Seeing genetic and epigenetic information without DNA denaturation using sequence-enabled reassembly (SEER)

Jason R. Porter, Sarah H. Lockwood, David J. Segal, Indraneel Ghosh

Research output: Chapter in Book/Report/Conference proceedingChapter

1 Scopus citations

Abstract

Virtually all methods for reading the sequence of bases in DNA rely on the ability to denature double-stranded DNA into single strands and then use Watson-Crick base-pairing rules to hybridize the strands with high specificity to another DNA primer or probe. However, nature frequently uses an alternative method, reading the sequence information directly from double-stranded DNA using sequence-specific DNA-binding proteins. Here we describe methods for the construction and testing of sequence probes based on engineered zinc finger DNA-binding proteins. Background is reduced using split-reporter molecules, and signal is amplified using enzymatic reporters. The resulting sequence-enabled reassembly (SEER) probes can be configured to detect DNA sequence (genetic) or DNA methylation (epigenetic) information.

Original languageEnglish (US)
Title of host publicationEngineered Zinc Finger Proteins
Subtitle of host publicationMethods and Protocols
EditorsJoel Mackay, David Segal
Pages365-382
Number of pages18
DOIs
StatePublished - Dec 1 2010

Publication series

NameMethods in Molecular Biology
Volume649
ISSN (Print)1064-3745

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Keywords

  • DNA sequence
  • Double-stranded DNA
  • beta-lactamase
  • diagnostics
  • engineered zinc finger proteins
  • fluorescent detection
  • green florescent protein
  • luciferase
  • methylation

ASJC Scopus subject areas

  • Molecular Biology
  • Genetics

Cite this

Porter, J. R., Lockwood, S. H., Segal, D. J., & Ghosh, I. (2010). Seeing genetic and epigenetic information without DNA denaturation using sequence-enabled reassembly (SEER). In J. Mackay, & D. Segal (Eds.), Engineered Zinc Finger Proteins: Methods and Protocols (pp. 365-382). (Methods in Molecular Biology; Vol. 649). https://doi.org/10.1007/978-1-60761-753-2_23