title: 7_wo_B6_males_pool2

city: Stony Brook

first: Leonard

middle: YM

last: Cheung

state: New York

address: 101 Nicolls Road

institute: Stony Brook University

description: single-cell RNAseq on 10x Genomics Chromium platform of 3 (D,E,F) 7-week-old C57BL/6 male mice as pool2

data_processing:
Illumina BCL file unpacked and demultiplexed using 10x Genomics Cellranger mkfastq software.
Fastq files analyzed by self-contained Cellranger count software from 10x Genomics.  Cellranger performs barcode processing, alignment to mm10 transcriptome using STAR, transcript counting, PCA analyses, tSNE, and k-means clustering to generate all processed data.
Genome_build: modified UCSC mm10 with additional gene "Prop1L" added to gtf file at chromosome 11: 50,948,572-50,949,192
Supplementary_files_format_and_content: matrix.mtx: tab-separated file with transcript counts for each gene in each cell.  It uses the row number from genes.tsv as the gene number in column 1, and the row number from barcodes.tsv as the cell number in column 2.  Column 3 is the count.
Supplementary_files_format_and_content: genes.tsv: tsv file with ENSEMBL ID and gene names
Supplementary_files_format_and_content: barcodes.tsv: tsv file with valid barcodes of the total 13663 cells from each pool aggregated together
Supplementary_files_format_and_content: projection.csv: csv file with calculations for tSNE analyses for 2 components for each barcode (i.e. cell) in experiment.  Used to draw tSNE plots described in article.

molecule: polyA RNA

source_name: 3 pituitary glands

extract_protocol:
The pituitary glands of six individually-housed 7-week-old male C57BL/6 mice were dissected, and the tissue dispersed as previously described14-16, as two pools of three glands each.  Briefly, the dissected tissue is placed into an enzyme mix containing 0.5% w/v collagenase type 2 (Lorne Laboratories, UK), 0.1x trypsin (Invitrogen), 50 mg/ml DNase I (Worthington Biochemical, USA), and 2.5 mg/ml Fungizone (Invitrogen) in Hank’s balanced salt solution (HBSS) (Invitrogen) for 4 hrs at 37°C.  The tissue and cells are gently triturated, counted on a hemocytometer, pelleted at 200g for 5 min, and resuspended at ~1000 cells per ml in 0.4% bovine serum albumin (BSA)/HBSS.
Single-cell 3’ library generation was performed on the 10x Genomics Chromium Controller following the manufacturers protocol for the v2 reagent kit (10X Genomics, Pleasanton, CA, USA)10.  Cell suspensions were loaded onto a Chromium Single-Cell A chip along with reverse transcription (RT) master mix and single cell 3’ gel beads, aiming for 10,000 cells per channel.  For each pool, 17,400 cells with 71-82 % viability were encapsulated into emulsion droplets at a concentration of 700-1200 cells/ul, which targets 10,000 single cells with an expected multiple cell rate rate of 7.6 %.  Following generation of single-cell gel bead-in-emulsions (GEMs), reverse transcription was performed and the resulting post GEM-RT product was cleaned up using DynaBeads MyOne Silane beads (ThermoFisher Scientific, Waltham, MA, USA).  The cDNA was amplified, SPRIselect (Beckman Coulter, Brea, CA, USA) cleaned and quantified then enzymatically fragmented and size selected using SPRIselect beads to optimize the cDNA amplicon size prior to library construction.  An additional round of double-sided SPRI bead cleanup is performed after end repair and A-tailing.  Another single-sided cleanup is done after adapter ligation.  Indexes were added during PCR amplification and a final double-sided SPRI cleanup was performed.  Libraries were quantified by Kapa qPCR for Illumina Adapters (Roche) and size was determined by Agilent tapestation D1000 tapes.  Read 1 primer sequence were added to the molecules during GEM incubation.  P5, P7 and sample index and read 2 primer sequence were added during library construction via end repair, A-tailing, adaptor ligation and PCR.  Libraries were generated with unique sample indices (SI) for each sample. Libraries were sequenced on a HiSeq 4000 (Illumina, San Diego, CA, USA), using a HiSeq 4000 PE Cluster Kit with HiSeq 4000 SBS Kit (100 cycles, reagents, loaded at 200 pM) following Illumina’s denaturing and dilution recommendations.  PhiX was spiked into the load at 1%.  The run configuration was 26 x 8 x 98 (115) cycles for Read 1, Index and Read 2, respectively.  Cell Ranger Single Cell Software Suite 2.1.1 was used to perform sample demultiplexing and barcode processing at the University of Michigan Biomedical Core Facilities DNA Sequencing Core.

value: pituitary gland

tag: strain

tag: age

tag: gender

value: male


title: Strain 904 (WT) S1

status: Public on Oct 19 2018

type: SRA

city: Cleveland

first: Hee-Sook

last: Kim

state: OH

address: 2399 Euclid Ave

country: USA

institute: Cleveland State University

description:
processed data file:
BP_MFAseq_sliding window.txt
2_wt_S1

library_source: genomic

data_processing:
sequence reads were quality trimmed using with parameter,  trim_galore --phred33 -q 20 -a GATCGGAAGAGCACACGTCTGAACTCCAGTCAC --stringency 5 --length 20 --e 0.1
bowtie aligned to the reference genome (Tb927v5 from the TritrypDB) with -p 8 -a --best -M 1 -v 2
Aligned reads were quantified in Seqmonk and RPKM or RPM values were generated
For VSG and all genes, Raw reads were obtained from Seqmonk, then analyzed with DEseq.
Genome_build: Tb927v5 (http://tritrypdb.org/tritrypdb); VSGnome (http://tryps.rockefeller.edu); 14 BES fasta [PMID 18953401]
Supplementary_files_format_and_content: tab-delimited text files include RPKM or RPM values and DEseq output files

molecule: genomic DNA

organism: Trypanosoma brucei

source_name: WT cell line_day0

growth_protocol: Trypanosome cells were grown in HMI-9 media at 37C

extract_protocol:
for genomic DNA seq, cells were sorted by cell cycle using flow cytometry and genomic DNA was isolated.
for stranded RNAseq, NEBNext Ultra Directional RNA Library Prep Kit for Illumina
for MFAseq, genomic DNA was fragmented using the NEBNext dsDNA fragmentase (NEB) and sequencing libraries were generated using the NEBNext Ultra DNA Library prep kit and NEB multiplex oligos for Illumina
stranded RNAseq, MFA-seq (or Sort-seq)

treatment_protocol: Cells were treated 0, 1, and 2 days with tetracycline

tag: strain background

value: Lister 427 antigenic type MITat1.2 clone 221a

tag: genotype/variation

value: WT cell line #904

tag: days with tetracycline


title: 7_wo_B6_males_pool1

description: single-cell RNAseq on 10x Genomics Chromium platform of 3 (A,B,C) 7-week-old C57BL/6 male mice as pool1

library_source: transcriptomic

organism: Mus musculus

tag: tissue


title: Strain 904 (WT) G1

description:
processed data file:
BP_MFAseq_sliding window.txt
1_wt_G1


title: Strain 660 day 0 G1

description:
processed data file:
BP_MFAseq_sliding window.txt
5_BPd0_G1

source_name: TbMCM-BP conditional knockout_day0

value: TbMCM-BP conditional knockout cell line # 660


title: T47D_Her2pos_populations.Rescue.seq.HER2.mut.Jan10_S171 [S171_L869R.Fulv]

city: Cambridge

first: Ofir

last: Cohen

state: Massachusetts

address: 415 Main St, Cambridge, MA 02142

department: Cancer Program

institute: Broad Institute

description: Parental cell line with HER2 L869R over-expression treated with 1µM Fulv

data_processing:
Library strategy: Smart-seq 2
mRNA-seq libraries were sequenced on an Illumina NextSeq 500 sequencer, V2 75 cycle kits
Reads were mapped with STAR aligner (Dobin et al. 2013)
Genome_build: hg19 of the human genome and build 12 of the GENCODE human genome annotation
Supplementary_files_format_and_content: rpkm and read counts

molecule: total RNA

organism: Homo sapiens

source_name: Breast Cancer

growth_protocol: T47D cells were infected de novo with lentivirus encoding HER2 mutants and controls (GFP, wild-type HER2, kinase-dead HER2, and ESR1 Y537S) in the pLX307 plasmid. Upon completion of selection with Puromycin, the cells were plated in 96-well plates in RPMI supplemented with 10% CSS for 48 hours.

