Description

SimpleChIP^? Human ID1 Promoter Primers contain a mix of forward and reverse PCR primers that are specific to a region of the human inhibitor of differentiation 1 (ID1) promoter. These primers can be used to amplify DNA that has been isolated using chromatin immunoprecipitation (ChIP). Primers have been optimized for use in SYBR^? Green quantitative real-time PCR and have been tested in conjunction with SimpleChIP^? Enzymatic Chromatin IP Kits.

Reactivity

Human

Background

The chromatin immunoprecipitation assay is a powerful and versatile technique used for probing protein-DNA interactions within the natural chromatin context of the cell. This assay can be used to either identify multiple proteins associated with a specific region of the genome or to identify the many regions of the genome bound by a particular protein. ChIP can be used to determine the specific order of recruitment of various proteins to a gene promoter or to "measure" the relative amount of a particular histone modification across an entire gene locus. In addition to histone proteins, the ChIP assay can be used to analyze binding of transcription factors and co-factors, DNA replication factors, and DNA repair proteins. When performing the ChIP assay, cells are first fixed with formaldehyde, a reversible protein-DNA cross-linking agent that "preserves" the protein-DNA interactions occurring in the cell. Cells are lysed and chromatin is harvested and fragmented using either sonication or enzymatic digestion. Fragmented chromatin is then immunoprecipitated with antibodies specific to a particular protein or histone modification. Any DNA sequences that are associated with the protein or histone modification of interest will co-precipitate as part of the cross-linked chromatin complex and the relative amount of that DNA sequence will be enriched by the immunoselection process. After immunoprecipitation, the protein-DNA cross-links are reversed and the DNA is purified. Standard PCR or quantitative real-time PCR are often used to measure the amount of enrichment of a particular DNA sequence by a protein-specific immunoprecipitation. Alternatively, the ChIP assay can be combined with genomic tiling micro-array techniques, high throughput sequencing, or cloning strategies, all of which allow for genome-wide analysis of protein-DNA interactions and histone modifications. SimpleChIP? primers have been optimized for amplification of ChIP-isolated DNA using real-time quantitative PCR and provide important positive and negative controls that can be used to confirm a successful ChIP experiment.

References

[1] Orlando, V. (2000) Trends Biochem Sci 25, 99-104.
[2] Kuo, M.H. and Allis, C.D. (1999) Methods 19, 425-33.
[3] Agalioti, T. et al. (2000) Cell 103, 667-78.
[4] Soutoglou, E. and Talianidis, I. (2002) Science 295, 1901-4.
[5] Mikkelsen, T.S. et al. (2007) Nature 448, 553-60.
[6] Lee, T.I. et al. (2006) Cell 125, 301-13.
[7] Weinmann, A.S. and Farnham, P.J. (2002) Methods 26, 37-47.
[8] Wells, J. and Farnham, P.J. (2002) Methods 26, 48-56.

Raw materials

Preparation Products