Substitution of the use of radioactivity by fluorescence for biochemical studies of RNA.
We present here the use of fluorescent methodologies for structural and functional studies of RNA in place of radioactivity. The methods are highly sensitive and quantitative with the use of an infrared fluorescence imaging system. IRD-700 and IRD-800 labels are used for fluorescence detection. Chemical probing methods are largely used for mapping RNA secondary structure and to monitor ligand interactions and conformational changes involving individual bases of RNA. The new fluorescent primer extension methodology allows simple and fast chemical probing of RNA with high sensitivity. IRD-700 and IRD-800 labeled primers can also be used to monitor protein-RNA interactions by fluorescent mobility shift assays. The speed and ease of these approaches are advantages over prior methods that used hazardous radioisotopes. Structural and biochemical investigations of RNA should benefit from the use of these fluorescent methodologies.
Ying
Bei-Wen
BW
Laboratoire de Chimie et Biologie Structurales, Gif-sur-Yvette, France.
Fourmy
Dominique
D
Yoshizawa
Satoko
S
D002232:Carbocyanines / Q000737:chemistry; D017931:DNA Primers / Q000737:chemistry*; D024202:Electrophoretic Mobility Shift Assay / Q000379:methods; D005453:Fluorescence; D005456:Fluorescent Dyes / Q000737:chemistry*; D007211:Indoles / Q000737:chemistry; D008024:Ligands; D009690:Nucleic Acid Conformation; D009943:Organophosphorus Compounds / Q000737:chemistry; D012313:RNA / Q000737:chemistry* / Q000378:metabolism; D012335:RNA, Ribosomal / Q000737:chemistry; D017423:Sequence Analysis, RNA
D002232:Carbocyanines; D017931:DNA Primers; D005456:Fluorescent Dyes; C525731:IRD 700; C427728:IRDye800; D007211:Indoles; D008024:Ligands; D009943:Organophosphorus Compounds; D012335:RNA, Ribosomal; D012313:RNA