Detection of the Excised, Damage-containing Oligonucleotide Products of Nucleotide Excision Repair in Human Cells.

Abstract

The human nucleotide excision repair system targets a wide variety of DNA adducts for removal from DNA, including photoproducts induced by UV wavelengths of sunlight. A key feature of nucleotide excision repair is its dual incision mechanism, which results in generation of a small, damage‐containing oligonucleotide approximately 24 to 32 nt in length. Detection of these excised oligonucleotides using cell‐free extracts and purified proteins with defined DNA substrates has provided a robust biochemical assay for excision repair activity in vitro. However, the relevance of a number of in vitro findings to excision repair in living cells in vivo has remained unresolved. Over the past few years, novel methods for detecting and isolating the excised oligonucleotide products of repair in vivo have therefore been developed. Here we provide a basic outline of a sensitive and versatile in vivo excision assay and discuss how the assay both confirms previous in vitro findings and offers a number of advantages over existing cell‐based DNA repair assays. Thus, the in vivo excision assay offers a powerful tool for readily monitoring the repair of DNA lesions induced by a large number of environmental carcinogens and anticancer compounds.