Phenylpyrrolocytosine as an unobtrusive base modification for monitoring activity and cellular trafficking of siRNA.

ABSTRACT

6-Phenylpyrrolocytosine (PhpC) is a cytosine mimic with excellent base-pairing fidelity, thermal stability, and high fluorescence. In this work, PhpC-containing small interfering RNAs (siRNAs) are shown to possess thermal stability and gene silencing activity that is virtually identical to that of natural siRNA. The emissive properties of PhpC allow the cellular trafficking of PhpC-containing siRNAs to be monitored by fluorescence microscopy. Accumulation in the cytoplasm of HeLa cells was observed using real time imaging. These findings demonstrate that PhpC-modified siRNAs retain the properties of natural siRNAs while allowing for fluorescence-based detection and monitoring, providing an ideal system for probing siRNA uptake and trafficking.