Chromatin immunoprecipitation (ChIP) is a type of immunoprecipitation experimental technique used to investigate the interaction between proteins and DNA in the cell. A summary of the protocol for ChIP is here. Briefly, the steps are:
- DNA and associated proteins on chromatin in living cells or tissues are crosslinked;
- The DNA-protein complexes are sheared into ~500 bp DNA fragments by sonication or nuclease digestion;
- Cross-linked DNA fragments associated with the protein(s) of interest are selectively immunoprecipitated from the cell debris using an appropriate protein-specific antibody;
- The associated DNA fragments are purified and their sequence is determined.
After step 3 (immunoprecipitation of the protein-DNA complex), it is my understanding that the protein–DNA cross-link is reversed and proteins are removed by digestion with proteinase K.
Can anyone tell me if during this step the DNA at site of cross-linking is included in further sequencing step or is washed away with the protein itself?