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Next-generation Sequencing of 16S Ribosomal RNA Gene Amplicons
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Double-digest RAD sequencing using Ion Proton semiconductor platform (ddRADseq-ion) with nonmodel organisms.

Hans Recknagel1, Arne Jacobs1, Pawel Herzyk2,3

  • 1Institute of Biodiversity, Animal Health & Comparative Medicine, College of Medical, Veterinary & Life Sciences, University of Glasgow, Glasgow, G12 8QQ, UK.

Molecular Ecology Resources
|March 27, 2015
PubMed
Summary

Researchers developed a new protocol for double-digest RAD sequencing on Ion Torrent platforms. This method enables cost-effective generation of genetic markers for nonmodel organisms in evolutionary biology research.

Keywords:
Ion ProtonIon TorrentRAD sequencingdouble-digestgenotyping by sequencingsemiconductor sequencingsingle nucleotide polymorphism (SNP) genotyping

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Area of Science:

  • Evolutionary Biology
  • Genomics
  • Population Genetics

Background:

  • Traditional molecular markers are being replaced by high-throughput SNP genotyping for nonmodel organisms.
  • Restriction site associated DNA sequencing (RADseq) is a key SNP genotyping method, primarily used on Illumina platforms.

Purpose of the Study:

  • To develop and validate a protocol for double-digest RAD sequencing (ddRADseq) specifically for Ion Torrent semiconductor sequencing platforms.
  • To enable rapid and cost-effective generation of genetic markers for nonmodel species.

Main Methods:

  • Developed a ddRADseq protocol and adapters for Ion Torrent (Ion Proton, Ion PGM).
  • Sequenced thirteen genomic libraries from Arctic charr, European whitefish, and common lizard.
  • Processed sequencing data using the Stacks bioinformatic tool.

Main Results:

  • Generated ~962 million single-end reads across all libraries.
  • Obtained an average of ~11,000 polymorphic loci per library.
  • Validated the method through replication, phylogenetic reconstruction, and genetic crosses.

Conclusions:

  • The developed ddRADseq protocol is effective for Ion Torrent platforms.
  • This method provides a rapid, cost-effective, and reproducible way to generate genetic markers for nonmodel vertebrates.
  • Facilitates advanced population genetics and phylogenetic studies in diverse species.