Abstract
RNA aptamers can be genetically encoded in cells to probe and manipulate cellular function. The usefulness of aptamers in mammalian cells is limited by low accumulation and degradation by ribonucleases. Expression of circular RNA aptamers using the Tornado expression system achieves high stability and an abundance of intracellular RNA aptamers. With this method, RNA aptamers with otherwise minimal activity become potent inhibitors. Here, we describe protocols to characterize circular RNA aptamers expressed using Tornado. Included are methods to assess stability, abundance, subcellular localization, and target binding by circular RNA aptamers.