extract_protocol: Plates with cell lysates were thawed and purified with 2.2x RNAClean SPRI beads (Beckman Coulter Genomics). The RNA captured beads were air-dried and processed immediately for RNA secondary structure denaturation (72˚C for three minutes) and cDNA synthesis. We performed SMART-Seq2 following the published protocol66

treatment_protocol: Cell were then treated with DMSO, 1μM fulvestrant, 1μM neratinib, 10μM palbociclib, 1μM fulvestrant + 1μM neratinib, or 1μM fulvestrant + 10μM palbociclib for 24 hours.

tag: cell line

value: HER2 L869R over-expression

tag: agent

value: 1microM Fulv


title: M3-wt-2

city: Houston

first: Samuel

last: Shelburne

state: TX

address: 1515 Holcombe Blvd

department: Infectious Diseases

institute: MD Anderson Cancer Center

data_processing:
The raw reads in FASTQ format were aligned to the reference genome, MGAS10870 (M3) or MGAS5005 (M1) using MOSAIK alignment software with parameters of -hs 15, -mmp 0.06, -mmal,   -minp 0.5, -act 35, -mhp 100, and -m all
The overlaps between aligned reads and annotated genes were counted using HTSeq software (http://www-huber.embl.de/users/anders/HTSeq/doc/overview.html). The gene counts were normalized using the scaling factor method implemented in the R package of DESeq2 (version 1.2.10). If the number of overlapped read of any given gene is less than one per million total mapped read for all samples, this gene was excluded from further analysis.
Please note that the MGAS10870 strain and its derivative strains get mapped to the publicly available MGAS315 genome  (NC_004070.1/AE014074.1).  For strain MGAS2221 and its derivatives, we map to the publicly available strain MGAS5005 (NZ_CP000017.1/CP000017.2).   MGAS10870 and MGAS2221 are very closely related to MGAS10870 and MGAS2221, respectively.
A negative binomial generalized linear model was fit to each gene expression with strain type as covariates. Then a likelihood-ratio test was applied to examine if there is any difference in the expression of a gene among six strains. The Benjamini-Hochberg method was used to control false discovery rate (FDR). The pairwise comparisons were performed to compare the gene expressions between WT vs any other strain using Wald's tests. The Holm's method was used to calculate adjusted p-value to correct for multiple testing. The calculation was conducted using R (version 3.0.1) with the package of DESeq2 (version 1.2.10)
Genome_build: MGAS315  (NC_004070.1/AE014074.1) and MGAS5005 (NZ_CP000017.1/CP000017.2) genomes
Supplementary_files_format_and_content: normalize counts

organism: Streptococcus sp. 'group A'

source_name: bacteria grown in THY

growth_protocol: GAS strains grown in THY late-exponential phase

extract_protocol:
Cells were lysed using a fast FastPrep approach and RNA was isolated from ruptured cell using an RNeasy Kit (Qiagen)
cDNA was fragmented to 200 bp (mean fragment size) with the S220 Focused-ultrasonicator (Covaris) and used to make barcoded sequencing libraries on the SPRI-TE Nucleic Acid Extractor (Beckman-Coulter).

treatment_protocol: RNA protect was added at 2x volumes to cells and then cells were harvested by centrifugation, snap frozen, and stored at -80 C.

tag: serotype

tag: molecule subtype

value: RNA after rRNA depletion


title: strain 660-2 day 2

description:
processed data file:
BP_strRNAseq_Chromosome_5kb1kb
BP_allgenes_rawreads.txt
BP_allgenes_d2vs0_DEseq_binom_naomit.txt
BP_VSG_rawReads.txt
BPvsg_d2vs0_DEseq_binom_naomit.txt
BP_BES_strRNA_1kb200bp.txt
TTS sites.txt
TSS sites.txt
HT sites.txt
25 TSS 30kb region.txt
21 tts 30kb region.txt
16 HT 30kb region.txt
BPday2_2_CTTGTA_10

source_name: TbMCM-BP conditional knockout_day2

value: TbMCM-BP conditional knockout cell line # 660, culture 2


title: strain 661-1 day 2

description:
processed data file:
BP_strRNAseq_Chromosome_5kb1kb
BP_allgenes_rawreads.txt
BP_allgenes_d2vs0_DEseq_binom_naomit.txt
BP_VSG_rawReads.txt
BPvsg_d2vs0_DEseq_binom_naomit.txt
BP_BES_strRNA_1kb200bp.txt
TTS sites.txt
TSS sites.txt
HT sites.txt
25 TSS 30kb region.txt
21 tts 30kb region.txt
16 HT 30kb region.txt
BPday2_3_GTCCGC_11

value: TbMCM-BP conditional knockout cell line # 661, culture 1


title: merged_D709_D501

first: Pierre

last: Larraufie

address: Box 289, Addenbrooke's Site

department: WT-MRC Institute of Metabolic Science

country: United Kingdom

institute: Metabolic Research Laboratories and MRC Metabolic Diseases Unit

data_processing:
data was demultiplexed using the Casava pipeline
data was mapped to the mouse genome GRCm38 using STAR v2.5.1 and raw counts generated simultaneously
Supplementary_files_format_and_content: Larraufie_VSG_EEC_mouse_rawcount.csv  is a csv file of number of counts for ensembl gene ids of each biological sample

growth_protocol: mice were grown on a 12h dark/light cycle with ad libitum access to water and standard chow

extract_protocol:
epithelium was single-cell digested and FACS sorted for positive cells. RNA was extracted using microplus RNeasy kit (Qiagen) and quality checked on a picochip bioanalyzer
2ng of RNA was used for library contruction using the SMARTer stranded total RNAseq kit v2 pico kit
RNAseq SE-50 on an Illumina HiSEQ 40000

treatment_protocol: VSG mice had ~70% of their stomach removed and sutured, sham were opened and had a suture around without pressure. A group of sham operated were weight matched for the VSG. All mice were fed on a standard chow except around the surgery for which they received liquid diet

tag: condition

value: duodenum

tag: cell type

value: enteroendocrine cells


title: T47D_Her2pos_populations.Rescue.seq.HER2.mut.Jan10_S207 [S207_wt.Fulv]

description: Parental cell line with HER2 wt over-expression treated with 1µM Fulv

