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

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

Research output: Contribution to journalArticle

7 Citations (Scopus)

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

Fingerprint

Venus
Nucleic Acids
Nucleic acid sequences
Methylation
DNA
Chemical modification
DNA-Binding Proteins
Biosensors
Oligonucleotides
Polymers
Proteins
Biosensing Techniques

Keywords

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

ASJC Scopus subject areas

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

Cite this

Systematic evaluation of split-fluorescent proteins for the direct detection of native and methylated DNA. / Furman, Jennifer L.; Badran, Ahmed H.; Shen, Shengyi; Stains, Clifford I; Hannallah, Jack; Segal, David J.; Ghosh, Indraneel.

In: Bioorganic and Medicinal Chemistry Letters, Vol. 19, No. 14, 15.07.2009, p. 3748-3751.

Research output: Contribution to journalArticle

Furman, Jennifer L. ; Badran, Ahmed H. ; Shen, Shengyi ; Stains, Clifford I ; Hannallah, Jack ; Segal, David J. ; Ghosh, Indraneel. / Systematic evaluation of split-fluorescent proteins for the direct detection of native and methylated DNA. In: Bioorganic and Medicinal Chemistry Letters. 2009 ; Vol. 19, No. 14. pp. 3748-3751.
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