Flow cytometric detection of specific RNAs in native human cells with quenched autoligating FRET probes.

ABSTRACT: We describe the use of modified fluorescent-labeled oligonucleotide probes in the sequence-specific detection of messenger RNAs in live human cells. To make this detection possible, we developed a previously undescribed probe design that combines earlier quenched autoligation chemistry with a previously undescribed fluorescence resonance energy transfer (FRET) strategy to lower background signals. The probe pairs consisted of a nucleophilic 3'-phosphorothioate probe carrying a Cy5 FRET acceptor, and an electrophilic probe containing the combination of a 5' end electrophile/quencher and a fluorescein FRET donor. Probes were introduced to HL-60 cells by use of the streptolysin O pore-forming peptide. Signals from three different messenger RNAs, as well as 28S ribosomal RNA, could be detected

SUBMITTER: Abe H 

PROVIDER: S-EPMC1326184 | biostudies-literature | 2006 Jan

REPOSITORIES: biostudies-literature