ChIP on Chip: surprising results are often artifacts.
BACKGROUND
The method of chromatin immunoprecipitation combined with microarrays (ChIP-Chip) is a powerful tool for genome-wide analysis of protein binding. However, a high background signal is a common phenomenon.


RESULTS
Reinvestigation of the chromatin immunoprecipitation procedure led us to discover four causes of high background: i) non-unique sequences, ii) incomplete reversion of crosslinks, iii) retention of protein in spin-columns and iv) insufficient RNase treatment. The chromatin immunoprecipitation method was modified and applied to analyze genome-wide binding of SeqA and sigma(32) in Escherichia coli.


CONCLUSIONS
False positive findings originating from these shortcomings of the method could explain surprising and contradictory findings in published ChIP-Chip studies. We present a modified chromatin immunoprecipitation method greatly reducing the background signal.
Waldminghaus
Torsten
T
Department of Cell Biology, Institute for Cancer Research, The Norwegian Radium Hospital, Oslo University Hospital and University of Oslo, 0310 Oslo, Norway.
Skarstad
Kirsten
K
D016477:Artifacts; D001425:Bacterial Outer Membrane Proteins / Q000378:metabolism; D001483:Base Sequence; D047369:Chromatin Immunoprecipitation / Q000379:methods*; D002876:Chromosomes, Bacterial / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D029968:Escherichia coli Proteins / Q000378:metabolism; D005189:False Positive Reactions; D016680:Genome, Bacterial / Q000235:genetics; D006360:Heat-Shock Proteins / Q000378:metabolism; D040081:Protein Array Analysis / Q000379:methods*; D012260:Ribonucleases / Q000378:metabolism; D012808:Sigma Factor / Q000378:metabolism
D001425:Bacterial Outer Membrane Proteins; D004268:DNA-Binding Proteins; D029968:Escherichia coli Proteins; D006360:Heat-Shock Proteins; C089746:SeqA protein, E coli; D012808:Sigma Factor; C069774:heat-shock sigma factor 32; D012260:Ribonucleases