value: HER2 wt over-expression


title: strain 661-2 day 2

description:
processed data file:
BP_strRNAseq_Chromosome_5kb1kb
BP_allgenes_rawreads.txt
BP_allgenes_d2vs0_DEseq_binom_naomit.txt
BP_VSG_rawReads.txt
BPvsg_d2vs0_DEseq_binom_naomit.txt
BP_BES_strRNA_1kb200bp.txt
TTS sites.txt
TSS sites.txt
HT sites.txt
25 TSS 30kb region.txt
21 tts 30kb region.txt
16 HT 30kb region.txt
BPday2_4_GTGAAA_12

value: TbMCM-BP conditional knockout cell line # 661, culture 2


title: strain 661-2 day 1

description:
processed data file:
BP_strRNAseq_Chromosome_5kb1kb
BP_allgenes_rawreads.txt
BP_allgenes_d1vs0_DEseq_binom_naomit.txt
BP_VSG_rawReads.txt
BPvsg_d1vs0_DEseq_binom_naomit.txt
BP_BES_strRNA_1kb200bp.txt
TTS sites.txt
TSS sites.txt
HT sites.txt
25 TSS 30kb region.txt
21 tts 30kb region.txt
16 HT 30kb region.txt
BPday1_4_CAGATC_8

source_name: TbMCM-BP conditional knockout_day1


title: T47D_Her2pos_populations.Rescue.seq.HER2.mut.Jan10_S87 [S87_ESR1.Fulv]

description: Parental cell line with ESR1 Y537S over-expression treated with 1µM Fulv

value: ESR1 Y537S over-expression


title: strain 661-1 day 1

description:
processed data file:
BP_strRNAseq_Chromosome_5kb1kb
BP_allgenes_rawreads.txt
BP_allgenes_d1vs0_DEseq_binom_naomit.txt
BP_VSG_rawReads.txt
BPvsg_d1vs0_DEseq_binom_naomit.txt
BP_BES_strRNA_1kb200bp.txt
TTS sites.txt
TSS sites.txt
HT sites.txt
25 TSS 30kb region.txt
21 tts 30kb region.txt
16 HT 30kb region.txt
BPday1_3_GCCAAT_7


title: strain 660-2 day 1

description:
processed data file:
BP_strRNAseq_Chromosome_5kb1kb
BP_allgenes_rawreads.txt
BP_allgenes_d1vs0_DEseq_binom_naomit.txt
BP_VSG_rawReads.txt
BPvsg_d1vs0_DEseq_binom_naomit.txt
BP_BES_strRNA_1kb200bp.txt
TTS sites.txt
TSS sites.txt
HT sites.txt
25 TSS 30kb region.txt
21 tts 30kb region.txt
16 HT 30kb region.txt
BPday1_2_ATGTCA_6


title: strain 661-2 day 0

description:
processed data file:
BP_strRNAseq_Chromosome_5kb1kb
BP_allgenes_rawreads.txt
BP_allgenes_d1vs0_DEseq_binom_naomit.txt
BP_allgenes_d2vs0_DEseq_binom_naomit.txt
BP_VSG_rawReads.txt
BPvsg_d1vs0_DEseq_binom_naomit.txt
BPvsg_d2vs0_DEseq_binom_naomit.txt
BP_BES_strRNA_1kb200bp.txt
TTS sites.txt
TSS sites.txt
HT sites.txt
25 TSS 30kb region.txt
21 tts 30kb region.txt
16 HT 30kb region.txt
BPday0_4_ACAGTG_4


title: T47D_Her2pos_populations.Rescue.seq.HER2.mut.Jan10_S99 [S99_GFP.Fulv]

description: Parental cell line with GFP over-expression treated with 1µM Fulv

value: GFP over-expression


title: strain 661-1 day 0

description:
processed data file:
BP_strRNAseq_Chromosome_5kb1kb
BP_allgenes_rawreads.txt
BP_allgenes_d1vs0_DEseq_binom_naomit.txt
BP_allgenes_d2vs0_DEseq_binom_naomit.txt
BP_VSG_rawReads.txt
BPvsg_d1vs0_DEseq_binom_naomit.txt
BPvsg_d2vs0_DEseq_binom_naomit.txt
BP_BES_strRNA_1kb200bp.txt
TTS sites.txt
TSS sites.txt
HT sites.txt
25 TSS 30kb region.txt
21 tts 30kb region.txt
16 HT 30kb region.txt
BPday0_3_AGTCAA_3


title: T47D_Her2pos_populations.Rescue.seq.HER2.mut.Jan10_S255 [S255_V777L.Fulv]

description: Parental cell line with HER2 V777L over-expression treated with 1µM Fulv

value: HER2 V777L over-expression


title: strain 660-2 day 0

description:
processed data file:
BP_strRNAseq_Chromosome_5kb1kb
BP_allgenes_rawreads.txt
BP_allgenes_d1vs0_DEseq_binom_naomit.txt
BP_allgenes_d2vs0_DEseq_binom_naomit.txt
BP_VSG_rawReads.txt
BPvsg_d1vs0_DEseq_binom_naomit.txt
BPvsg_d2vs0_DEseq_binom_naomit.txt
BP_BES_strRNA_1kb200bp.txt
TTS sites.txt
TSS sites.txt
HT sites.txt
25 TSS 30kb region.txt
21 tts 30kb region.txt
16 HT 30kb region.txt
BPday0_2_TGACCA_2


title: mRNA_E-15_non-Vh11

city: Rotterdam

first: Ralph

last: Stadhouders

address: Dr. Molewaterplein 50

department: Pulmonary Medicine

country: Netherlands

institute: Erasmus Medical Center

description:
s_2_1_I12-1297-26_E-15_non-Vh11
I12-1297-26_E-15_non-Vh11/ FPKM

data_processing:
Reads were aligned using Bowtie (http://bowtie-bio.sourceforge.net/index.shtml) with default parameters.
Sorted and indexed bam files were parsed to HOMER to create tag directories without duplicates and the generation of RPKM values per gene.
Sample scaling and statistical analysis were performed using the R package DESeq2 implementation in HOMER
Genome_build: mm9
Supplementary_files_format_and_content: RPKM expression values

growth_protocol: To obtain aIgM-stimulated B cells, purified naïve WT B cells were cultured in culture medium (RPMI 1640 (life technologies)/ 10% FCS (gibco) / 50 μg/mL gentamycin(life technologies) / 0,05 mM ß-mercaptoethanol (Sigma)) in the presence of 10 μg/ml F(ab’)2 anti-IgM (Jackson Immunoresearch) for 12h.

extract_protocol:
Splenic single-cell suspensions were prepared in magnetic-activated cell sorting (MACS) buffer (PBS/2mM EDTA/0.5%BSA) and naïve splenic B cells from 8–12 week-old WT C57BL/6 mice were purified by MACS. Non-B cells, B-1 cells, GC B cells and plasma cells were first labeled with biotinylated antibodies (BD Biosciences) to CD5 (53–7.3), CD11b (M1-70), CD43 (S7), CD95 (Jo2), CD138 (281-2), Gr-1 (RB6-8C5) and TER-119 (PK136) and subsequently with streptavidin-conjugated magnetic beads (Miltenyi Biotec). Purity of MACS-sorted naïve B cells was confirmed by flow cytometry (typically > 99% CD19+ cells). CLL cells were purified from primary tumors from IgH.TEmu mice using CD19+IgMb+ MACS purification.
RNA-Seq: standard Illumina TruSeq library construction.

