INTRODUCTIONKnowledge of the DNA-binding specificity of a transcription factor aids in understanding the function of that factor in the regulation of gene transcription. One popular method of identifying the genomic DNA sites bound by a given protein in vivo is the Chromatin ImmunoPrecipitation with microarray analysis (ChIP-chip) technique. However, this method reveals a binding pattern influenced by in vivo phenomena that may mask the actual DNA-binding specificity of the factor, such as chromatin effects and competitive or cooperative protein-protein interactions. ChIP-chip also requires adequate expression of the protein in the cell type chosen to create the extract used for immunoprecipitation. DNA Immunoprecipitation (DIP) is an alternative technique that allows one to take advantage of the favorable properties of both in vivo and traditional in vitro techniques (e.g., electromobility shift assay [EMSA] and binding site selection [SELEX]). DIP utilizes naked genomic DNA as a binding substrate for one or more purified recombinant proteins. Because genomic DNA is used as a template in DIP reactions, the results are directly comparable to ChIP-chip or ChIP-seq data. DIP can be carried out in the presence of cofactors such as heterodimer partners, competitors, or small molecule binding inhibitors. After DNA is isolated by DIP, it is most efficiently detected using a highly parallel genomic technique such as a DNA microarray (DIP-chip) or high-throughput sequencing (DIP-seq). In this protocol, we describe a DIP of a yeast (Saccharomyces cerevisiae) protein with yeast genomic DNA.
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Evidence ID | Analyze ID | Gene/Complex | Systematic Name/Complex Accession | Qualifier | Gene Ontology Term ID | Gene Ontology Term | Aspect | Annotation Extension | Evidence | Method | Source | Assigned On | Reference |
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Evidence ID | Analyze ID | Gene | Gene Systematic Name | Phenotype | Experiment Type | Experiment Type Category | Mutant Information | Strain Background | Chemical | Details | Reference |
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Evidence ID | Analyze ID | Gene | Gene Systematic Name | Disease Ontology Term | Disease Ontology Term ID | Qualifier | Evidence | Method | Source | Assigned On | Reference |
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Evidence ID | Analyze ID | Regulator | Regulator Systematic Name | Target | Target Systematic Name | Direction | Regulation of | Happens During | Regulator Type | Direction | Regulation Of | Happens During | Method | Evidence | Strain Background | Reference |
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Site | Modification | Modifier | Source | Reference |
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Evidence ID | Analyze ID | Interactor | Interactor Systematic Name | Interactor | Interactor Systematic Name | Allele | Assay | Annotation | Action | Phenotype | SGA score | P-value | Source | Reference | Note |
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Evidence ID | Analyze ID | Interactor | Interactor Systematic Name | Interactor | Interactor Systematic Name | Assay | Annotation | Action | Modification | Source | Reference | Note |
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Complement ID | Locus ID | Gene | Species | Gene ID | Strain background | Direction | Details | Source | Reference |
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Evidence ID | Analyze ID | Dataset | Description | Keywords | Number of Conditions | Reference |
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