IN-VITRO SELECTION METHODOLOGIES TO PROBE RNA FUNCTION AND STRUCTURE

In vitro selection, or SELEX, has been used both to characterize the i nteraction of natural nucleic acids with proteins and to generate nove l nucleic acid-binding species, or aptamers. Although numerous reports have demonstrated the power of the technique, they have not expanded on the methodologies that can be used for selection. This review focus es on the considerations and problems involved in selecting protein-bi nding aptamers from a random-sequence RNA pool. As an illustration, we describe two approaches to selecting aptamers to a particular target, the HTLV-I Rex protein. In the first, complete randomization is used to find an artificial, high-affinity RNA binding site. In the second, the contributions of individual nucleotides and ! or base pairs to the natural Rex-binding element are determined by mutating the wild-type sequence and selecting active binding variants.