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Fluorescence-Based High-Throughput Screening of Dicer Cleavage Activity

Publikace na Přírodovědecká fakulta |
2014

Tento text není v aktuálním jazyce dostupný. Zobrazuje se verze "en".Abstrakt

Production of small RNAs by ribonuclease III Dicer is a key step in microRNA and RNA interference pathways, which employ Dicer-produced small RNAs as sequence-specific silencing guides. Further studies and manipulations of microRNA and RNA interference pathways would benefit from identification of small-molecule modulators.

Here, we report a study of a fluorescence-based in vitro Dicer cleavage assay, which was adapted for high-throughput screening. The kinetic assay can be performed under single-turnover conditions (35 nM substrate and 70 nM Dicer) in a small volume (5 mu L), which makes it suitable for high-throughput screening in a 1536-well format.

As a proof of principle, a small library of bioactive compounds was analyzed, demonstrating potential of the assay.