DNA sustains a wide variety of damage, such as the formation of abasic sites, pyrimidine dimers, alkylation adducts, or oxidative lesions, upon exposure to UV radiation, alkylating agents, or oxidative conditions. Since such damage may be acutely toxic or mutagenic and potentially carcinogenic, it is of interest to gain insight into how their structures impact biochemical processing of DNA, such as synthesis, transcription, and repair. Lesion-specific molecular probes have been used to study polymerase-mediated translesion DNA synthesis of abasic sites and TT dimers, while other probes have been developed to specifically investigate the alkylation adduct O 6 -Bn-G and the oxidative lesion 8-oxo-G. In this review recent examples of lesion-specific molecular probes are surveyed; their specificities of incorporation opposite target lesions compared to unmodified nucleotides are discussed, and limitations of their applications under physiologically relevant conditions are assessed.Within years of the discovery that DNA 1 was responsible for transmitting hereditary information (1), and before publication of the double helical structure of DNA (2), researchers began to probe its susceptibility to various types of damage (3). Exposure of DNA to UV radiation, alkylating agents, and reactive oxidative species leads to the formation of abasic sites, pyrimidine dimers, alkylation adducts, and oxidative damage products. Each of these types of lesions can be mutagenic, sometimes carcinogenic, and this chemical process, therefore, is highly relevant to cellular biology and disease. For example, UV radiation damageinduced skin cancer was estimated to result in over one million new cases of the disease in the United States in 2008 (4). Further, DNA alkylation by tobacco-derived carcinogens contributes to lung cancer, which is estimated will result in about 160,000 deaths in the United States in 2009 (5,6). These examples of the biological consequences of DNA damage underscore the continued need for understanding the underlying molecular mechanisms.