value: 8-12 weeks

value: wild type

value: spleen

value: MACS-purified lymphocyte

tag: cell subtype

value: non-VH11 chronic lymphocytic leukemia (CLL)


title: mRNA_E-29_non-Vh11

description:
s_2_1_I12-1297-27_E-29_non-Vh11
I12-1297-27_E-29_non-Vh11/ FPKM


title: mRNA_ET-06_non-Vh11

description:
s_2_1_I12-1297-25_ET-06_non-Vh11
I12-1297-25_ET-06_non-Vh11/ FPKM


title: T47D_Her2pos_populations.Rescue.seq.HER2.mut.Jan10_S111 [S111_wt.Fulv]


title: T47D_Her2pos_populations.Rescue.seq.HER2.mut.Jan10_S219 [S219_D845A.Fulv]

description: Parental cell line with HER2 D845A over-expression treated with 1µM Fulv

value: HER2 D845A over-expression


title: mRNA_EA-04_Vh11

description:
s_2_1_I12-1297-23_EA-04_Vh11
I12-1297-23_EA-04_Vh11/ FPKM

value: VH11 chronic lymphocytic leukemia (CLL)


title: strain 660-1 day 2

description:
processed data file:
BP_strRNAseq_Chromosome_5kb1kb
BP_allgenes_rawreads.txt
BP_allgenes_d2vs0_DEseq_binom_naomit.txt
BP_VSG_rawReads.txt
BPvsg_d2vs0_DEseq_binom_naomit.txt
BP_BES_strRNA_1kb200bp.txt
TTS sites.txt
TSS sites.txt
HT sites.txt
25 TSS 30kb region.txt
21 tts 30kb region.txt
16 HT 30kb region.txt
BPday2_1_CCGTCC_9

value: Lister 427 antigenic type MITat1.2 clone 221a

value: TbMCM-BP conditional knockout cell line # 660, culture 1


title: T47D_Her2pos_populations.Rescue.seq.HER2.mut.Jan10_S266 [S266_L869R.DMSO]

description: Parental cell line with HER2 L869R over-expression treated with DMSO

value: HER2 L869R over-expression

value: DMSO


title: strain 660-1 day 0

description:
processed data file:
BP_strRNAseq_Chromosome_5kb1kb
BP_allgenes_rawreads.txt
BP_allgenes_d1vs0_DEseq_binom_naomit.txt
BP_allgenes_d2vs0_DEseq_binom_naomit.txt
BP_VSG_rawReads.txt
BPvsg_d1vs0_DEseq_binom_naomit.txt
BPvsg_d2vs0_DEseq_binom_naomit.txt
BP_BES_strRNA_1kb200bp.txt
TTS sites.txt
TSS sites.txt
HT sites.txt
25 TSS 30kb region.txt
21 tts 30kb region.txt
16 HT 30kb region.txt
BPday0_1_CGATGT_1


title: strain 660-1 day 1

description:
processed data file:
BP_strRNAseq_Chromosome_5kb1kb
BP_allgenes_rawreads.txt
BP_allgenes_d1vs0_DEseq_binom_naomit.txt
BP_VSG_rawReads.txt
BPvsg_d1vs0_DEseq_binom_naomit.txt
BP_BES_strRNA_1kb200bp.txt
TTS sites.txt
TSS sites.txt
HT sites.txt
25 TSS 30kb region.txt
21 tts 30kb region.txt
16 HT 30kb region.txt
BPday1_1_AGTTCC_5


title: T47D_Her2pos_populations.Rescue.seq.HER2.mut.Jan10_S278 [S278_ESR1.DMSO]

description: Parental cell line with ESR1 Y537S over-expression treated with DMSO


title: T47D_Her2pos_populations.Rescue.seq.HER2.mut.Jan10_S134 [S134_S653C.DMSO]

description: Parental cell line with HER2 S653C over-expression treated with DMSO

value: HER2 S653C over-expression


title: mRNA_E-06_Vh11

description:
s_2_1_I12-1297-21_E-06_Vh11
I12-1297-21_E-06_Vh11/ FPKM


title: mRNA_EA-02_Vh11

description:
s_2_1_I12-1297-22_EA-02_Vh11
I12-1297-22_EA-02_Vh11/ FPKM


title: T47D_Her2pos_populations.Rescue.seq.HER2.mut.Jan10_S242 [S242_L755S.DMSO]

description: Parental cell line with HER2 L755S over-expression treated with DMSO

value: HER2 L755S over-expression


title: mRNA_EQ_wt3_aIgM

description:
s_7_1_I12-1297-11_EQ_wt3_aIgM
I12-1297-11_EQ_wt3_aIgM/ FPKM

source_name: aIgM-stimulated B cells

value: aIgM-stimulated WT B cells


title: mRNA_EQ_wt1_aIgM

description:
s_7_1_I12-1297-09_EQ_wt1_aIgM
I12-1297-09_EQ_wt1_aIgM/ FPKM


title: mRNA_EQ_wt4_unst

description:
s_6_1_I12-1297-04_EQ_wt4_unst
I12-1297-04_EQ_wt4_unst/ FPKM

source_name: unstimulated B cells

value: naïve WT B cells


title: T47D_Her2pos_populations.Rescue.seq.HER2.mut.Jan10_S74 [S74_L869R.DMSO]


title: mRNA_EQ_wt2_unst

description:
s_6_1_I12-1297-02_EQ_wt2_unst
I12-1297-02_EQ_wt2_unst/ FPKM


title: T47D_Her2pos_populations.Rescue.seq.HER2.mut.Jan10_S194 [S194_GFP.DMSO]

description: Parental cell line with GFP over-expression treated with DMSO

value: GFP over-expression


title: mRNA_EQ_wt1_unst

description:
s_6_1_I12-1297-01_EQ_wt1_unst
I12-1297-01_EQ_wt1_unst/ FPKM


title: mRNA_EQ_wt3_unst

description:
s_6_1_I12-1297-03_EQ_wt3_unst
I12-1297-03_EQ_wt3_unst/ FPKM


title: Nascent_RNAseq_siEsrrb_R2

city: Los Angeles

first: Siavash

middle: K

last: Kurdistani

state: CA

address: 615 Charles E Young Dr South

department: Biological Chemistry

institute: UCLA

data_processing:
Basecalls performed using CASAVA version 1.4
For RNA-seq, sequenced reads were mapped to mm9 using TopHat2 with option –g 1 –p 4 –N 2. For mRNA-seq, gene transcription levels were normalized to RPKM using SAMMate version 2.7.4 with Read Assignment Expectation Maximiation (RAEM). For Nascent RNA-seq, RNA levels in the entire gene body were normalized by calculating RPKM using a custom script.
Genome_build: mm9
Supplementary_files_format_and_content: RPKM for each gene.

growth_protocol: E14 mESCs were cultured on 0.2% gelatinized tissue culture plates in mESC media containing DMEM-KO (Invitrogen, 10829-018) supplemented with 15% fetal bovine serum, 1000 U/ml LIF, 100 µM nonessential amino acids (Invitrogen, 11140-050), 2 mM L-glutamine (Invitrogen, 25030-081) and 8 nL/ml of 2-mercaptoethanol at 37oC in a humidified 5% CO2/95% air incubator.

