10.5061/DRYAD.36775
Wielstra, Ben
University of Sheffield
Duijm, Elza
Naturalis Biodiversity Center
Lagler, Patricia
University of Natural Resources and Life Sciences
Lammers, Youri
Naturalis Biodiversity Center
Meilink, Willem R. M.
Vrije Universiteit Brussel
Ziermann, Janine M.
Naturalis Biodiversity Center
Arntzen, Jan W.
Naturalis Biodiversity Center
Data from: Parallel tagged amplicon sequencing of transcriptome-based
genetic markers for Triturus newts with the Ion Torrent next-generation
sequencing platform
Dryad
dataset
2014
Triturus
2014-02-19T21:24:23Z
2014-02-19T21:24:23Z
en
https://doi.org/10.1111/1755-0998.12242
1898243020 bytes
1
CC0 1.0 Universal (CC0 1.0) Public Domain Dedication
Next-generation sequencing is a fast and cost-effective way to obtain
sequence data for non-model organisms for many markers and for many
individuals. We describe a protocol through which we obtain orthologous
markers for the crested newts (Amphibia: Salamandridae: Triturus),
suitable for analysis of interspecific hybridization. We use transcriptome
data of a single Triturus species and design 96 primer pairs that amplify
c. 180 bp fragments positioned in 3-prime untranslated regions. Next these
markers are tested with uniplex PCR for a set of species spanning the
taxonomical width of the genus Triturus. The 52 markers that consistently
show a single band of expected length at gel electrophoreses for all
tested crested newt species are then amplified in five multiplex PCRs
(with a plexity of ten or eleven) for 132 individual newts: a set of 84
representing the seven (candidate) species and a set of 48 from a presumed
hybrid population. After pooling multiplexes per individual, unique tags
are ligated to link amplicons to individuals. Subsequently individuals are
pooled equimolar and sequenced on the Ion Torrent next-generation
sequencing platform. A bioinformatics pipeline identifies the alleles and
recodes these to a genotypic format. Next we test the utility of our
markers. BAPS allocates the 84 crested newt individuals representing
(candidate) species to their expected (candidate) species, confirming the
markers are suitable for species delineation. NewHybrids, a hybrid index
and HIest confirm the 48 individuals from the presumed hybrid population
to be genetically admixed, illustrating the potential of the markers to
identify interspecific hybridization. We expect the set of markers we
designed to provide a high resolving power for analysis of hybridization
in Triturus.
Triturus transcriptome-based gene models in FASTA formatTriturus
transcriptome-based gene models in FASTA
formatTriturus_transcriptome_contigs_sorted_to_length.fasExcel sheet and
macro to compose multiplexesExcel sheet and macro to compose
multiplexesExcel_multiplex_check.zipRaw Ion Torrent reads in FASTQ
formatRaw Ion Torrent reads in FASTQ formatFASTQ_IonTorrent.zipBWA
alignments in SAM formatBWA alignments in SAM formatSAM_alignments.zipRaw
SNP reports in VCF formatRaw SNP reports in VCF
formatVCF_raw_SNP_reports.zipFASTA files of reconstructed sequencesFASTA
files of reconstructed sequencesReconstructed_sequences_FASTA.zipInput
files for the BAPS analysis (in GENEPOP format), NewHybrids and
HIest.Input files for the BAPS analysis (in GENEPOP format), NewHybrids
and HIest.input_files.zipScript associated with the bioinformatics
pipelineScript associated with the bioinformatics pipeline. The first
script strings the programs together (used in Linux environment). The
second script determines the SNPs and allelic variants. The third script
creates the consensus sequences.Scripts.zip
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