ChIP–seq and beyond: new and improved methodologies to detect and characterize protein–DNA interactions

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Abstract
Chromatin immunoprecipitation followed by sequencing (ChIP–seq) detects protein–DNA binding events and chemical modifications of histone proteins. Recent technological advances in the ChIP–seq protocol have enabled assaying samples with limited cells, increased precision of the genomic location of binding events, and assaying multiple binding events. However, technical and analytical challenges remain. Open chromatin assays — such as DNase–seq, formaldehyde-assisted identification of regulatory elements (FAIRE–seq) and DNaseI footprinting — offer complementary methods to identify genomic regions bound by regulatory proteins. Chromatin conformation capture (3C) and chromatin interaction analysis with paired-end tag sequencing (ChIA-PET) experiments detect three-dimensional chromatin interactions between bound proteins. Protein binding efficiency varies across sites within a single genome due to differences in the underlying genomic sequences and chromatin state. These differences affect the functionality of transcription factors within cells. SNPs in protein–DNA binding sites can affect binding efficiency across individuals and can be detected by allelic biases in sequences produced from high-throughput sequencing assays.