 1, 2, 3 and 4 of the side-necked turtle, Phrynops geoffroanus (TXT 6 KB)

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de Carvalho, V.T., Martínez, J.G., Hernández-Rangel, S.M. et al. Giving IDs to turtles: SNP markers for assignment of individuals to lineages of the geographically structured Phrynops geoffroanus (Chelidae: Testudines). Conservation Genet Resour 9, 157–163 (2017). https://doi.org/10.1007/s12686-016-0626-8

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