Additional file 7 of Sex-biased and parental allele-specific gene regulation by KDM6A
Creators: name: Ma, Wenxiu
name: Fang, He
name: Pease, Nicolas
name: Filippova, Galina N.
name: Disteche, Christine M.
name: Berletch, Joel B.
Publisher: name: figshare
Year: 2022
Subjects: subject: Genetics
subject: FOS: Biological sciences

description: Additional file 7: Fig. S7. Allelic epigenetic features of genes regulated by KDM6A. (A) Scatter plots of H3K27me3 read coverage in log2 scale on maternal and paternal alleles in male BC Kdm6aΔE1 cells versus wt. Allele-specific ChIP-seq reads around promoters (± 2 kb of the TSS) were used to calculate promoter coverage normalized by sequencing depth and allele differences (see also Fig. 5A, D). (B) Same analysis as in (A) but for DEGs that overlap between exonic KO clones only (Kdm6aΔE1 and Kdm6aΔE3) and Kdm6aΔY clones (Kdm6aΔE1, Kdm6aΔE3 and Kdm6aΔP4). (C) Scatter plots of ATAC-seq read coverage in log2 scale on maternal and paternal alleles in male BC Kdm6aΔE1 cells versus wt. Allele-specific reads around promoters (± 2 kb of the TSS) were used to calculate promoter coverage normalized by sequencing depth and allele differences (see also Fig. 5B, E). (D) Same analysis as in (C) but for DEGs that overlap between exonic KO clones only (Kdm6aΔE1 and Kdm6aΔE3) and Kdm6aΔY clones (Kdm6aΔE1, Kdm6aΔE3 and Kdm6aΔP4). (E, F) Same analysis as in (A) and (B), but for upregulated DEGs. (G, H) same analysis as in (C) and (D), but for upregulated DEGs. (I, J) Scatter plots of ratios of ATAC-seq read coverage versus ratios of H3K27me3 ChIP-seq read coverage between CB clone Kdm6aΔE2.5 and wt CB cells at promoter regions (± 2 kb of the TSS) of downregulated groups on maternal and paternal alleles. (K, L) Same analysis as in (I) and (J) but for upregulated groups. See also Fig. 5G, H. Unedited image figure legend. (A) Raw images for electrophoresis gels shown in Additional file 1A for BC and CB clones. Left panel shows screening of CRISPR exon deletion clones by PCR. The top right gel shows screening of CRISPR exon deletion clones in CB cells by PCR (gDNA). The associated control Actb lanes are in (C) lower right gel (gDNA). For all panels, primer names correspond to those in Additional file 1A. (B) Unedited images of western blots and controls for KDM6A. Top two are film images after 30 s and 5 min exposure (30 s image is shown in Additional file 1A). Lower image shows a picture of the Ponceau S stain used for loading control. For all images the clones are labelled at the top. (C) Raw gels for Kdm6a expression changes in exon deletion clones. Left panel shows Kdm6a expression changes in BC clones. The center panel shows a repeat electrophoresis run of the Actb controls which is shown in Additional file 1A. For CB clones, Kdm6a expression in controls and KO cells are shown in the lower right gel in (A) (cDNA). The right panel shows the Actb control (cDNA). Primer names correspond to those in Additional file 1A. (D) Raw images for electrophoresis gels shown in Additional file 1B for BC and CB clones. Left panel shows promoter deletion screening by PCR in BC clones (gDNA). The center and right panels show the same for the CB promoter clone (gDNA). For all panels, primer names correspond to those in Additional file 1B. (E) Raw gels for Kdm6a expression changes in control and promoter deletion clones. Top gel shows expression changes in BC KO clones and the lower gel shows expression changes in the CB KO clone. Primer names correspond to those in Additional file 1B. (F) Gel image of PCR screening for the off-target effect seen in CRISPR KO BC exon clones ΔE1, and ΔE3. Primer names correspond to those in Additional file 1F. (G) Gel images showing expression changes of Meg3 (left) and Rian (center) in controls and BC KO clones. The right panel shows the Actb controls.