Orthogonal RNA-regulated destabilization domains for three-color RNA imaging with minimal RNA perturbation

ABSTRACT RNA-regulated destabilization domain allows live-cell imaging of single RNA. However, this method is limited to single-color RNA imaging and its long RNA tag induces destabilization of the tagged RNA. Here, we describe two orthogonal RNA-regulated destabilization domains that enable three-color mRNA imaging in living mammalian cells. One of these destabilizations can be combined with a short RNA tag (9XMS2) for single-molecule RNA imaging without perturbation of RNA stability.