Abstract
Restriction-site Associated DNA (RAD) markers are rapidly becoming a standard for SNP discovery and genotyping studies even in organisms without a sequenced reference genome. It is difficult, however, to identify genes nearby RAD markers of interest or move from SNPs identified by RAD to a high-throughput genotyping assay. Paired-end sequencing of RAD fragments can alleviate these problems by generating a set of paired sequences that can be locally assembled into high-quality contigs up to 1 kb in length. These contigs can then be used for SNP identification, homology searching, or high-throughput assay primer design. In this chapter, we offer suggestions on how to design a RAD paired-end (RAD-PE) sequencing project and the protocol for creating paired-end RAD libraries suitable for Illumina sequencers.
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Acknowledgments
The authors thank the University of Oregon researchers who have helped troubleshoot preliminary versions of this protocol. The project described was supported by grant R21HG003834 from the National Human Genome Research Institute (E.A.J.).
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© 2012 Springer Science+Business Media New York
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Etter, P.D., Johnson, E. (2012). RAD Paired-End Sequencing for Local De Novo Assembly and SNP Discovery in Non-model Organisms. In: Pompanon, F., Bonin, A. (eds) Data Production and Analysis in Population Genomics. Methods in Molecular Biology, vol 888. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-61779-870-2_9
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DOI: https://doi.org/10.1007/978-1-61779-870-2_9
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Publisher Name: Humana Press, Totowa, NJ
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