DYKDDDDK Tag CUTANA™ CUT&RUN Antibody
55 in stock
Powered by BiozDYKDDDDK antibody is useful for studies utilizing FLAG®*-tagged target proteins. This antibody meets EpiCypher’s “CUTANA Compatible” criteria for performance in Cleavage Under Targets and Release Using Nuclease (CUT&RUN) and/or Cleavage Under Targets and Tagmentation (CUT&Tag) approaches to genomic mapping. Every lot of a CUTANA Compatible antibody is tested in the indicated approach using EpiCypher optimized protocols and determined to yield peaks that show a genomic distribution pattern consistent with reported function(s) of the target. DYKDDDDK Tag antibody produces CUT&RUN peaks (Figures 2-3) that overlap with GATA3 in breast cancer cells expressing 3xFLAG-tagged GATA3 transcription factor [1]**.
*FLAG® is a registered trademark of Merck KGaA, Darmstadt, Germany.
**Thanks to Dr. Takaku (UND) for 3xFLAG-GATA3-3xHA MDA-MB-231 cells.
Intellectual Property
Clone BLRE00G is provided for commercial sale under license from Bethyl Laboratories, Inc. with additional lot-specific validation in CUT&RUN performed by EpiCypher.
Background References:
[1] Takaku et al. Genome Biol. (2016). PMID: 26922637
Figure 1: Defined nucleosome spike-ins provide an in-assay control for DYKDDDDK Tag antibody in CUT&RUN
CUT&RUN was preformed as described in Figure 7. CUT&RUN sequencing reads were aligned to the unique DNA barcodes corresponding to each nucleosome in the SNAP-CUTANA™ DYKDDDDK Tag Panel (EpiCypher 19-5001). Data are expressed as a percent relative to on-target recovery (DYKDDDDK Tag set to 100%) or total counts (IgG/GATA3). IgG/GATA3 show no preferential binding to unmodified or DYKDDDDK spike-in nucleosomes. DYKDDDDK Tag antibody selectively enriches the DYKDDDDK Tag spike-in nucleosome, validating the antibody in CUT&RUN.
Figure 2: DYKDDDDK-tagged protein peaks in CUT&RUN
Figure 3: CUT&RUN representative browser tracks for DYKDDDDK-tagged protein
Figure 4: Western blot data
Figure 5: Immunoprecipitation data
Figure 6: Immunocytochemistry data
Figure 7: CUT&RUN methods
CUT&RUN was performed on 500k MDA-MB-231 native cells either stably overexpressing 3xFLAG-tagged GATA3 [1] or containing vector control. Either DYKDDDDK Tag (0.05 µg), H3K4me3 (0.5 µg; EpiCypher 13-0041), GATA3 (0.5 µg; CST 5852), or IgG (0.5 µg, EpiCypher 13-0042) antibodies were used with the CUTANA™ ChIC/CUT&RUN Kit v3 (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). Libraries were run on an Illumina NextSeq2000 with paired-end sequencing (2×50 bp). Sample sequencing depth (vector control cells/overexpressing cells) was 5.6/4.5 million reads (IgG), 7.8/6.7 million reads (H3K4me3), 5.8/4.7 million reads (GATA3), and 6.2/5.5 million reads (DYKDDDDK Tag). Data were aligned to the hg19 genome using Bowtie2. Data were filtered to remove duplicates, multi-aligned reads, and ENCODE DAC Exclusion List regions.
- Type: Monoclonal
- Host: Rabbit
- Applications: CUT&RUN, ELISA, WB, IP, ICC
- Reactivity: FLAG® Epitope (DYKDDDDK)
- Format: Purified
- Target Size: N/A
- CUT&RUN: 0.05 µg per reaction (0.5 µL of 100 µg/mL antibody)
- Enzyme Immunosorbent Assay (ELISA): 1:1,000 – 1:75,000; for coating plates 1:100 – 1:500
- Western Blot (WB): 1:1,000
- Immunoprecipitation (IP): 20 µL/mg lysate
- Immunocytochemistry (ICC): 1:1,000 – 1:5,000. Epitope retrieval with citrate buffer pH 6.0 is recommended for FFPE cell sections
