Quenched autoligation probes.
Methods are described for preparation and use of quenched autoligation (QUAL) probes. These modified oligonucleotide fluorescent probes can be used to detect DNA and RNA in solution, on solid surfaces, and in fixed and living bacterial and human cells. They are quenched probes, and thus provide a "lighting up" signal in a single step, without removing unbound or unreacted probes from the analyte. QUAL probe signals can be detected by fluorescence spectrometer, fluorescence microscope, or flow cytometry. These probes can distinguish between very small variations, including single nucleotide differences, in nucleic acid targets. The described method includes a description of how to prepare the needed dabsyl quencher linker, how to prepare the QUAL probes by DNA synthesizer, and how to employ them in detecting nucleic acids in solution and in detecting RNAs in bacterial and human cells.
Silverman
Adam P
AP
Department of Chemistry, Stanford University, Stanford, CA, USA.
Abe
Hiroshi
H
Kool
Eric T
ET
D001483:Base Sequence; D002478:Cells, Cultured; D005434:Flow Cytometry / Q000379:methods*; D005456:Fluorescent Dyes / Q000737:chemistry*; D006090:Gram-Negative Bacteria / Q000235:genetics; D018922:HL-60 Cells; D006801:Humans; D008969:Molecular Sequence Data; D015345:Oligonucleotide Probes / Q000737:chemistry*; D012333:RNA, Messenger / Q000032:analysis* / Q000737:chemistry; D012335:RNA, Ribosomal / Q000032:analysis* / Q000737:chemistry
D005456:Fluorescent Dyes; D015345:Oligonucleotide Probes; D012333:RNA, Messenger; D012335:RNA, Ribosomal