RNA G-quadruplex (G4) structures are thought to affect biological processes, including translation and pre-mRNA splicing, but it is not possible at present to demonstrate that they form naturally at specific sequences in long functional RNA molecules. We developed a new strategy, footprinting of long 7-deazaguanine-substituted RNAs (FOLDeR), that allows the formation of G4s to be confirmed in long RNAs and under functional conditions.