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Fluorophore optimization for FRET based ribozyme studies
Author(s) -
Zaborowicz Matthew,
Rohlman Christopher
Publication year - 2010
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.24.1_supplement.882.8
Subject(s) - ribozyme , fluorophore , förster resonance energy transfer , rna , substrate (aquarium) , rna splicing , intron , chemistry , biophysics , group ii intron , hairpin ribozyme , tetrahymena , fluorescence , biochemistry , biology , gene , physics , ecology , quantum mechanics
The focus of this research is fluorophore optimization for FRET analysis of group I introns. Group I introns are ribonucleic acids. These introns have the ability to splice themselves from the larger RNA to which they are bound and, once modified into ribozymes, can act as reaction catalysts. FRET is used to study the kinetics and folding of these Group I introns. Current protocols for the synthesis of fluorescently labeled substrates require a large excess of the fluorophore relative to the RNA substrate used in the ribozyme study. To develop an optimized reaction, a Tetrahymena ribozyme substrate was designed with both a fluorescein label and the ability to be modified with the fluorophore tetramethyl rhodamine. A study was conducted to determine the optimum conditions necessary to achieve a 90% yield and a functional FRET substrate. Results from this study are generally applicable to synthesis of FRET probes for RNA folding and catalysis. This research was supported by the Albion College Foundation for Undergraduate Research, Scholarly and Creative Activity.