ChIP-on-Chip

A ChIP -on-chip or ChIP - chip (of English. Chromatin Immunoprecipitation chip) is a biochemical method for the determination of protein-DNA interactions. The chip -on-chip is a combination of a chromatin immunoprecipitation and hybridisation with a DNA microarray ( DNA chip synonymously ). DNA -protein interactions occur in binding sequences for transcription factors, promoters, enhancers, repressors, silencers, insulators, as well as binding sequences for the DNA replication.

Properties

The purified recombinant protein is either mixed with the purified DNA (in vitro) or in vivo the recombinant protein is produced in a GMO and released after binding to the cellular DNA by cell disruption. Then, the protein and the binding of DNA to be reversibly cross-linked with formaldehyde. By sonication or by digestion with DNase from Micrococcus sp. the DNA is fragmented. Then immunoprecipitation of the crosslinked protein -DNA complexes is carried out with an antibody against the recombinant protein or protein tag. By heating, the DNA is released from the complexes, and hybridized to the microarray and colored. Finally, the sequences of the hybridized DNA can be determined by their position on the microarray.

One problem with the method is a lack of purity of the DNA -binding protein, which can lead to false -positive results of immunoprecipitation. Similarly, a high specificity of the antibody is necessary as heterophilic antibodies may also lead to false positive results. When microarray can be a lack of efficiency of hybridization lead to false negative results.

An alternative method is the chip Seq connecting the chromatin immunoprecipitation with the DNA sequencing in a high throughput. Systematic errors of the methods can be partially offset by a parallel use of both methods.