extract_protocol:
mESCs were harvested, washed with DPBS and lysed in 200 μl of ice-cold lysis buffer (10 mM Tris-HCl pH 7.5, 0.1% NP40, 150 mM NaCl). The cell lysate was gently layered over 500 μl of chilled sucrose cushion (24% RNAse-free sucrose in lysis buffer) in a new Eppendorf tube and centrifuged for 10 min at 4°C, 10,000xg. The supernatant (cytoplasmic fraction) was removed and the pellet (nuclei) was washed once with 200 μl of ice-cold 1 × PBS/1 mM EDTA. The nuclear pellet was resuspended in 100 μl of pre-chilled glycerol buffer (20 mM Tris-HCl pH 7.9, 75 mM NaCl, 0.5 mM EDTA, 0.85 mM DTT, 0.125 mM PMSF, 50% glycerol) by gentle flicking of the tube. An equal volume (100 μl) of cold nuclear lysis buffer (10 mM HEPES pH 7.6, 1 mM DTT, 7.5 mM MgCl2, 0.2 mM EDTA, 0.3 M NaCl, 1 M urea, 1% NP-40) was added. The mix was vortexed vigorously for 2 s. The sample was incubated for 2 min on ice, and then centrifuged for 2 min at 4°C, 10,000xg. The supernatant (nuclear fraction/nucleoplasm) was removed and the pellet (chromatin) was gently rinsed with ice-cold 1×PBS/1 mM EDTA. 1 ml of Trizol reagent was added to the chromatin and incubated for 30 min at 50oC to dissolve it. Nascent RNA was extracted following the manufacturer’s instructions.
Libraries were constructed using KAPA Stranded RNA-Seq Kits with RiboErase (HMR).
Nascent RNA-seq libraries were constructed using KAPA Stranded RNA-Seq Kits with RiboErase (HMR).

treatment_protocol: siRNA was transfected into mESCs using Lipofectamine RNAiMAX following manufacturer's protocol. Cells were harvested and analyzed 72 h post transfection

value: embryonic stem cells (mESC)

value: siEsrrb

value: nuclear RNA


title: mRNA_EQ_wt4_aIgM

description:
s_7_1_I12-1297-12_EQ_wt4_aIgM
I12-1297-12_EQ_wt4_aIgM/ FPKM

value: 8-12 weeks

value: wild type

value: spleen


title: Nascent_RNAseq_siCtrl_R2

value: siCtrl


title: mRNA_EQ_wt2_aIgM

description:
s_7_1_I12-1297-10_EQ_wt2_aIgM
I12-1297-10_EQ_wt2_aIgM/ FPKM

value: MACS-purified lymphocyte


title: T47D_Her2pos_populations.Rescue.seq.HER2.mut.Jan10_S62 [S62_V777L.DMSO]

description: Parental cell line with HER2 V777L over-expression treated with DMSO

value: HER2 V777L over-expression


title: T47D_Her2pos_populations.Rescue.seq.HER2.mut.Jan10_S254 [S254_V777L.DMSO]


title: T47D_Her2pos_populations.Rescue.seq.HER2.mut.Jan10_S26 [S26_D845A.DMSO]

description: Parental cell line with HER2 D845A over-expression treated with DMSO

value: HER2 D845A over-expression


title: 4C_Tbx3_siCtrl_Rep2

data_processing:
Basecalls performed using CASAVA version 1.4
For ChIP-seq, sequenced reads were mapped to mm9 using Bowtie version 1.1.1. Only those reads aligning to a unique position with no more than 2 mismatches were kept for downstream processing. After mapping, clonal reads in each sample were removed. To call peaks, the mouse genome was segmented into 50-bp windows. A Poisson p-value was calculated for each window based on the IP and normalized input counts in this window. A significant peak was retained only when p-values of its own window and the 2 neighboring windows were all below 0.001. If different IPs (e.g. siCtrl vs. siEsrrb) of the same protein were compared in downstream analyses, their total counts were first shuffled to the same number before peak calling. Their inputs were also shuffled similarly.
For RNA-seq, sequenced reads were mapped to mm9 using TopHat2 with option –g 1 –p 4 –N 2. For mRNA-seq, gene transcription levels were normalized to RPKM using SAMMate version 2.7.4 with Read Assignment Expectation Maximiation (RAEM). For Nascent RNA-seq, RNA levels in the entire gene body were normalized by calculating RPKM using a custom script.
For 4C-seq analysis using 4Cseqpipe, the fastq file containing all the reads was first converted to the format compatible for downstream processing using the “-fastq2raw” function of the pipeline, followed by de-multiplexing based on the barcode and reading primer sequence for each library. Using the “–map” function, reads were mapped to fragmented mm9 genome digested by the restriction enzyme in silico. Normalizing and graphing were performed using the “-nearcis” function. Default settings were used in all steps. For analysis by 4C-Ker, a reduced genome was built using Bowtie2. The reduced genome contains only sequences flanking the primary restriction enzyme sites (51 bp on both sides). Reads were de-multiplexed and barcode and primer sequences were trimmed using a custom script. Trimmed reads were mapped to the reduced genome by Bowtie2 with option –p 12 –N 0. To remove self-ligated and undigested fragments, the fragments before and after the bait fragment were removed. Near-cis interactions were analyzed using the “nearBaitAnalysis” function in 4C-Ker with default settings.
Genome_build: mm9
Supplementary_files_format_and_content: For ChIP-seq, bigWig files are provided containing normalized IP reads. For RNA-seq, xlsx files are provided containing RPKM for each gene. For 4C-seq, bedGraph files are provided containing nearbait normalized counts.

