Dye selection for live cell imaging of intact siRNA

Publication type:
Journal article
Metadata:
Autoren
  • Markus Hirsch
  • Dennis Strand
  • Mark Helm
Autoren-URL
https://www.webofscience.com/api/gateway?GWVersion=2&SrcApp=fis-test-1&SrcAuth=WosAPI&KeyUT=WOS:000300741400003&DestLinkType=FullRecord&DestApp=WOS_CPL
DOI
10.1515/BC-2011-256
eISSN
1437-4315
Externe Identifier
  • Clarivate Analytics Document Solution ID: 898KN
  • PubMed Identifier: 22628296
ISSN
1431-6730
Ausgabe der Veröffentlichung
1-2
Zeitschrift
BIOLOGICAL CHEMISTRY
Schlüsselwörter
  • confocal fluorescence microscopy
  • fluorescence resonance energy transfer (FRET)
  • RNA interference (RNAi)
  • siRNA degradation
  • siRNA integrity
  • small interfering RNA (siRNA)
Paginierung
23 - 35
Datum der Veröffentlichung
2012
Status
Published
Titel
Dye selection for live cell imaging of intact siRNA
Sub types
  • Article
Ausgabe der Zeitschrift
393

Data source: Web of Science (Lite)

Other metadata sources:
Abstract
<jats:title>Abstract</jats:title> <jats:p>Investigations into the fate of small interfering RNA (siRNA) after transfection may unravel new ways to improve RNA interference (RNAi) efficiency. Because intracellular degradation of RNA may prevent reliable observation of fluorescence-labeled siRNA, new tools for fluorescence microscopy are warranted to cover the considerable duration of the RNAi effect. Here, the characterization and application of new fluorescence resonance energy transfer (FRET) dye pairs for sensing the integrity of duplex siRNA is reported, which allows an assessment of the degradation status of an siRNA cell population by live cell imaging. A panel of high-yield fluorescent dyes has been investigated for their suitability as FRET pairs for the investigation of RNA inside the cell. Nine dyes in 13 FRET pairs were evaluated based on the performance in assays of photostability, cross-excitation, bleed-through, as well as on quantified changes of fluorescence as a consequence of, e.g., RNA strand hybridization and pH variation. The Atto488/Atto590 FRET pair has been applied to live cell imaging, and has revealed first aspects of unusual trafficking of intact siRNA. A time-lapse study showed highly dynamic movement of siRNA in large perinuclear structures. These and the resulting optimized FRET labeled siRNA are expected to have significant impact on future observations of labeled RNAs in living cells.</jats:p>
Autoren
  • Markus Hirsch
  • Dennis Strand
  • Mark Helm
DOI
10.1515/bc-2011-256
eISSN
1437-4315
ISSN
1431-6730
Ausgabe der Veröffentlichung
1-2
Zeitschrift
Biological Chemistry
Paginierung
23 - 35
Datum der Veröffentlichung
2012
Status
Published
Herausgeber
Walter de Gruyter GmbH
Herausgeber URL
http://dx.doi.org/10.1515/bc-2011-256
Datum der Datenerfassung
2021
Titel
Dye selection for live cell imaging of intact siRNA
Ausgabe der Zeitschrift
393

