Self-reporting hybridisation assay for miRNA analysis
Self-reporting hybridisation assay for miRNA analysis
Hybridisation assays, which are commonly used to analyse oligonucleotides such as siRNAs and miRNAs, often employ detection probes with fluorescent tags. The signal emitted by a fluorescent tag covers a broad range of wavelengths and this limits the multiplexing potential due to overlapping signals. A novel method of indirect oligonucleotide analysis has been developed which combines a hybridisation assay with cleavable small molecule mass tags using HPLC-ESI MS detection. A self-reporting detection probe has been designed which incorporates a DNA/RNA chimeric oligonucleotide sequence in the reporter region, which generates small nucleotide products upon RNase cleavage of the ribose-phosphate backbone. These small nucleotides can then serve as mass tags for the indirect detection of oligonucleotide analytes. The narrow mass range covered by a small molecule mass tag combined with the wide range of possible mass tags provides a high degree of multiplexing potential. This approach has been demonstrated for the analysis of a synthetic miRNA.
1088-1092
Riley, Jo-Anne
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Brown, Tom
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Gale, Nittaya
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Herniman, Julie
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Langley, G. John
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2014
Riley, Jo-Anne
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Brown, Tom
a64aae36-bb30-42df-88a2-11be394e8c89
Gale, Nittaya
eead6253-2431-407b-ab6b-92e35d41c3ef
Herniman, Julie
530b1a36-1386-4602-8df7-defa6eb3512b
Langley, G. John
7ac80d61-b91d-4261-ad17-255f94ea21ea
Riley, Jo-Anne, Brown, Tom, Gale, Nittaya, Herniman, Julie and Langley, G. John
(2014)
Self-reporting hybridisation assay for miRNA analysis.
Analyst, 139 (5), .
(doi:10.1039/C3AN01825C).
(PMID:24404560)
Abstract
Hybridisation assays, which are commonly used to analyse oligonucleotides such as siRNAs and miRNAs, often employ detection probes with fluorescent tags. The signal emitted by a fluorescent tag covers a broad range of wavelengths and this limits the multiplexing potential due to overlapping signals. A novel method of indirect oligonucleotide analysis has been developed which combines a hybridisation assay with cleavable small molecule mass tags using HPLC-ESI MS detection. A self-reporting detection probe has been designed which incorporates a DNA/RNA chimeric oligonucleotide sequence in the reporter region, which generates small nucleotide products upon RNase cleavage of the ribose-phosphate backbone. These small nucleotides can then serve as mass tags for the indirect detection of oligonucleotide analytes. The narrow mass range covered by a small molecule mass tag combined with the wide range of possible mass tags provides a high degree of multiplexing potential. This approach has been demonstrated for the analysis of a synthetic miRNA.
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Accepted/In Press date: 23 December 2013
e-pub ahead of print date: 2 January 2014
Published date: 2014
Organisations:
Characterisation and Analytics, Faculty of Natural and Environmental Sciences, Chemistry
Identifiers
Local EPrints ID: 362872
URI: http://eprints.soton.ac.uk/id/eprint/362872
ISSN: 0003-2654
PURE UUID: 81b15642-fb1a-48bd-b0da-494f4e007120
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Date deposited: 07 Mar 2014 13:53
Last modified: 15 Mar 2024 03:00
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Author:
Jo-Anne Riley
Author:
Nittaya Gale
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