extract_protocol:
Crosslinked cells were lysed for 15 min on ice in 10 mM Tris-HCl pH 8.0, 10 mM NaCl, and 0.2% NP-40 supplemented with cOmplete protease inhibitor. Nuclei were isolated by centrifugation and by removing the supernatant. Nuclei corresponding to 10 million cells were resuspended in 500 μl of the primary restriction enzyme buffer. SDS was added to a final concentration of 0.3% and samples were incubated for 1 h at 37oC at 1,200 rpm on a thermo-mixer. SDS was quenched by addition of Triton X-100 to a final concentration of 3%. Primary restriction enzyme (400 units) was added and samples were digested for 6 h on a thermo-mixer followed by addition of 400 additional units of the primary restriction enzyme and overnight digestion. The primary restriction enzyme was inactivated by heating to 65oC for 20 min. Samples were diluted to a total volume of 8 ml in ligation buffer (66 mM Tris-HCl pH 7.5, 5 mM MgCl2, 5 mM DTT, 1 mM ATP). Proximity ligation was carried out by adding 4000 units of T4 DNA ligase and incubating at room temperature overnight. After reversal of -crosslinking and RNA removal, DNA was extracted by phenol/chloroform and purified by ethanol precipitation. A secondary digestion was performed overnight in a volume of 500 μl with 200 units of the secondary restriction enzyme. For secondary ligation, following inactivation of the restriction enzyme, each sample was diluted to 14 ml with ligation buffer. T4 DNA ligase (4000 units) was added and samples were incubated overnight at room temperature. DNA was extracted by phenol/chloroform and purified by ethanol precipitation. To remove salts, DNA was further purified using QIAquick PCR Cleanup kit.
To construct 4C-seq libraries, reading and non-reading primers were designed for each viewpoint. PCR was performed in sixteen 25-μl PCR reactions using Platinum Taq DNA Polymerase High Fidelity. Reaction conditions were as in the manufacturer’s instructions except 1.4 μM of each primer and 200-400 ng of template were used in each reaction. The PCR program was as follows: (1) 95oC, 5 min; (2) 95oC, 30 s; (3) 55oC, 1 min; (4) 68oC, 3 min; (5) go to (2), 30 cycles; (6) 68oC, 7 min. PCR products were first purified using illustra GFX PCR DNA and Gel Band Purification Kit (GE Healthcare) to remove primer dimers and then further purified using QIAquick PCR Purification Kit to remove salt.
For ChIP-seq, crosslinked cells were resuspended in swelling buffer (25 mM HEPES pH 7.9, 1.5 mM MgCl2, 10 mM KCl, 0.1% NP-40, and 1X cOmplete protease inhibitor cocktail [Roche]) and incubated for 10 min on ice. Cells were centrifuged and the cell pellet was resuspended in Buffer A (50 mM HEPES-KOH pH 7.9, 140 mM NaCl, 1 mM EDTA, 1% Triton X-100, 0.1% Na-deoxycholate, 0.1% SDS) plus 1X cOmplete protease inhibitor cocktail. Cell sonication was performed on a Qsonica Q800R2 sonicator with 20% amplitude for 20 cycles at 10s each with 30s between cycles at 4oC. Sonicated lysate was pre-cleared by incubating with Dynabeads Protein A/G. Part of the pre-cleared lysate was used as input and the remainder was incubated overnight at 4°C with 2-10 micrograms of antibody. DNA/Protein-antibody conjugates were precipitated using Dynabeads Protein A/G blocked with 5 mg/ml BSA in PBS. Beads were washed twice each with Buffer A, Buffer B (50 mM HEPES-KOH pH 7.9, 500 mM NaCl, 1 mM EDTA, 1% Triton X-100, 0.1% Na-deoxycholate, 0.1% SDS), LiCl buffer (20 mM Tris-HCl pH 8.0, 250 mM LiCl, 1 mM EDTA, 0.5% Na-deoxycholate, 0.5% NP-40) and TE buffer (10 mM Tris-HCl pH 8.0, 1 mM EDTA). DNA was eluted in elution buffer (10 mM Tris-HCl pH 8.0, 1 mM EDTA, 1% SDS). Cross-links were reversed overnight at 65°C. RNA and protein were digested using RNase A and Proteinase K, respectively, and DNA was purified by phenol-chloroform extraction and ethanol precipitation.
For mRNA-seq, total RNA was extracted from mESCs using TRIzol reagent (Invitrogen) according to manufacturer’s instruction. To remove genomic DNA contamination, RNA samples were treated with DNaseI and purified again by phenol/chloroform extraction and ethanol precipitation.
For Nascent RNA-seq, mESCs were harvested, washed with DPBS and lysed in 200 μl of ice-cold lysis buffer (10 mM Tris-HCl pH 7.5, 0.1% NP40, 150 mM NaCl). The cell lysate was gently layered over 500 μl of chilled sucrose cushion (24% RNAse-free sucrose in lysis buffer) in a new Eppendorf tube and centrifuged for 10 min at 4°C, 10,000xg. The supernatant (cytoplasmic fraction) was removed and the pellet (nuclei) was washed once with 200 μl of ice-cold 1 × PBS/1 mM EDTA. The nuclear pellet was resuspended in 100 μl of pre-chilled glycerol buffer (20 mM Tris-HCl pH 7.9, 75 mM NaCl, 0.5 mM EDTA, 0.85 mM DTT, 0.125 mM PMSF, 50% glycerol) by gentle flicking of the tube. An equal volume (100 μl) of cold nuclear lysis buffer (10 mM HEPES pH 7.6, 1 mM DTT, 7.5 mM MgCl2, 0.2 mM EDTA, 0.3 M NaCl, 1 M urea, 1% NP-40) was added. The mix was vortexed vigorously for 2 s. The sample was incubated for 2 min on ice, and then centrifuged for 2 min at 4°C, 10,000xg. The supernatant (nuclear fraction/nucleoplasm) was removed and the pellet (chromatin) was gently rinsed with ice-cold 1×PBS/1 mM EDTA. 1 ml of Trizol reagent was added to the chromatin and incubated for 30 min at 50oC to dissolve it. Nascent RNA was extracted following the manufacturer’s instructions.
For 4C-seq, crosslinked cells were lysed for 15 min on ice in 10 mM Tris-HCl pH 8.0, 10 mM NaCl, and 0.2% NP-40 supplemented with cOmplete protease inhibitor. Nuclei were isolated by centrifugation and by removing the supernatant. Nuclei corresponding to 10 million cells were resuspended in 500 μl of the primary restriction enzyme buffer. SDS was added to a final concentration of 0.3% and samples were incubated for 1 h at 37oC at 1,200 rpm on a thermo-mixer. SDS was quenched by addition of Triton X-100 to a final concentration of 3%. Primary restriction enzyme (400 units) was added and samples were digested for 6 h on a thermo-mixer followed by addition of 400 additional units of the primary restriction enzyme and overnight digestion. The primary restriction enzyme was inactivated by heating to 65oC for 20 min. Samples were diluted to a total volume of 8 ml in ligation buffer (66 mM Tris-HCl pH 7.5, 5 mM MgCl2, 5 mM DTT, 1 mM ATP). Proximity ligation was carried out by adding 4000 units of T4 DNA ligase and incubating at room temperature overnight. After reversal of -crosslinking and RNA removal, DNA was extracted by phenol/chloroform and purified by ethanol precipitation. A secondary digestion was performed overnight in a volume of 500 μl with 200 units of the secondary restriction enzyme. For secondary ligation, following inactivation of the restriction enzyme, each sample was diluted to 14 ml with ligation buffer. T4 DNA ligase (4000 units) was added and samples were incubated overnight at room temperature. DNA was extracted by phenol/chloroform and purified by ethanol precipitation. To remove salts, DNA was further purified using QIAquick PCR Cleanup kit.
For ChIP-seq, libraries were prepared using a KAPA LTP kit following manufacturer's instructions.
For mRNA-seq, libraries were prepared with KAPA stranded mRNA-Seq kits.
For Nascent RNA-seq, libraries were constructed using KAPA Stranded RNA-Seq Kits with RiboErase (HMR).
For 4C-seq libraries construction, reading and non-reading primers were designed for each viewpoint. PCR was performed in sixteen 25-μl PCR reactions using Platinum Taq DNA Polymerase High Fidelity. Reaction conditions were as in the manufacturer’s instructions except 1.4 μM of each primer and 200-400 ng of template were used in each reaction. The PCR program was as follows: (1) 95oC, 5 min; (2) 95oC, 30 s; (3) 55oC, 1 min; (4) 68oC, 3 min; (5) go to (2), 30 cycles; (6) 68oC, 7 min. PCR products were first purified using illustra GFX PCR DNA and Gel Band Purification Kit (GE Healthcare) to remove primer dimers and then further purified using

treatment_protocol:
siRNA was transfected into mESCs using Lipofectamine RNAiMAX following manufacturer's protocol. Cells were harvested and analyzed 72 h post transfection
For knockdown by siRNA, siRNA was transfected into mESCs using Lipofectamine RNAiMAX following manufacturer's protocol. Cells were harvested and analyzed 72 h post transfection.
For knockdown by shRNA, plasmids expressing shRNA were transfected into mESCs using Lipofectamine 2000 following manufacturer's protocol. Selection with puromycin started 24 h post transfection. After selection for 72 h, cells were harvested for downstream analysis.


