title: Detection of mRNA of the cyclin D1 breast cancer marker by a novel duplex-DNA probe.
abstract: Previously, we have described 5-((4-methoxy-phenyl)-trans-vinyl)-2'-deoxy-uridine, 6, as a fluorescent uridine analogue exhibiting a 3000-fold higher quantum yield (Φ 0.12) and maximum emission (478 nm) which is 170 nm red-shifted as compared to uridine. Here, we utilized 6 for preparation of labeled oligodeoxynucleotide (ODN) probes based on MS2 and cyclin D1 (a known breast cancer mRNA marker) sequences. Cyclin D1-derived labeled-ssODN showed a 9.5-fold decrease of quantum yield upon duplex formation. On the basis of this finding, we developed the ds-NIF (nucleoside with intrinsic fluorescence)-probe methodology for detection of cyclin D1 mRNA, by which the fluorescent probe is released upon recognition of target mRNA by the relatively dark NIF-duplex-probe. Indeed, we successfully detected, a ss-deoxynucleic acid (DNA) variant of cyclin D1 mRNA using a dark NIF-labeled duplex-probe, and monitoring the recognition process by fluorescence spectroscopy and gel electrophoresis. Furthermore, we successfully detected cyclin D1 mRNA in RNA extracted from cancerous human cells, using ds-NIF methodology.
authors.affiliation: Department of Chemistry, Bar-Ilan University, Ramat-Gan 52900, Israel.
authors.forename: Meirav
authors.initials: M
authors.lastname: Segal
authors.forename: Eylon
authors.initials: E
authors.lastname: Yavin
authors.forename: Pinhas
authors.initials: P
authors.lastname: Kafri
authors.forename: Yaron
authors.initials: Y
authors.lastname: Shav-Tal
authors.forename: Bilha
authors.initials: B
authors.lastname: Fischer
mesh_terms: D001483:Base Sequence; D014408:Biomarkers, Tumor / Q000235:genetics* / Q000378:metabolism*; D001943:Breast Neoplasms / Q000473:pathology*; D045744:Cell Line, Tumor; D019938:Cyclin D1 / Q000235:genetics* / Q000378:metabolism*; D015342:DNA Probes / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D015336:Molecular Probe Techniques; D009838:Oligodeoxyribonucleotides / Q000235:genetics / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D013050:Spectrometry, Fluorescence; D013696:Temperature
chemical_list: D014408:Biomarkers, Tumor; D015342:DNA Probes; D009838:Oligodeoxyribonucleotides; D012333:RNA, Messenger; D019938:Cyclin D1