[41] Selection of protein binding sites from random nucleic acid sequences

Abstract

This chapter describes the selection of protein binding sites from random nucleic acid sequences. A variety of strategies have been developed by which specific DNA or RNA sequences that can be bound by a protein, or by a protein complex, can be isolated by selecting these molecules in vitro from libraries of random sequences. In principle, these protocols make it possible to evaluate the capability of any protein to recognize specific nucleic acid sequences, thus providing information which can be invaluable for investigations of how that protein functions. These techniques have a number of experimental applications that can be useful for study of various proteins that have been implicated in oncogenesis. Either a biological or a biochemical selection scheme can be employed to isolate protein binding sites from a random sequence nucleic acid library, and each strategy can potentially yield different information. The power of current random sequence selection techniques derives from a central concept: that the selection procedure itself is coupled with a step in which the selected molecules are amplified by the polymerase chain reaction (PCR). The amplification step makes it possible to perform multiple sequential rounds of selection and amplification to isolate specific binding sequences.