AP-2 binding to double-stranded DNA microarray probes

Copyright information:Taken from "Proof of concept for microarray-based detection of DNA-binding oncogenes in cell extracts"Nucleic Acids Research 2005;33(8):e79-e79.Published online 12 May 2005PMCID:PMC1110745.© The Author 2005. Published by Oxford University Press. All rights reserved () AP-2 DNA-binding specificity. Microarrays were spotted with probes in the form of 50 bp oligonucleotides (O58–O19, black columns) or of 120 bp PCR products (grey columns) generated as described in the legend to . Binding sites are located either in central (EcoRV site, E58–E19) or in terminal (HincII, H58–H19 and NotI, N58–N19) position on the 120 bp probes. Probe numbers indicate the profile score of the binding sites computed as in . Microarrays were incubated with purified GST-AP-2 fusion and bound proteins were detected with Cy3-labeled polyclonal antisera as described in Materials and methods. Binding activities determined from the Cy3 fluorescence were normalized to the Cy5 signal corresponding to the spotted DNA. The white column indicates background binding to a control PCR fragment into which no AP-2 site has been inserted. Values depict the average and SD of three data points. () Protein dose–response curves. Normalized binding activities were determined as above using the indicated amounts in nanograms of GST-AP-2 protein. High-affinity sequences (matrix values >33) are given as closed symbols (diamonds N58, triangles N38, squares N33), and low-affinity sequences (matrix values