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Ancient DNA pp 107-120 | Cite as

Generating RNA Baits for Capture-Based Enrichment

  • Noah Snyder-Mackler
  • Tawni Voyles
  • Jenny TungEmail author
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1963)

Abstract

Capture-based enrichment techniques have revolutionized genomic analysis of species and populations for which only low-quality or contaminated DNA samples (e.g., ancient DNA, noninvasively collected DNA, environmental DNA) are available. This chapter outlines an optimized laboratory protocol for generating RNA “baits” for genome-wide capture of target DNA from a larger pool of DNA. This method relies on the in vitro transcription of biotinylated RNA baits, which has the dual benefit of eliminating the high cost of synthesizing custom baits and producing a bait set that targets the majority of regions genome-wide. We provide a detailed protocol for the three main steps involved in bait library construction: (1) making a DNA library from a high-quality DNA sample for the organism of interest or a closely related species; (2) using duplex-specific nuclease digestion to reduce the representation of repetitive regions in the DNA library; and (3) performing in vitro transcription of the repetitive region-depleted DNA library to generate biotinylated RNA baits. Where applicable, we include notes and recommendations based on our own experiences.

Key words

Whole-genome capture Biotinylated RNA baits Targeted enrichment Repetitive regions Capture-based enrichment Genome resequencing 

Notes

Acknowledgments

This work was supported by National Science Foundation grants DEB-1405308 (to J.T.) and SMA-1306134 (to J.T. and N.S.M.). We thank Jacob Gordon, Amanda Shaver, and Michael Yuan for key contributions to the protocol design and optimization and Arielle Fogel and Jen Tinsman for comments on an earlier draft of this chapter.

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Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2019

Authors and Affiliations

  • Noah Snyder-Mackler
    • 1
    • 2
  • Tawni Voyles
    • 1
  • Jenny Tung
    • 1
    Email author
  1. 1.Department of Evolutionary AnthropologyDuke UniversityDurhamUSA
  2. 2.Department of PsychologyUniversity of WashingtonSeattleUSA

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