| Description | |
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| Catalogue Number | 17-375 |
| Brand Family | Upstate |
| Trade Name |
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| Description | EZ-Zyme™ Chromatin Prep Kit |
| Overview | Chromatin Immunoprecipitation (ChIP) is a widely utilized experimental technique to monitor the association of proteins with specific DNA sequences. This technique has been used to study patterns of histone modifications and to map binding sites of various proteins on a genome-wide scale with the aid of microarray technology. ChIP has also been used to determine the temporal regulation underlying the occupation of the particular chromatin locus by multiple proteins. In addition, because formaldehyde efficiently crosslinks protein to DNA as well as protein to protein, this method can, in principle, detect proteins that bind DNA directly as well as those that bind DNA indirectly through other factors. When performing ChIP, chromatin from cells and tissues needs to be fragmented so that it becomes soluble and resolution can be achieved in detecting protein-DNA interaction at specific loci. Sonication is a common method for producing sheared chromatin. Although this method is very efficient in releasing chromatin into the solution, it has a number of limitations. For example, foaming and overheating associated with sonication can result in protein denaturation or incomplete chromatin fragmentation. Also, sonication conditions can be physically harsh and can negatively impact immunoprecipitation efficiency. These and other limitations in the procedure can make sample sonication difficult to optimize and perform reproducibly. The EZ-Zyme™ Chromatin Prep kit allows ChIP analysis at nucleosome resolution by performing complete or partial digestions with a proprietary enzymatic cocktail to obtain chromatin fragments of on average one to a few nucleosomes in length. Compared to sonication, this procedure not only allows mapping the protein-DNA association at a higher resolution, but also renders the subsequent immunoprecipitation more efficient due to its mild digestion condition. In addition, enzymatic shearing is compatible with non-crosslinked ChIP (native ChIP), which allows analysis of histone modifications not only on cultured cells, but also on freshly dissected and frozen tissues. Use: The EZ-Zyme™ kit contains reagents optimized for generating enzymatically processed chromatin from mammalian cells for use in chromatin immunoprecipitation. While the kit contains a full complement of buffers required for chromatin shearing, careful attention must be paid to the details of the instructions. Follow all the instructions carefully, especially with regard to incubation times and temperatures. |
| Materials Required but Not Delivered | • Cells, stimulated or treated as needed for the experimental system • 37% Formaldehyde • Liquid nitrogen (N2 ) or dry ice/alcohol bath • Vortex mixer • Timer • Variable volume (2-1000 μL) pipettes + tips • Refrigerated Microcentrifuge • Variable temperature water bath • Cell scraper • Microfuge tubes, 1.5 mL • Filter-tip pipette tips |
| Product Information | |
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| Components |
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| Quality Level | MQ100 |
| Applications | |
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| Application | Contains proprietary reagents optimized for the enzymatic shearing of chromatin from mammalian cells at higher resolution than sonication for use in chromatin immunoprecipitation (ChIP) assays. |
| Product Usage Statements | |
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| Usage Statement |
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| Storage and Shipping Information | |
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| Storage Conditions | Upon receipt, store components at the temperatures indicated on the labels. Storage temperatures are also indicated on page 3 of this manual. Kit components are stable for 6 months from date of shipment when stored as directed. |
| Packaging Information | |
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| Material Size | 1 kit |
| Material Package | Sufficient reagents for 22 enzymatic chromatin preparations |