Aptamers are short, single stranded nucleic acids which bind a wide range of different ligands with extraordinary high binding affinity and specificity. The steadily increasing number of aptamers is accompanied by an expanding range of applications in biotechnology. We will describe new developments in the field including the use of aptamers for conditional gene regulation and as biosensors. In addition, we will discuss the potential of aptamers as tags to visualize RNA and protein distribution in living cells and as therapeutics. Furthermore, we will consider biotechnological applications of riboswitches for gene regulation and as drug target.