A novel method for the genome-wide high resolution analysis of DNA damage

Abstract

DNA damage occurs via endogenous and exogenous genotoxic agents and compromises a genome's integrity. Knowing where damage occurs within a genome is crucial to understanding the repair mechanisms which protect this integrity. This paper describes a new development based on microarray technology which uses ultraviolet light induced DNA damage as a paradigm to determine the position and frequency of DNA damage and its subsequent repair throughout the entire yeast genome.

Figure 1.

The CPD distribution in a part of Chromosome 4. Black: CPD level detected by the CPD antibody. Error bars shown on the black line are 1 SD from the mean from the two individual experiments. Red: theoretical CPD distribution. For the genome representations here and in the supplementary figures: yellow boxes show ORF positions (arrows indicate direction of transcription), green boxes are ARSs and blue boxes are centromeres and telomeres. Grey dots show probe positions.

Figure 2.

An outline showing the detectable genomic region for a probe (not to scale). All detectable fragments are in blue.

Formula 1.

Calculation of CPD probe values. s = dipyrimidine site, n = number of dipyrimidine sites, f = mean fragment length, d = distance of dipyrimidine site from probe, P = probability of a CPD occurring at the dipyrimidine site (TT/AA = 0.68, TC/GA = 0.16, CT/AG = 0.13, CC/GG = 0.03).

Figure 3.

Scatter plot of CPD levels before repair (CPD 0 h) versus at 2 h repair (CPD 2 h). Red symbols: mitochondrial DNA (probes for all coding region in mitochondrial DNA where the GC content is closer to than in nuclear DNA); Grey symbols: nuclear DNA. Each data point is for a specific genomic probe.

Figure 4.

Relative CPD repair rate in a part of chromosome 4. Black: NER proficient cells; Red: NER deficient rad4 cells. Nomenclature for the genome is as per Figure 1.