FOXA1/HNF3A CUTANA™ CUT&RUN Antibody

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Figure 2: FOXA1 transcription factor binding motif analysis in CUT&RUN
(A) Homer analysis determined that the FOXA1 consensus motif, represented as a sequence logo position weight matrix, was enriched under FOXA1 CUT&RUN peaks. (B) The number of FOXA1 peaks containing consensus motifs from panel A is represented by a Venn Diagram. (C) Gene browser shots were generated using the Integrative Genomics Viewer (IGV, Broad Institute). Three representative loci show overlap of FOXA1 peaks with consensus motifs.

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Figure 3: Western blot data
Western analysis of FOXA1 in whole cell lysates from MCF-7 and HeLa cells using NETN lysis buffer. The indicated amounts (µg) of lysate were loaded onto 4-20% SDS-PAGE gel and analyzed under standard western blot conditions using FOXA1 antibody (0.1 µg/mL).

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Figure 4: Immunoprecipitation of human FOXA1
EpiCypher FOXA1 antibody (6 µg) was used to immunoprecipitate whole cell lysates isolated from HeLa cells using NETN lysis buffer (1 mg per IP). A negative control IgG antibody and positive control antibody to a different FOXA1 epitope (Bethyl Laboratories) were also used to demonstrate specificity of the IP. Immunoprecipitates were loaded onto 4-20% SDS-PAGE gel (20% of IP loaded) and probed via western blot with EpiCypher FOXA1 antibody (1 µg/mL).

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Figure 5: CUT&RUN Methods
CUT&RUN was performed on 500k MCF7 cells with 0.5 µg of either FOXA1/HNF3A or IgG negative control (EpiCypher 13-0042) antibodies using the CUTANA™ ChIC/CUT&RUN Kit v2.0 (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. Libraries were run on an Illumina NextSeq2000 with paired-end sequencing (2x50 bp). Sample sequencing depth was 2.5 million reads (FOXA1) and 2.4 million reads (IgG). Data were aligned to the hg19 genome using Bowtie2. Data were filtered to remove duplicates, multi-aligned reads, and ENCODE DAC Exclusion List regions.