Provided are polynucleotides that find use, e.g., for detecting RNase activity. In certain embodiments, the polynucleotide comprises a tRNA operably linked to a pre-microRNA (pre-miRNA), and an aptamer that binds to a detectable label. According to some embodiments, the detectable label is a fluorescent dye, non-limiting examples of which include malachite green, tetramethylrosamine, sulforhodamine B, and a triphenylmethane dye. Also provided are compositions that include the polynucleotides of the present disclosure. Also provided are kits that include a polynucleotide of the present disclosure and a detectable label to which the aptamer of the polynucleotide binds.