Solid-phase detection of RNA using bispyrene-modified RNA probe.
Bispyrene modified 2'-O-methyl oligoRNA (OMUpy2), which is a useful fluorescent probe for specific RNA detection, was immobilized on a glass substrate via various linker molecules to develop a more convenient RNA detection chip for gene expression study. As the fluorescence intensity of OMUpy2 enhanced greatly when it hybridized with its complementary RNA, it was expected that RNA detection by OMUpy2-immobilized chip (RNA-chip) required neither fluorescent labels to target nucleic acid nor washing protocol after the hybridization. As the linker molecules, oligo(dT), agarose, and poly(ethylene glycol) (PEG) were chosen, and the fluorescence from the chip was measured by a fluorescence microscope. Results indicated that the RNA-chip was able to detect the complementary oligoribonucleotide (cORN) without target labeling and washing protocols. Furthermore, the background emission was reduced as the length of the linker increased. In the case of PEG linker, 25-fold enhancement of the fluorescence intensity of the OMUpy2 was observed upon the addition of cORN.
Sakamoto
Takashi
T
Department of Biomolecular Engineering, Kyoto Institute of Technology, Sakyo-ku, Kyoto 606-8585, Japan.
Kobori
Akio
A
Murakami
Akira
A
D005456:Fluorescent Dyes / Q000737:chemistry*; D008856:Microscopy, Fluorescence; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods*; D012313:RNA / Q000032:analysis* / Q000737:chemistry*; D015347:RNA Probes / Q000737:chemistry*
D005456:Fluorescent Dyes; D015347:RNA Probes; D012313:RNA