Skip to main content

Rapid Retrieval of DNA Target Sequences by Primer Extension Capture

  • Protocol
  • First Online:

Part of the book series: Methods in Molecular Biology ((MIMB,volume 772))

Abstract

There is a widespread need for methods to enrich DNA samples for sequences of interest prior to high-throughput sequencing and to reduce the costs associated with a shotgun approach. While useful for targeting megabase-sized regions in a few samples, hybridization capture approaches such as those using microarrays currently involve bulky handling steps, long incubation times, and high per-sample costs. In contrast, the primer extension capture (PEC) method allows direct selection of small genomic regions from DNA sources within 2 h, with low costs for use with parallel samples. PEC promises useful applications in studies such as ancient DNA or forensic sequencing, taxonomic surveying of metagenomic samples, or genomic mapping of repetitive elements.

This is a preview of subscription content, log in via an institution.

Buying options

Protocol
USD   49.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD   89.00
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever
Softcover Book
USD   119.99
Price excludes VAT (USA)
  • Compact, lightweight edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info
Hardcover Book
USD   169.99
Price excludes VAT (USA)
  • Durable hardcover edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info

Tax calculation will be finalised at checkout

Purchases are for personal use only

Learn about institutional subscriptions

Springer Nature is developing a new tool to find and evaluate Protocols. Learn more

References

  1. Briggs AW, Good JM, Green RE et al (2009) Targeted retrieval and analysis of five Neandertal mtDNA genomes. Science 325:318–321

    Article  PubMed  CAS  Google Scholar 

  2. Bentley DR (2006) Whole-genome re-­sequencing. Curr Opin Genet Dev 16:545–552

    Article  PubMed  CAS  Google Scholar 

  3. Margulies M, Egholm M, Altman WE et al (2005) Genome sequencing in microfabricated high-­density picolitre reactors. Nature 437:376–380

    PubMed  CAS  Google Scholar 

  4. Krause J, Briggs AW, Kircher M et al (2010) A complete mtDNA genome of an early modern human from Kostenki, Russia. Curr Biol 20:231–236

    Article  PubMed  CAS  Google Scholar 

  5. Gnirke A, Melnikov A, Maguire J et al (2009) Solution hybrid selection with ultra-long oligonucleotides for massively parallel targeted sequencing. Nat Biotechnol 27:182–189

    Article  PubMed  CAS  Google Scholar 

  6. Hodges E, Xuan Z, Balija V et al (2007) Genome-wide in situ exon capture for selective resequencing. Nat Genet 39:1522–1527

    Article  PubMed  CAS  Google Scholar 

  7. Rohland N, Hofreiter M (2007) Ancient DNA extraction from bones and teeth. Nat Protoc 2:1756–1762

    Article  PubMed  CAS  Google Scholar 

  8. Maricic T, Pääbo S (2009) Optimization of 454 sequencing library preparation from small amounts of DNA permits sequence determination of both DNA strands. Biotechniques 46:51–57

    Article  PubMed  CAS  Google Scholar 

  9. Briggs AW, Stenzel U, Meyer M et al (2010) Removal of deaminated cytosines and detection of in vivo methylation in ancient DNA. Nucleic Acids Res 38:e87

    Article  PubMed  Google Scholar 

  10. Meyer M, Briggs AW, Maricic T et al (2008) From micrograms to picograms: quantitative PCR reduces the material demands of ­high-throughput sequencing. Nucleic Acids Res 36:e5

    Article  PubMed  Google Scholar 

  11. Rozen S, Skaletsky H (2000) Primer3 on the WWW for general users and for biologist programmers. Methods Mol Biol 132:365–386

    PubMed  CAS  Google Scholar 

  12. Meyer M, Kircher M (2010) Illumina sequencing library preparation for highly multiplexed target capture and sequencing Cold Spring Harb Protoc doi:10.1101/pdb.prot5448

  13. Dapprich J, Ferriola D, Magira EE et al (2008) SNP-specific extraction of haplotype-resolved targeted genomic regions. Nucleic Acids Res 36:e94

    Article  Google Scholar 

  14. Meyer M, Stenzel U, Hofreiter M (2008) Parallel tagged sequencing on the 454 platform. Nat Protoc 3:267–278

    Article  PubMed  CAS  Google Scholar 

Download references

Acknowledgments

I thank Matthias Meyer for laboratory advice, Svante Pääbo for support and supervision, and the whole Neanderthal genome group for discussions. This work was supported by the Max Planck Society.

Author information

Authors and Affiliations

Authors

Corresponding author

Correspondence to Adrian W. Briggs .

Editor information

Editors and Affiliations

Rights and permissions

Reprints and permissions

Copyright information

© 2012 Springer Science+Business Media, LLC

About this protocol

Cite this protocol

Briggs, A.W. (2012). Rapid Retrieval of DNA Target Sequences by Primer Extension Capture. In: Orgogozo, V., Rockman, M. (eds) Molecular Methods for Evolutionary Genetics. Methods in Molecular Biology, vol 772. Humana Press. https://doi.org/10.1007/978-1-61779-228-1_8

Download citation

  • DOI: https://doi.org/10.1007/978-1-61779-228-1_8

  • Published:

  • Publisher Name: Humana Press

  • Print ISBN: 978-1-61779-227-4

  • Online ISBN: 978-1-61779-228-1

  • eBook Packages: Springer Protocols

Publish with us

Policies and ethics