Revision 1
#9002
Store at 4C and -20C
877-616-CELL (2355)
877-678-TECH (8324)
3 Trask Lane | Danvers | Massachusetts | 01923 | USA
For Research Use Only. Not for Use in Diagnostic Procedures.
Applications:
ChIP
Storage
Specificity/Sensitivity
Description
Cells are fixed with formaldehyde and lysed, and chromatin is fragmented by partial digestion with Micrococcal Nuclease to obtain chromatin fragments of 1 to 5 nucleosomes. Enzymatic fragmentation of chromatin is much milder than sonication and eliminates problems resulting from variability in sonication power and emulsification of chromatin during sonication, which can result in incomplete fragmentation of chromatin or loss of antibody epitopes due to protein denaturation and degradation. Chromatin immunoprecipitations are performed using ChIP-validated antibodies and ChIP-Grade Protein G Agarose Beads. After reversal of protein-DNA cross-links, the DNA is purified using DNA purification spin columns, allowing for easy and efficient recovery of DNA and removal of protein contaminants without the need for phenol/chloroform extractions and ethanol precipitations. The enrichment of particular DNA sequences during immunoprecipitation can be analyzed by a variety of methods, including standard PCR and quantitative real-time PCR. Please note that this kit is not compatible with ChIP-seq because the ChIP-Grade Protein G Agarose Beads are blocked with sonicated salmon sperm DNA, which interferes with downstream sequencing.
The SimpleChIP® Kit also provides important controls to ensure a successful ChIP experiment. The kit contains a positive control Histone H3 Antibody, a negative control Normal Rabbit IgG Antibody and primer sets for PCR detection of the human and mouse ribosomal protein L30 (RPL30) genes. Histone H3 is a core component of chromatin and is bound to most DNA sequences throughout the genome, including the RPL30 locus. Thus, the Histone H3 Antibody provides a universal positive control that should enrich for almost any locus examined.
Background
Background References
- Orlando, V. (2000) Trends Biochem Sci 25, 99-104.
- Liu, Q. et al. (2000) Genes Dev 14, 1448-59.
- Zhao, H. and Piwnica-Worms, H. (2001) Mol Cell Biol 21, 4129-39.
- Kuo, M.H. and Allis, C.D. (1999) Methods 19, 425-33.
- Jiang, K. et al. (2003) J Biol Chem 278, 25207-17.
- Agalioti, T. et al. (2000) Cell 103, 667-78.
- Martin, S.A. and Ouchi, T. (2008) Mol Cancer Ther 7, 2509-16.
- Soutoglou, E. and Talianidis, I. (2002) Science 295, 1901-4.
- Chen, M.S. et al. (2003) Mol Cell Biol 23, 7488-97.
- Mikkelsen, T.S. et al. (2007) Nature 448, 553-60.
- Zeng, Y. et al. (1998) Nature 395, 507-10.
- Lee, T.I. et al. (2006) Cell 125, 301-13.
- Löffler, H. et al. (2006) Cell Cycle 5, 2543-7.
- Weinmann, A.S. and Farnham, P.J. (2002) Methods 26, 37-47.
- Zachos, G. et al. (2007) Dev Cell 12, 247-60.
- Wells, J. and Farnham, P.J. (2002) Methods 26, 48-56.
- Garber, K. (2005) J Natl Cancer Inst 97, 1026-8.
Species Reactivity
Species reactivity is determined by testing in at least one approved application (e.g., western blot).
Applications Key
ChIP: Chromatin IP
Cross-Reactivity Key
H: Human M: Mouse R: Rat Hm: Hamster Mk: Monkey Vir: Virus Mi: Mink C: Chicken Dm: D. melanogaster X: Xenopus Z: Zebrafish B: Bovine Dg: Dog Pg: Pig Sc: S. cerevisiae Ce: C. elegans Hr: Horse GP: Guinea Pig Rab: Rabbit G: Goat All: All Species Expected
Trademarks and Patents
Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc.
SimpleChIP is a registered trademark of Cell Signaling Technology, Inc.
All other trademarks are the property of their respective owners. Visit cellsignal.com/trademarks for more information.
Limited Uses
Except as otherwise expressly agreed in a writing signed by a legally authorized representative of CST, the following terms apply to Products provided by CST, its affiliates or its distributors. Any Customer's terms and conditions that are in addition to, or different from, those contained herein, unless separately accepted in writing by a legally authorized representative of CST, are rejected and are of no force or effect.
Products are labeled with For Research Use Only or a similar labeling statement and have not been approved, cleared, or licensed by the FDA or other regulatory foreign or domestic entity, for any purpose. Customer shall not use any Product for any diagnostic or therapeutic purpose, or otherwise in any manner that conflicts with its labeling statement. Products sold or licensed by CST are provided for Customer as the end-user and solely for research and development uses. Any use of Product for diagnostic, prophylactic or therapeutic purposes, or any purchase of Product for resale (alone or as a component) or other commercial purpose, requires a separate license from CST. Customer shall (a) not sell, license, loan, donate or otherwise transfer or make available any Product to any third party, whether alone or in combination with other materials, or use the Products to manufacture any commercial products, (b) not copy, modify, reverse engineer, decompile, disassemble or otherwise attempt to discover the underlying structure or technology of the Products, or use the Products for the purpose of developing any products or services that would compete with CST products or services, (c) not alter or remove from the Products any trademarks, trade names, logos, patent or copyright notices or markings, (d) use the Products solely in accordance with CST Product Terms of Sale and any applicable documentation, and (e) comply with any license, terms of service or similar agreement with respect to any third party products or services used by Customer in connection with the Products.
Revision 1