Cloning DNA Binding Proteins from cDNA Expression Libraries Using Oligonucleotide Binding Site Probes

Central to the regulation of transcription of eukaryotic genes is the interaction of specific DNA binding proteins with promoter and enhancer elements. The molecular cloning of such DNA binding factors is an important step toward understanding this process and a number of strategies have been devised to meet this aim. Oligonucleotide probes derived from peptide sequence data may be used to screen cDNA libraries by hybridization or, alternatively, cDNA expression libraries may be screened immunologically, using antibodies raised against the factor of interest. The protocol described here, which is based on a method described by Singh et al. (1 ), is analogous to immunological screening but uses a labeled DNA binding site oligonucleotide probe to screen a cDNA expression library; DNA binding factors are thus cloned by virtue of their binding activity.