Surveillance of siRNA integrity by FRET imaging.

Surveillance of siRNA integrity by FRET imaging. Research Abstract Details 

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Surveillance of siRNA integrity by FRET imaging. Abstract Text:

Anne ,Julius ,Il-Han Kim,Karl Rohr,Caroline MacLean,Gert Fricker,Ulrich Massing,Florian Eberle,Alexander Dalpke,Roger Fischer,Michael F Trendelenburg,Mark Helm,

Techniques for investigation of exogenous small interfering RNA (siRNA) after penetration of the cell are of substantial interest to the development of efficient transfection methods as well as to potential medical formulations of siRNA. A FRET-based visualization method including the commonplace dye labels fluorescein and tetramethylrhodamin (TMR) on opposing strands of siRNA was found compatible with RNA interference (RNAi). Investigation of spectral properties of three labelled siRNAs with differential FRET efficiencies in the cuvette, including pH dependence and FRET efficiency in lipophilic environments, identified the ratio of red and green fluorescence (R/G-ratio) as a sensitive parameter, which reliably identifies samples containing >90% un-degraded siRNA. Spectral imaging of siRNAs microinjected into cells showed emission spectra indistinguishable from those measured in the cuvette. These were used to establish a calibration curve for assessing the degradation state of siRNA in volume elements inside cells. An algorithm, applied to fluorescence images recorded in standard green and red fluorescence channels, produces R/G-ratio images of high spatial resolution, identifying volume elements in the cell with high populations of intact siRNA with high fidelity. To demonstrate the usefulness of this technique, the movement of intact siRNA molecules are observed after introduction into the cytosol by microinjection, standard transfection and lipofection with liposomes.

Surveillance of siRNA integrity by FRET imaging. Publishing Authors By Initials

A ,J ,IH Kim,K Rohr,C MacLean,G Fricker,U Massing,F Eberle,A Dalpke,R Fischer,MF Trendelenburg,M Helm,

For similar investigative techniques: genetic techniques: gene transfer techniques: transfection research abstracts see: investigative techniques: genetic techniques: gene transfer techniques: transfection research

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Surveillance of siRNA integrity by FRET imaging. Journal Published:

PUBLICATION TYPE: Research Support, Non-U.S. Gov

Journal: Nucleic acids research

VOLUME: 35

Page Numbers: e124

Journal Abbreviation: Nucleic Acids Res.

ISSN: 1362-4962

DAY: 22

MONTH: 09

YEAR: 2007

Surveillance of siRNA integrity by FRET imaging. Information

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LANGUAGE: eng

NlmUniqueID: 411011

Surveillance of siRNA integrity by FRET imaging. Keywords Mesh Terms:

KEYWORDS: Transfection

MESH TERMS: chemistry

Chemical & Substance for Abstract: Surveillance of siRNA integrity by FRET imaging. Information

Substance Name: tetramethylrhodamine

Registry Number: 62669-72-1

Grant and Affiliation Information for Surveillance of siRNA integrity by FRET imaging.

AFFILIATION: Department of Chemistry, Institute for Pharmacy and Molecular Biotechnology (IPMB), University of Heidelberg, Im Neuenheimer Feld 364, D-69120 Heidelberg, Germany.

Country: England

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MEDLINETA: Nucleic Acids Res

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