Products /
CUT&RUN Kits & Reagents /
CUT&RUN pAG-MNase and Spike-In DNA

CUT&RUN pAG-MNase and Spike-In DNA #40366

Name: CUT&RUN pAG-MNase and Spike-In DNA
Price: 320.00 USD
Availability: InStock

Filter:

Product Information

Product Usage Information

For the pAG-MNase Enzyme, after cell permeabilization and primary antibody binding, resuspend cells in 50 μl of digitonin buffer containing 1.5 μl of pAG-MNase Enzyme (33X dilution). Incubate cell samples with rotation at 4°C for 1 hour, wash cells with digitonin buffer, and then perform the chromatin digestion.

Sample Normalization Spike-In DNA can be added directly to the digestion stop buffer. For sample normalization with NG-seq, add 5 μl (50 pg) of Sample Normalization Spike-In DNA

to each reaction. When using 100,000 cells or 1mg of tissue per reaction this ensures that the normalization reads are around 0.5% of the total sequencing reads. If more or less than 100,000 cells or 1mg of tissue are used per reaction, proportionally scale the volume of Sample Normalization Spike-In DNA up or down to adjust normalization reads to around 0.5% of total reads.

When performing sample normalization, be sure to map the CUT&RUN sequencing data for all samples to both the test reference genome (e.g. human) and the sample normalization genome (S. cerevisiae).

Storage

pAG-MNase Enzyme is supplied in 10 mM Tris-HCl (pH 7.5), 150 mM NaCl, 1mM EDTA, 0.1mM PMSF and 50% glycerol. Do not aliquot the product. Sample Normalization Spike-In DNA is supplied in 10 mM Tris-HCl (pH 8.0). Store both products at –20°C. These products are stable for at least 12 months.

Protocol

Product Description

The CUT&RUN pAG-MNase and Spike-In DNA kit provides enough enzyme and spike-in DNA to support 50 CUT&RUN assays. The pAG-MNase Enzyme #57813 is a fusion of Protein A and Protein G to Micrococcal Nuclease, and is recombinantly produced in E. coli. The pAG-MNase is compatible with multiple species of antibodies, including both rabbit and mouse. The Sample Normalization Spike-In DNA (10 pg/µl) #29987 is fragmented genomic DNA from the yeast S. cerevisiae that can be added to a CUT&RUN reaction to facilitate normalization between samples during NG-seq analysis.

Background

Like the chromatin immunoprecipitation (ChIP) assay, Cleavage Under Targets and Release Using Nuclease (CUT&RUN) is a powerful and versatile technique used for probing protein-DNA interactions within the natural chromatin context of the cell (1-4). CUT&RUN provides a rapid, robust, and true low cell number assay for detection of protein-DNA interactions in the cell. Unlike the ChIP assay, CUT&RUN is free from formaldehyde cross-linking, chromatin fragmentation, and immunoprecipitation, making it a much faster and more efficient method for enriching protein-DNA interactions and identifying target genes. CUT&RUN can be performed in less than one day, from live cells to purified DNA, and has been shown to work with as few as 500-1,000 cells per assay (1,2). Instead of fragmenting all of the cellular chromatin as done in ChIP, CUT&RUN utilizes an antibody-targeted digestion of chromatin, resulting in much lower background signal than seen in the ChIP assay. As a result, CUT&RUN requires only 1/10th the sequencing depth that is required for ChIP-seq assays (1,2). Finally, the inclusion of simple spike-in control DNA allows for accurate quantification and normalization of target-protein binding that is not possible with the ChIP method. This provides for effective normalization of signal between samples and between experiments.

限制使用

除非 CST 的合法授书代表以书面形式书行明确同意，否书以下条款适用于 CST、其关书方或分书商提供的书品。任何书充本条款或与本条款不同的客书条款和条件，除非书 CST 的合法授书代表以书面形式书独接受，否书均被拒书，并且无效。

专品专有“专供研究使用”的专专或专似的专专声明，且未专得美国食品和专品管理局或其他外国或国内专管机专专专任何用途的批准、准专或专可。客专不得将任何专品用于任何专断或治专目的，或以任何不符合专专声明的方式使用专品。CST 专售或专可的专品提供专作专最专用专的客专，且专用于研专用途。将专品用于专断、专防或治专目的，或专专售（专独或作专专成）或其他商专目的而专专专品，均需要 CST 的专独专可。客专：(a) 不得专独或与其他材料专合向任何第三方出售、专可、出借、捐专或以其他方式专专或提供任何专品，或使用专品制造任何商专专品，(b) 不得复制、修改、逆向工程、反专专、反专专专品或以其他方式专专专专专品的基专专专或技专，或使用专品开专任何与 CST 的专品或服专专争的专品或服专， (c) 不得更改或专除专品上的任何商专、商品名称、徽专、专利或版专声明或专专，(d) 只能根据 CST 的专品专售条款和任何适用文档使用专品， (e) 专遵守客专与专品一起使用的任何第三方专品或服专的任何专可、服专条款或专似专专

For Research Use Only. Not for Use in Diagnostic Procedures.

Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc.

SimpleChIP is a registered trademark of Cell Signaling Technology, Inc.

XP is a registered trademark of Cell Signaling Technology, Inc.

All other trademarks are the property of their respective owners. Visit our Trademark Information page.