Systematic evaluation of split-fluorescent proteins for the direct detection of native and methylated DNA

Jennifer L. Furman, Ahmed H. Badran, Shengyi Shen, Cliff I. Stains, Jack Hannallah, David Segal, Indraneel Ghosh

Research output: Contribution to journalArticlepeer-review

7 Scopus citations

Abstract

In order to directly detect nucleic acid polymers, we have designed biosensors comprising sequence-specific DNA binding proteins tethered to split-reporter proteins, which generate signal upon binding a predetermined nucleic acid target, in an approach termed SEquence-Enabled Reassembly (SEER). Herein we demonstrate that spectroscopically distinct split-fluorescent protein variants, GFPuv, EGFP, Venus, and mCherry, function effectively in the SEER system, providing sensitive DNA detection and the ability to simultaneously detect two target oligonucleotides. Additionally, a methylation-specific SEER-Venus system was generated, which was found to clearly distinguish between methylated versus non-methylated target DNA. These results will aid in refinement of the SEER system for the detection of user defined nucleic acid sequences and their chemical modifications as they relate to human disease.

Original languageEnglish (US)
Pages (from-to)3748-3751
Number of pages4
JournalBioorganic and Medicinal Chemistry Letters
Volume19
Issue number14
DOIs
StatePublished - Jul 15 2009

Keywords

  • Detection
  • DNA
  • GFP
  • Methylation
  • Sensor
  • Split-protein
  • Zinc-finger

ASJC Scopus subject areas

  • Pharmaceutical Science
  • Drug Discovery
  • Organic Chemistry
  • Molecular Medicine
  • Molecular Biology
  • Clinical Biochemistry
  • Biochemistry

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