Direct fluorescent monitoring of RNA levels

The same user who was looking for destabilized fluorescent proteins was also looking for ways to directly monitor RNA levels fluorescently. It turns out that two nice reviews on this subject have recently been published: [1] and [2]. Traditionally, this has been done by using RNA binding proteins that bind to a specific RNA sequence, as in the MS2 system. More recently, however, an RNA aptamer has been selected that binds an analog of the GFP chromophore. The chromophore is non-fluorescent in water, but becomes brightly fluorescent on binding to the aptamer [3]. The chromophore, DFHBI, is now commercially available, which means that this system should be pretty easy to use. I’ve not seen anyone use it yet to monitor RNA levels in cells, but I’ll be curious to see if it takes off. Interestingly, this is not the first fluorogenic RNA aptamer synthesized, but none of the others seem to have been that successful.

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  1. I’ve tried and tried to get this to work, Kurt… no luck. An 18x aptamer sequence in the RNA I was testing wasn’t enough to see any fluorescence on an epi scope. If anyone else wants to try the aptamer at UCSF, or anywhere else, I have the constructs.

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