MLL1/KMT2A CUTANA™ CUT&RUN Antibody

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Figure 2: MLL1 CUT&RUN representative browser tracks
CUT&RUN was performed as described in Figure 5. Gene browser shots were generated using the Integrative Genomics Viewer (IGV, Broad Institute). Two representative loci show overlap of MLL1 and H3K4me3 peaks.

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Figure 3: Immunoprecipitation data
EpiCypher MLL1 antibody (6 µL/mg lysate) was used to immunoprecipitate whole cell lysates (1 mg per IP reaction; 20% of IP loaded) isolated from HEK293T cells using NETN lysis buffer. A negative control IgG antibody and positive control antibodies targeting MLL1 (Bethyl Laboratories) were also used to demonstrate specificity of the IP. EpiCypher 13-2004, Bethyl A700-010-1, and Bethyl A300-086-6 target the same epitope, while Bethyl A300-087-3 targets a different epitope (between amino acids 1320 and 1380) For blotting immunoprecipitates, EpiCypher MLL1 antibody was used at a dilution of 1:1,000.

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Figure 4: Western blot data
Western analysis of MLL1 in whole cell extracts from HEK293T, MCF-7, A-549, and SW620 cells. Fifty micrograms of lysate was resolved via SDS-PAGE and detected with a 1:1,000 dilution of MLL1 antibody.

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Figure 5: CUT&RUN Methods
CUT&RUN was performed on 500k K562 cells with 0.5 µg of either MLL1, IgG negative control (EpiCypher 13-0042), or H3K4me3 positive control (EpiCypher 13-0041) antibodies using the CUTANA™ ChIC/CUT&RUN Kit v2.0 (EpiCypher 14-1048). Library preparation was performed with 5 ng 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. Libraries were run on an Illumina NextSeq2000 with paired-end sequencing (2x50 bp). Sample sequencing depth was 10.0 million reads (IgG), 13.5 million reads (MLL1), and 8.8 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.