title: T47D_Her2pos_populations.Rescue.seq.HER2.mut.Jan10_S2 [S2_GFP.DMSO]


title: 4C_Klf4_siEsrrb_Rep2


title: 4C_Tbx3_siCtrl_Rep1


title: 4C_Klf4_siCtrl_Rep2


title: 4C_Tbx3_siEsrrb_Rep1


title: T47D_Her2pos_populations.Rescue.seq.HER2.mut.Jan10_S38 [S38_S653C.DMSO]


title: 4C_Klf4_siEsrrb_Rep1


title: 4C_Tbx3_siEsrrb_Rep2


title: T47D_Her2pos_populations.Rescue.seq.HER2.mut.Jan10_S158 [S158_V777L.DMSO]


title: 4C_Klf4_siCtrl_Rep1


title: 4C_Cdyl2_siEsrrb_Rep2


title: 4C_Cdyl2_siEsrrb_Rep1


title: T47D_Her2pos_populations.Rescue.seq.HER2.mut.Jan10_S98 [S98_GFP.DMSO]


title: 4C_Cdyl2_siCtrl_Rep2


title: T47D_Her2pos_populations.Rescue.seq.HER2.mut.Jan10_S50 [S50_L755S.DMSO]


title: 4C_Cdly2_siCtrl_Rep1


title: T47D_Her2pos_populations.Rescue.seq.HER2.mut.Jan10_S122 [S122_D845A.DMSO]


title: T47D_Her2pos_populations.Rescue.seq.HER2.mut.Jan10_S230 [S230_S653C.DMSO]


title: 4C_Sik1_siEsrrb_Rep1

value: sisiEsrrb


title: 4C_Sik1_siEsrrb_Rep2


title: 4C_Sik1_siCtrl_Rep2

value: sisiCtrl


title: 4C_Mitf_siCtrl_Rep2


title: 4C_Sik1_siCtrl_Rep1


title: T47D_Her2pos_populations.Rescue.seq.HER2.mut.Jan10_S86 [S86_ESR1.DMSO]

value: ESR1 Y537S over-expression

value: DMSO


title: 4C_Mitf_siCtrl_Rep1


title: T47D_Her2pos_populations.Rescue.seq.HER2.mut.Jan10_S14 [S14_wt.DMSO]

description: Parental cell line with HER2 wt over-expression treated with DMSO

value: HER2 wt over-expression


title: Nascent_RNAseq_siEsrrb

extract_protocol:
mESCs were harvested, washed with DPBS and lysed in 200 μl of ice-cold lysis buffer (10 mM Tris-HCl pH 7.5, 0.1% NP40, 150 mM NaCl). The cell lysate was gently layered over 500 μl of chilled sucrose cushion (24% RNAse-free sucrose in lysis buffer) in a new Eppendorf tube and centrifuged for 10 min at 4°C, 10,000xg. The supernatant (cytoplasmic fraction) was removed and the pellet (nuclei) was washed once with 200 μl of ice-cold 1 × PBS/1 mM EDTA. The nuclear pellet was resuspended in 100 μl of pre-chilled glycerol buffer (20 mM Tris-HCl pH 7.9, 75 mM NaCl, 0.5 mM EDTA, 0.85 mM DTT, 0.125 mM PMSF, 50% glycerol) by gentle flicking of the tube. An equal volume (100 μl) of cold nuclear lysis buffer (10 mM HEPES pH 7.6, 1 mM DTT, 7.5 mM MgCl2, 0.2 mM EDTA, 0.3 M NaCl, 1 M urea, 1% NP-40) was added. The mix was vortexed vigorously for 2 s. The sample was incubated for 2 min on ice, and then centrifuged for 2 min at 4°C, 10,000xg. The supernatant (nuclear fraction/nucleoplasm) was removed and the pellet (chromatin) was gently rinsed with ice-cold 1×PBS/1 mM EDTA. 1 ml of Trizol reagent was added to the chromatin and incubated for 30 min at 50oC to dissolve it. Nascent RNA was extracted following the manufacturer’s instructions.
Libraries were constructed using KAPA Stranded RNA-Seq Kits with RiboErase (HMR).
For ChIP-seq, crosslinked cells were resuspended in swelling buffer (25 mM HEPES pH 7.9, 1.5 mM MgCl2, 10 mM KCl, 0.1% NP-40, and 1X cOmplete protease inhibitor cocktail [Roche]) and incubated for 10 min on ice. Cells were centrifuged and the cell pellet was resuspended in Buffer A (50 mM HEPES-KOH pH 7.9, 140 mM NaCl, 1 mM EDTA, 1% Triton X-100, 0.1% Na-deoxycholate, 0.1% SDS) plus 1X cOmplete protease inhibitor cocktail. Cell sonication was performed on a Qsonica Q800R2 sonicator with 20% amplitude for 20 cycles at 10s each with 30s between cycles at 4oC. Sonicated lysate was pre-cleared by incubating with Dynabeads Protein A/G. Part of the pre-cleared lysate was used as input and the remainder was incubated overnight at 4°C with 2-10 micrograms of antibody. DNA/Protein-antibody conjugates were precipitated using Dynabeads Protein A/G blocked with 5 mg/ml BSA in PBS. Beads were washed twice each with Buffer A, Buffer B (50 mM HEPES-KOH pH 7.9, 500 mM NaCl, 1 mM EDTA, 1% Triton X-100, 0.1% Na-deoxycholate, 0.1% SDS), LiCl buffer (20 mM Tris-HCl pH 8.0, 250 mM LiCl, 1 mM EDTA, 0.5% Na-deoxycholate, 0.5% NP-40) and TE buffer (10 mM Tris-HCl pH 8.0, 1 mM EDTA). DNA was eluted in elution buffer (10 mM Tris-HCl pH 8.0, 1 mM EDTA, 1% SDS). Cross-links were reversed overnight at 65°C. RNA and protein were digested using RNase A and Proteinase K, respectively, and DNA was purified by phenol-chloroform extraction and ethanol precipitation.
For mRNA-seq, total RNA was extracted from mESCs using TRIzol reagent (Invitrogen) according to manufacturer’s instruction. To remove genomic DNA contamination, RNA samples were treated with DNaseI and purified again by phenol/chloroform extraction and ethanol precipitation.
For Nascent RNA-seq, mESCs were harvested, washed with DPBS and lysed in 200 μl of ice-cold lysis buffer (10 mM Tris-HCl pH 7.5, 0.1% NP40, 150 mM NaCl). The cell lysate was gently layered over 500 μl of chilled sucrose cushion (24% RNAse-free sucrose in lysis buffer) in a new Eppendorf tube and centrifuged for 10 min at 4°C, 10,000xg. The supernatant (cytoplasmic fraction) was removed and the pellet (nuclei) was washed once with 200 μl of ice-cold 1 × PBS/1 mM EDTA. The nuclear pellet was resuspended in 100 μl of pre-chilled glycerol buffer (20 mM Tris-HCl pH 7.9, 75 mM NaCl, 0.5 mM EDTA, 0.85 mM DTT, 0.125 mM PMSF, 50% glycerol) by gentle flicking of the tube. An equal volume (100 μl) of cold nuclear lysis buffer (10 mM HEPES pH 7.6, 1 mM DTT, 7.5 mM MgCl2, 0.2 mM EDTA, 0.3 M NaCl, 1 M urea, 1% NP-40) was added. The mix was vortexed vigorously for 2 s. The sample was incubated for 2 min on ice, and then centrifuged for 2 min at 4°C, 10,000xg. The supernatant (nuclear fraction/nucleoplasm) was removed and the pellet (chromatin) was gently rinsed with ice-cold 1×PBS/1 mM EDTA. 1 ml of Trizol reagent was added to the chromatin and incubated for 30 min at 50oC to dissolve it. Nascent RNA was extracted following the manufacturer’s instructions.
For 4C-seq, crosslinked cells were lysed for 15 min on ice in 10 mM Tris-HCl pH 8.0, 10 mM NaCl, and 0.2% NP-40 supplemented with cOmplete protease inhibitor. Nuclei were isolated by centrifugation and by removing the supernatant. Nuclei corresponding to 10 million cells were resuspended in 500 μl of the primary restriction enzyme buffer. SDS was added to a final concentration of 0.3% and samples were incubated for 1 h at 37oC at 1,200 rpm on a thermo-mixer. SDS was quenched by addition of Triton X-100 to a final concentration of 3%. Primary restriction enzyme (400 units) was added and samples were digested for 6 h on a thermo-mixer followed by addition of 400 additional units of the primary restriction enzyme and overnight digestion. The primary restriction enzyme was inactivated by heating to 65oC for 20 min. Samples were diluted to a total volume of 8 ml in ligation buffer (66 mM Tris-HCl pH 7.5, 5 mM MgCl2, 5 mM DTT, 1 mM ATP). Proximity ligation was carried out by adding 4000 units of T4 DNA ligase and incubating at room temperature overnight. After reversal of -crosslinking and RNA removal, DNA was extracted by phenol/chloroform and purified by ethanol precipitation. A secondary digestion was performed overnight in a volume of 500 μl with 200 units of the secondary restriction enzyme. For secondary ligation, following inactivation of the restriction enzyme, each sample was diluted to 14 ml with ligation buffer. T4 DNA ligase (4000 units) was added and samples were incubated overnight at room temperature. DNA was extracted by phenol/chloroform and purified by ethanol precipitation. To remove salts, DNA was further purified using QIAquick PCR Cleanup kit.
For ChIP-seq, libraries were prepared using a KAPA LTP kit following manufacturer's instructions.
For mRNA-seq, libraries were prepared with KAPA stranded mRNA-Seq kits.
For Nascent RNA-seq, libraries were constructed using KAPA Stranded RNA-Seq Kits with RiboErase (HMR).
For 4C-seq libraries construction, reading and non-reading primers were designed for each viewpoint. PCR was performed in sixteen 25-μl PCR reactions using Platinum Taq DNA Polymerase High Fidelity. Reaction conditions were as in the manufacturer’s instructions except 1.4 μM of each primer and 200-400 ng of template were used in each reaction. The PCR program was as follows: (1) 95oC, 5 min; (2) 95oC, 30 s; (3) 55oC, 1 min; (4) 68oC, 3 min; (5) go to (2), 30 cycles; (6) 68oC, 7 min. PCR products were first purified using illustra GFX PCR DNA and Gel Band Purification Kit (GE Healthcare) to remove primer dimers and then further purified using QIAquick PCR Purification Kit to remove salt.

