Article ID | Journal | Published Year | Pages | File Type |
---|---|---|---|---|
10533573 | Analytical Biochemistry | 2005 | 12 Pages |
Abstract
The design of oligonucleotides for gene silencing requires a rational method for identifying hybridization-accessible sequences within the target RNA. To this end, we have developed stem-loop self-quenching reporter molecules (SQRMs) as probes for such sequence. SQRMs have a 5â² fluorophore, a quenching moiety on the 3â² end, an intervening sequence that forms an â¼5-basepaired stem, and a loop sequence of â¼20-30 bases. We have previously described a mapping strategy employing SQRMs to locate stem-loop structures in the target mRNA molecule. We now show that the original design constraint of a basepaired stem is not needed, either in vitro or in vivo. We propose that stemless probes possess sufficient signal-to-noise for use in vivo and that this ratio is an indication of hybridization of the probe to its target. Data showing that these SQRMs can specifically target and reduce c-Myb protein synthesis and can be used for real-time in vivo assays are presented.
Related Topics
Physical Sciences and Engineering
Chemistry
Analytical Chemistry
Authors
Lida K. Gifford, David Jordan, Vikram Pattanayak, Kathy Vernovsky, Bao T. Do, Alan M. Gewirtz, Ponzy Lu,