Data source: Crossref

Abstract
Investigations into the fate of small interfering RNA (siRNA) after transfection may unravel new ways to improve RNA interference (RNAi) efficiency. Because intracellular degradation of RNA may prevent reliable observation of fluorescence-labeled siRNA, new tools for fluorescence microscopy are warranted to cover the considerable duration of the RNAi effect. Here, the characterization and application of new fluorescence resonance energy transfer (FRET) dye pairs for sensing the integrity of duplex siRNA is reported, which allows an assessment of the degradation status of an siRNA cell population by live cell imaging. A panel of high-yield fluorescent dyes has been investigated for their suitability as FRET pairs for the investigation of RNA inside the cell. Nine dyes in 13 FRET pairs were evaluated based on the performance in assays of photostability, cross-excitation, bleed-through, as well as on quantified changes of fluorescence as a consequence of, e.g., RNA strand hybridization and pH variation. The Atto488/Atto590 FRET pair has been applied to live cell imaging, and has revealed first aspects of unusual trafficking of intact siRNA. A time-lapse study showed highly dynamic movement of siRNA in large perinuclear structures. These and the resulting optimized FRET labeled siRNA are expected to have significant impact on future observations of labeled RNAs in living cells.
Addresses
  • Institute of Pharmacy and Molecular Biotechnology, Heidelberg University, Im Neuenheimer Feld 364, D-69120 Heidelberg, Germany.
Autoren
  • Markus Hirsch
  • Dennis Strand
  • Mark Helm
DOI
10.1515/bc-2011-256
eISSN
1437-4315
Externe Identifier
  • PubMed Identifier: 22628296
Open access
false
ISSN
1431-6730
Ausgabe der Veröffentlichung
1-2
Zeitschrift
Biological chemistry
Schlüsselwörter
  • Brain
  • Cells, Cultured
  • Endothelial Cells
  • Animals
  • Rats
  • RNA, Small Interfering
  • Fluorescent Dyes
  • Microscopy, Confocal
  • Fluorescence Resonance Energy Transfer
  • Cell Survival
  • Hydrogen-Ion Concentration
Sprache
eng
Medium
Print
Paginierung
23 - 35
Datum der Veröffentlichung
2012
Status
Published
Datum der Datenerfassung
2012
Titel
Dye selection for live cell imaging of intact siRNA.
Sub types
  • Research Support, Non-U.S. Gov't
  • Journal Article
Ausgabe der Zeitschrift
393

Data source: Europe PubMed Central

Abstract
Investigations into the fate of small interfering RNA (siRNA) after transfection may unravel new ways to improve RNA interference (RNAi) efficiency. Because intracellular degradation of RNA may prevent reliable observation of fluorescence-labeled siRNA, new tools for fluorescence microscopy are warranted to cover the considerable duration of the RNAi effect. Here, the characterization and application of new fluorescence resonance energy transfer (FRET) dye pairs for sensing the integrity of duplex siRNA is reported, which allows an assessment of the degradation status of an siRNA cell population by live cell imaging. A panel of high-yield fluorescent dyes has been investigated for their suitability as FRET pairs for the investigation of RNA inside the cell. Nine dyes in 13 FRET pairs were evaluated based on the performance in assays of photostability, cross-excitation, bleed-through, as well as on quantified changes of fluorescence as a consequence of, e.g., RNA strand hybridization and pH variation. The Atto488/Atto590 FRET pair has been applied to live cell imaging, and has revealed first aspects of unusual trafficking of intact siRNA. A time-lapse study showed highly dynamic movement of siRNA in large perinuclear structures. These and the resulting optimized FRET labeled siRNA are expected to have significant impact on future observations of labeled RNAs in living cells.
Date of acceptance
2011
Autoren
  • Markus Hirsch
  • Dennis Strand
  • Mark Helm
Autoren-URL
https://www.ncbi.nlm.nih.gov/pubmed/22628296
DOI
10.1515/BC-2011-256
eISSN
1437-4315
Ausgabe der Veröffentlichung
1-2
Zeitschrift
Biol Chem
Schlüsselwörter
  • Animals
  • Brain
  • Cell Survival
  • Cells, Cultured
  • Endothelial Cells
  • Fluorescence Resonance Energy Transfer
  • Fluorescent Dyes
  • Hydrogen-Ion Concentration
  • Microscopy, Confocal
  • RNA, Small Interfering
  • Rats
Sprache
eng
Country
Germany
Paginierung
23 - 35
PII
/j/bchm.2012.393.issue-1-2/bc-2011-256/bc-2011-256.xml
Datum der Veröffentlichung
2012
Status
Published
Datum, an dem der Datensatz öffentlich gemacht wurde
2013
Titel
Dye selection for live cell imaging of intact siRNA.
Sub types
  • Journal Article
  • Research Support, Non-U.S. Gov't
Ausgabe der Zeitschrift
393

Data source: PubMed

Beziehungen:
Property of

Tools