Sensing of transcription factor binding via cyanine dye pair fluorescence lifetime changes.

ABSTRACT: We designed and synthesized sensors for imaging transcription factor-DNA interactions using a complementary pair of 21-base pair long oligonucleotides (ODNs) carrying two internucleoside phosphate-linked cyanine fluorophores that can either engage in Förster's resonance energy transfer (FRET) with fluorescence emission or assemble into a ground state quenched dimer with short fluorescence lifetimes (FL). Cyanine fluorophores were linked to ODNs within the NF-κB binding site. These sensors were tested in the presence of recombinant p50 and p65 NF-κB proteins or constitutively NF-κB activating HeLa cell lysates. By using a coherent light excitation source we followed changes in fluorescence lifetime of the donor (Cy5.5) at the donor's excitation and emission light wavelengths, as well as the

SUBMITTER: Bogdanov AA 

PROVIDER: S-EPMC3544408 | biostudies-literature | 2012 Aug

REPOSITORIES: biostudies-literature