CUTANA™

CUTANA™ ChIC/CUT&RUN Kit

$945.00
SKU: 14-1048
Pack size: 48 Reactions

Description

The CUTANA™ ChIC/CUT&RUN Kit enables streamlined chromatin profiling of histone post-translational modifications (PTMs) and chromatin associated proteins.

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The CUT&RUN Kit Version 4 (v4) now uses SPRI magnetic beads for DNA purification instead of DNA spin columns, enabling multi-channel pipetting throughout for increased throughput and reproducibility. Positive (H3K4me3) and negative (IgG) control antibodies are included to pair with SNAP-CUTANA™ spike-in controls for assay optimization and continuous assay monitoring (Figure 2). E. coli DNA is included for data normalization. The kit is compatible with a variety of inputs including cells or nuclei derived from native, cryopreserved, or cross-linked samples. While it is recommended to start with 500,000 cells, comparable data can be generated using as few as 5,000 cells. The inclusion of controls, as well as compatibility with diverse target types, sample inputs, and low cell numbers, make this kit the go-to solution for chromatin mapping experiments. To place bulk orders, contact info@epicypher.com.

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Validation Data

Figure 1: CUT&RUN DNA fragment size distribution analysis
CUT&RUN was performed as described in Figure 5. Library DNA was analyzed by Agilent Tapestation®. This analysis confirmed that mononucleosomes were predominantly enriched in CUT&RUN (~300 bp peaks represent 150 bp nucleosomes + sequencing adapters).

Figure 2: SNAP-CUTANA™ K-MetStat Spike-in controls
DNA-barcoded designer nucleosomes (dNucs) representing 16 K-methyl PTMs: mono-, di-, and tri-methylation at H3K4, H3K9, H3K27, H3K36, and H4K20, as well as unmodified control, were spiked into CUT&RUN reactions prior to the addition of antibodies (IgG, H3K4me3). Instances of each spike-in barcode were counted and normalized from raw fastq files using the shell script and analysis excel sheet available on the spike-in product page (EpiCypher 19-1002). Barcodes for IgG (top; normalized to total reads) and H3K4me3 (bottom; normalized to on-target) antibodies are shown. The spike-ins confirmed optimal experimental conditions (H3K4me3 antibody specifically recovered the target dNuc, while IgG showed no preferential enrichment).

Figure 3: CUT&RUN genome-wide heatmaps
CUT&RUN was performed as described in Figure 5. Heatmaps show two replicates (“Rep”) of IgG (left) and H3K4me3 (right) kit control antibodies in aligned rows ranked by intensity (top to bottom) and colored such that red indicates high localized enrichment and blue denotes background signal.

Figure 4: Representative gene browser tracks
CUT&RUN was performed as described in Figure 5. A representative 174 kb window at the TRMT2A gene is shown for two replicates (“Rep”) of IgG and H3K4me3 kit control antibodies. Representative tracks are also shown for antibodies to H3K27me3 and the transcription factor CTCF. The CUT&RUN kit produced the expected genomic distribution for each target. Images were generated using the Integrative Genomics Viewer (IGV, Broad Institute).

Figure 5: CUT&RUN methods
CUT&RUN was performed using the CUTANA™ ChIC/CUT&RUN Kit starting with 500k K562 cells with 0.5 µg of either IgG (EpiCypher 13-0042), H3K4me3 (EpiCypher 13-0041), H3K27me3 (EpiCypher 13-0055), or 0.125 µg of CTCF (EpiCypher 13-2014) antibodies in duplicate. Library preparation was performed with 5 ng of DNA (or the total amount recovered if less than 5 ng) using the CUTANA™ Library Prep Kit (EpiCypher 14-1001/14-1002). Libraries were run on an Illumina NextSeq2000 with paired-end sequencing (2x50 bp). Sample sequencing depth was 3.5 million reads (IgG Rep 1), 3.8 million reads (IgG Rep 2), 4.7 million reads (H3K4me3 Rep 1), 6.9 million reads (H3K4me3 Rep 2), 6.6 million reads (H3K27me3 Rep 1), 4.7 million reads (H3K27me3 Rep 2), 3.9 million reads (CTCF Rep 1) and 4.6 million reads (CTCF Rep 2). Data were aligned to the hg19 genome using Bowtie2. Data were filtered to remove duplicates, multi-aligned reads, and ENCODE DAC Exclusion List regions.

Kit Contents

Item Cat. No.
CUTANA™ pAG-MNase for ChIC/CUT&RUN Workflows 15-1016
H3K4me3 Antibody, SNAP-Certified™ for CUT&RUN 13-0041k
CUTANA™ Rabbit IgG CUT&RUN Negative Control Antibody 13-0042k
SNAP-CUTANA™ K-MetStat Panel 19-1002k
CUTANA™ Concanavalin A Conjugated Paramagnetic Beads 21-1401
CUTANA™ E. coli Spike-in DNA 18-1401
CUTANA™ CUT&RUN 8-Strip 0.2 mL Tubes 10-0009a
Bead Activation Buffer 21-1001
Pre-Wash Buffer 21-1002
Stop Buffer 21-1003
5% Digitonin 21-1004k
1 M Spermidine 21-1005
0.5 M EDTA 21-1006
100 mM Calcium Chloride 21-1007
0.1X TE Buffer 21-1025
SPRIselect reagent manufactured by Beckman Coulter, Inc. 21-1405

Recommended Accessory Products

Item Cat. No.
CUTANA™ CUT&RUN Library Prep Kit 14-1001/14-1002
SNAP-CUTANA™ K-MetStat Panel 19-1002
CUT&RUN Antibodies See the list
Magnetic Separation Rack, 0.2 mL Tubes 10-0008
Magnetic Separation Rack, 1.5 mL Tubes 10-0012
CUTANA™ Nuclei Extraction Buffer 21-1026

Technical Information

Storage
OPEN KIT IMMEDIATELY and store components at room temperature, 4°C, and -20°C as indicated (see User Manual corresponding to Kit Version 4). Stable for 6 months upon date of receipt.
Instructions for Use
See User Manual corresponding to Kit Version 4. This kit is not compatible with previous user manuals. Version 4 contains SPRI magnetic beads for DNA clean up, while previous versions used DNA spin columns..

Additional Information

Beckman Coulter, the stylized logo, and SPRIselect are trademarks or registered trademarks of Beckman Coulter, Inc. in the United States and other countries.

Documents & Resources

Intellectual Property

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