Triplex-Mediated Genome Targeting and Editing

  • Faisal Reza
  • Peter M. Glazer
Protocol
First Online:
Part of the Methods in Molecular Biology book series (MIMB, volume 1114)

Abstract

Genome targeting and editing in vitro and in vivo can be achieved through an interplay of exogenously introduced molecules and the induction of endogenous recombination machinery. The former includes a repertoire of sequence-specific binding molecules for targeted induction and appropriation of this machinery, such as by triplex-forming oligonucleotides (TFOs) or triplex-forming peptide nucleic acids (PNAs) and recombinagenic donor DNA, respectively. This versatile targeting and editing via recombination approach facilitates high-fidelity and low-off-target genome mutagenesis, repair, expression, and regulation.

Herein, we describe the current state-of-the-art in triplex-mediated genome targeting and editing with a perspective towards potential translational and therapeutic applications. We detail several materials and methods for the design, delivery, and use of triplex-forming and recombinagenic molecules for mediating and introducing specific, heritable, and safe genomic modifications. Furthermore we denote some guidelines for endogenous genome targeting and editing site identification and techniques to test targeting and editing efficiency.

Key words

Genome targeting Genome editing Recombination Triplex-forming oligonucleotide (TFO) Peptide nucleic acid (PNA) Recombinagenic donor DNA Molecular delivery 
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Notes

Acknowledgments

We gratefully acknowledge members of the Glazer Laboratory for helpful discussions. This work was supported by a National Institutes of Health (NIH) grant R01HL082655 and by a Doris Duke Innovations in Clinical Research Award (to P.M.G.). A National Institute of Diabetes and Digestive and Kidney Diseases Experimental and Human Pathobiology Postdoctoral Fellowship from NIH grant T32DK007556 also provided support (to F.R.). The authors declared no conflict of interest.

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Copyright information

© Springer Science+Business Media, LLC 2014

Authors and Affiliations

  • Faisal Reza
    • 1
  • Peter M. Glazer
    • 2
    • 3
  1. 1.Departments of Therapeutic RadiologyYale University School of MedicineNew HavenUSA
  2. 2.Department of Therapeutic RadiologyYale University School of MedicineNew HavenUSA
  3. 3.Department of GeneticsYale University School of MedicineNew HavenUSA

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