918博天堂

A. Rapid increase of histone acetylation in silent genes caused by HDAC inhibitor treatment.ChIP assays were performed using H3K9ac and H4K16ac antibodies with chromatin from cells treated in the absence or presence of HDAC inhibitors. The ChIP DNA was analyzed using qPCR.

B. The ChIP-Seq signals for the DIPA gene (active) and FOSL1 gene (silent) were displayed.The signals highlighted in blue are not statistically significant as determined using peak-finding programs.

(Wang Z, Zang C, et al. Genome-wide mapping of HATs and HDACs reveals distinct functions in active and inactive genes [J]. Cell. 2009 Sep 4;138(5):1019-31.)

The EMSA results of binding of AlgR to the rsmX/Y/Z promoter sequence.

(Li M, Yan J, Yan Y. The Pseudomonas transcriptional regulator AlgR controls LipA expression via the noncoding RNA RsmZ in Pseudomonas protegens Pf-5 [J]. Biochem Biophys Res Commun. 2017 May 20;487(1):173-180.)

A. Methylation specific PCR of CASP8, MASPIN, TMS1 and MDR1 in prostate cancer cells. DNA from prostate cancer cells were Bisulfite modified using EZ methylation kit (Zymo Research) and then PCR done using methylation specific primers and unmethylation specific primers. Because in the cells sometimes methylated and unmethylated both copies are present, so depending upon the number copies they get amplified.

B. Methylation specific PCR of CASP8, MASPIN, TMS1 and MDR1 in 5-aza-dC (5?M/48hrs) treated prostate cancer cells.

C. Bisulfite sequencing chromatogram of TMS1 gene showing methylation in LNCaP, DU145 and PC3, partial methylation in PC3 and Unmethylation in RWPE1.

D. Diagrammatic representation of methylation pattern of TMS1 and MDR1 gene at different promoter positions in RWPE1, LNCaP, DU145 and PC3. Dark circles (●) represents methylated sites, shadowed circles (An external file that holds a picture, illustration, etc.Object name is nihms159165ig1.jpg) represent partially methylated sites and blank circles (○) represents unmethylated sites in TMS1 and MDR1 promoter region.

(Mishra DK, Chen Z, et al. Global methylation pattern of genes in androgen-sensitive and androgen-independent prostate cancer cells [J]. Mol Cancer Ther. 2010 Jan;9(1):33-45.)

Rad23b and Ddit3 methylation determined by BSP. Bisulfite treated DNA was amplified by PCR with BSP primers. PCR products were cloned into the pUC57 vector, and five clones selected and sequenced from each sample. Methylation level was defined as the ratio of methylated CpG sites in all clones.

A. Typical sequencing results;

B. early effects, tissues were collected 2 h postirradiation;

C. delay effects, 1 month postirradiation. *P,0.05 versus control.

(Wang J, Zhang Y, et al. Genome-wide screen of DNA methylation changes induced by low dose X-ray radiation in mice [J]. PLoS One. 2014 Mar 10;9(3):e90804.)

Examples of the liquid chemiluminescent DNA pull-down assay. (A–C) Light emission was measured from the liquid chemiluminescent DNA pull-down assay to indicate binding. The histograms indicate relative light emission. Data are shown as means ± x001D. (A) Effect of polychlorinated biphenyl (PCB) on thyroid hormone receptor β1 (TRβ1)-thyroid hormone response element (TRE) binding. Glutathione S-transferase (GST)-TRβ1 was incubated with digoxigenin (DIG)-labeled TRE with or without T3 and/or PCB. (B) Effect of PCB on steroid receptor coactivator-1 (SRC-1)-TRβ1-TRE binding. GST-SRC-1 was incubated with in vitro-translated TRβ1 and retinoid X receptor (RXR), then SRC-1-TRβ1/RXR complex was incubated with DIG-labeled TRE with or without T3 and/or PCB for 1 h at room temperature. (C) Effect of tamoxifen on silent mediator for retinoid and thyroid hormone (SMRT)-estrogen receptor α(ERα )-estrogen response element (ERE) binding. GST-SMRT was incubated with ERα, then SMRT-ERα complex was incubated with DIG-labeled ERE with or without 17β-estradiol (E2) and/or tamoxifen (TAM). After detection reaction, light emission was measured to indicate binding as described above. Data are shown as means ±SEM.

(Iwasaki T, Miyazaki W, et al. Liquid chemiluminescent DNA pull-down assay to measure nuclear receptor-DNA binding in solution [J]. Biotechniques. 2008 Oct;45(4):445-8.)

Description of the Y1H screens approach conducted with Zat12 promoter segments as baits.

(Ben Daniel BH, Cattan E, et al. Identification of novel transcriptional regulators of Zat12 using comprehensive yeast one-hybrid screens [J]. Physiol Plant. 2016 Aug;157(4):422-41.)

Figure. HiC-DC identifies identifies interactions associated with regulatory and structural elements at the sub-TAD level.

a. Raw Hi-C count matrix from the Rao et al.11 GM12878 data set for aB700 kb region including the BLC2 locus. Sub-TAD regions as called by Rao et al.11 are shown as blue squares.

b. Significant interactions (–log10 P values) for the same region as estimated by HiC-DC.

c. Epigenomic tracks for GM12878 for the same region, showing DNase I hypersensitive sites, H3K27ac, ChIP-seq for components of the cohesin complex, and Hi-C hotspots as estimated by HiC-DC.

d. Sashimi plot depiction of significant interactions called by HiC-DC, showing chromatin looping between hotspots associated with regulatory and structural elements.

(Carty M, Zamparo L, et al. An integrated model for detecting significant chromatin interactions from high-resolution Hi-C data [J]. Nat Commun. 2017 May 17;8:15454.)