We describe a fast and simple one-step affinity-purification method for the isolation of specific RNA-binding
proteins. An in vitro-transcribed hybrid RNA consisting of an aptamer sequence with high binding specificity to
the antibiotic streptomycin and a putative protein-binding RNA sequence is incubated with crude extract. After
complex formation, the sample is applied to an affinity column containing streptomycin immobilized to Sepharose.
The binding of the in vitro-assembled RNA-protein complex to streptomycin-Sepharose is mediated by the aptamer
RNA and the specifically bound proteins are recovered from the affinity matrix by elution with the antibiotic.
Employing two well-characterized RNA-protein interactions, we tested the performance of this new method. The
spliceosomal U1A protein and the bacteriophage MS2 coat protein could be isolated via their appropriate RNA
motif containing hybrid RNA from crude yeast extracts in high yield and purity after only one round of affinity
purification. As the purification principle is independent of the extract source, this new affinity chromatography
strategy that makes use of an in vitro-selected antibiotic-binding RNA as a tag, "StreptoTag," should be applicable
to extracts from other organisms as well. Therefore, we propose StreptoTag to be a versatile tool for the isolation
of unknown RNA-binding proteins.