| Glowing locked nucleic acids: brightly fluorescent probes for detection of nucleic acids in cells. | |
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MedLine Citation:
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PMID: 20845923 Owner: NLM Status: MEDLINE |
Abstract/OtherAbstract:
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Fluorophore-modified oligonucleotides have found widespread use in genomics and enable detection of single-nucleotide polymorphisms, real-time monitoring of PCR, and imaging of mRNA in living cells. Hybridization probes modified with polarity-sensitive fluorophores and molecular beacons (MBs) are among the most popular approaches to produce hybridization-induced increases in fluorescence intensity for nucleic acid detection. In the present study, we demonstrate that the 2'-N-(pyren-1-yl)carbonyl-2'-amino locked nucleic acid (LNA) monomer X is a highly versatile building block for generation of efficient hybridization probes and quencher-free MBs. The hybridization and fluorescence properties of these Glowing LNA probes are efficiently modulated and optimized by changes in probe backbone chemistry and architecture. Correctly designed probes are shown to exhibit (a) high affinity toward RNA targets, (b) excellent mismatch discrimination, (c) high biostability, and (d) pronounced hybridization-induced increases in fluorescence intensity leading to formation of brightly fluorescent duplexes with unprecedented emission quantum yields (Φ(F) = 0.45-0.89) among pyrene-labeled oligonucleotides. Finally, specific binding between messenger RNA and multilabeled quencher-free MBs based on Glowing LNA monomers is demonstrated (a) using in vitro transcription assays and (b) by quantitative fluorometric assays and direct microscopic observation of probes bound to mRNA in its native form. These features render Glowing LNA as promising diagnostic probes for biomedical applications. |
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Authors:
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Michael E Østergaard; Pallavi Cheguru; Madhusudhan R Papasani; Rodney A Hill; Patrick J Hrdlicka |
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Publication Detail:
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Type: Journal Article; Research Support, U.S. Gov't, Non-P.H.S. |
Journal Detail:
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Title: Journal of the American Chemical Society Volume: 132 ISSN: 1520-5126 ISO Abbreviation: J. Am. Chem. Soc. Publication Date: 2010 Oct |
Date Detail:
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Created Date: 2010-10-07 Completed Date: 2011-01-20 Revised Date: - |
Medline Journal Info:
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Nlm Unique ID: 7503056 Medline TA: J Am Chem Soc Country: United States |
Other Details:
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Languages: eng Pagination: 14221-8 Citation Subset: IM |
Affiliation:
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Department of Chemistry, University of Idaho, Moscow, Idaho 83844-2343, USA. |
Export Citation:
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| MeSH Terms | |
Descriptor/Qualifier:
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Fluorescence Fluorescent Dyes* Nucleic Acid Hybridization Nucleic Acids / chemistry* Polymerase Chain Reaction |
| Chemical | |
Reg. No./Substance:
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0/Fluorescent Dyes; 0/Nucleic Acids |
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