CUTANA™

CUTANA™ IgG Negative Control Antibody for CUT&RUN and CUT&Tag

$105.00
SKU: 13-0042
Pack Size: 100 µg
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  • Type:  Polyclonal
  • Host:  Rabbit
  • Applications:  CUT&RUN, CUT&Tag
  • Reactivity: Negative Control
  • Format:  Affinity Purified IgG
  • Target Size:  N/A

Description

Cleavage Under Targets & Release Using Nuclease (CUT&RUN) and Cleavage Under Targets and Tagmentation (CUT&Tag) are next generation assays for genomic mapping of chromatin features to replace Chromatin Immunoprecipitation (ChIP-seq) [Skene & Henikoff, 2017]. Unlike in ChIP-seq, where baseline signal is established by sequencing the fragmented input chromatin, reactions performed with this rabbit IgG antibody are used to determine background signal arising from non-specific MNase digestion and Tn5 tagmentation. Pair with the H3K4me3 positive control antibody (EpiCypher 13-0060) for a well-controlled experiment.

Validation Data - CUT&RUN

13-0042 Heat Map (Click to enlarge)

Figure 1: CUT&RUN genome wide enrichment
CUT&RUN was performed as described in Figure 3. Heatmaps show H3K4me3 peaks relative to IgG antibody in aligned rows ranked by intensity (top to bottom) and colored such that red indicates high localized enrichment and blue denotes background signal. H3K4me3 antibody showed expected enrichment around the TSS. Despite some observable background in the IgG control, differential enrichment with the target antibody is as expected.

13-0042 CUT&RUN Data (Click to enlarge)

Figure 2: Rabbit IgG CUT&RUN tracks
CUT&RUN was performed as described in Figure 3. Gene browser shots were generated using the Integrative Genomics Viewer (IGV, Broad Institute). Two gene loci show H3K4me3 peaks and minimal background in the IgG track, as expected.

13-0042-immunoprecipitation (Click to enlarge)

Figure 3: CUT&RUN methods
CUT&RUN was performed on 500k native K562 cells with 0.5 µg of either IgG or H3K4me3 (EpiCypher 13-0060) antibodies in duplicate using the CUTANA™ ChIC/CUT&RUN Kit v4 (EpiCypher 14-1048). Library preparation was performed with 5 ng of DNA (or the total amount recovered if less than 5 ng) using the CUTANA™ CUT&RUN Library Prep Kit (EpiCypher 14-1001/14-1002). Both kit protocols were adapted for high throughput Tecan liquid handling. Sample sequencing depth was 7.8/6.8 million reads (IgG Rep 1/Rep 2) and 12.6/10.9 million reads (H3K4me3 Rep 1/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.

Validation Data - CUT&Tag

13-0042 Heat Map (Click to enlarge)

Figure 4: CUT&Tag genome wide enrichment
CUT&Tag was performed as described in Figure 6. Heatmaps show H3K4me3 peaks relative to IgG antibody in aligned rows ranked by intensity (top to bottom) and colored such that red indicates high localized enrichment and blue denotes background signal. H3K4me3 antibody showed expected enrichment around the TSS, while the IgG antibody displayed minimal background, as expected.

13-0042 CUT&RUN Data (Click to enlarge)

Figure 5: Rabbit IgG CUT&Tag tracks
CUT&Tag was performed as described in Figure 6. Gene browser shots were generated using the Integrative Genomics Viewer (IGV, Broad Institute). Two gene loci show H3K4me3 peaks and minimal background in the IgG track, as expected.

13-0042-immunoprecipitation (Click to enlarge)

Figure 6: CUT&Tag methods
CUT&Tag was performed on 100k native K562 nuclei with 0.5 µg of either IgG or H3K4me3 (EpiCypher 13-0041) antibodies in singlicate using the CUTANA™ CUT&Tag Kit v1 (EpiCypher 14-1102/14-1103). Libraries were run on an Illumina NextSeq2000 with paired-end sequencing (2x50bp). Sample sequencing depth was 0.9 million reads (IgG) and 2.9 million reads (H3K4me3). Data were aligned to the hg19 genome using Bowtie2. Data were filtered to remove duplicates, multi-aligned reads, and ENCODE DAC Exclusion List regions.

Technical Information

Immunogen
None
Storage
Stable for 1 year at 4°C from date of receipt
Formulation
Affinity-purified antibody in PBS pH 7.6

Recommended Dilution

CUT&RUN
0.5 µg per reaction
CUT&Tag
0.5 µg per reaction

Documents & Resources

Technical Datasheet
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