UV-induced cross-linking of proteins to plasmid pBR322 containing 8-azidoadenine 2'-deoxyribonucleotides

An efficient method of cross-linking DNA to protein is described. The method is based on the incorporation of photoactive 8-azidoadenine 2'-deoxyribonucleotides into DNA. We have found that 8-N3dATP is a substrate for E. coli DNA polymerase I and that 8-N3dATP can be incorporated into plasmid pBR322 by nick-translation. Subsequently we were able to cross-link a set of different proteins to 8-azido-2'-deoxyadenosine-containing pBR322 by UV irradiation (366nm). No DNA-protein photocross-linking was observed under the same conditions when the non-photoactive plasmid pBR322 was used.