value: nulcear RNA


title: T47D_Her2pos_populations.Rescue.seq.HER2.mut.Jan10_S146 [S146_L755S.DMSO]

value: HER2 L755S over-expression


title: mRNAseq_shCtrl_Rep2

extract_protocol:
Total RNA was extracted from mESCs using TRIzol reagent (Invitrogen) according to manufacturer’s instruction. To remove genomic DNA contamination, RNA samples were treated with DNaseI and purified again by phenol/chloroform extraction and ethanol precipitation.
Libraries were prepared with KAPA stranded mRNA-Seq kits
For ChIP-seq, crosslinked cells were resuspended in swelling buffer (25 mM HEPES pH 7.9, 1.5 mM MgCl2, 10 mM KCl, 0.1% NP-40, and 1X cOmplete protease inhibitor cocktail [Roche]) and incubated for 10 min on ice. Cells were centrifuged and the cell pellet was resuspended in Buffer A (50 mM HEPES-KOH pH 7.9, 140 mM NaCl, 1 mM EDTA, 1% Triton X-100, 0.1% Na-deoxycholate, 0.1% SDS) plus 1X cOmplete protease inhibitor cocktail. Cell sonication was performed on a Qsonica Q800R2 sonicator with 20% amplitude for 20 cycles at 10s each with 30s between cycles at 4oC. Sonicated lysate was pre-cleared by incubating with Dynabeads Protein A/G. Part of the pre-cleared lysate was used as input and the remainder was incubated overnight at 4°C with 2-10 micrograms of antibody. DNA/Protein-antibody conjugates were precipitated using Dynabeads Protein A/G blocked with 5 mg/ml BSA in PBS. Beads were washed twice each with Buffer A, Buffer B (50 mM HEPES-KOH pH 7.9, 500 mM NaCl, 1 mM EDTA, 1% Triton X-100, 0.1% Na-deoxycholate, 0.1% SDS), LiCl buffer (20 mM Tris-HCl pH 8.0, 250 mM LiCl, 1 mM EDTA, 0.5% Na-deoxycholate, 0.5% NP-40) and TE buffer (10 mM Tris-HCl pH 8.0, 1 mM EDTA). DNA was eluted in elution buffer (10 mM Tris-HCl pH 8.0, 1 mM EDTA, 1% SDS). Cross-links were reversed overnight at 65°C. RNA and protein were digested using RNase A and Proteinase K, respectively, and DNA was purified by phenol-chloroform extraction and ethanol precipitation.
For mRNA-seq, total RNA was extracted from mESCs using TRIzol reagent (Invitrogen) according to manufacturer’s instruction. To remove genomic DNA contamination, RNA samples were treated with DNaseI and purified again by phenol/chloroform extraction and ethanol precipitation.
For Nascent RNA-seq, mESCs were harvested, washed with DPBS and lysed in 200 μl of ice-cold lysis buffer (10 mM Tris-HCl pH 7.5, 0.1% NP40, 150 mM NaCl). The cell lysate was gently layered over 500 μl of chilled sucrose cushion (24% RNAse-free sucrose in lysis buffer) in a new Eppendorf tube and centrifuged for 10 min at 4°C, 10,000xg. The supernatant (cytoplasmic fraction) was removed and the pellet (nuclei) was washed once with 200 μl of ice-cold 1 × PBS/1 mM EDTA. The nuclear pellet was resuspended in 100 μl of pre-chilled glycerol buffer (20 mM Tris-HCl pH 7.9, 75 mM NaCl, 0.5 mM EDTA, 0.85 mM DTT, 0.125 mM PMSF, 50% glycerol) by gentle flicking of the tube. An equal volume (100 μl) of cold nuclear lysis buffer (10 mM HEPES pH 7.6, 1 mM DTT, 7.5 mM MgCl2, 0.2 mM EDTA, 0.3 M NaCl, 1 M urea, 1% NP-40) was added. The mix was vortexed vigorously for 2 s. The sample was incubated for 2 min on ice, and then centrifuged for 2 min at 4°C, 10,000xg. The supernatant (nuclear fraction/nucleoplasm) was removed and the pellet (chromatin) was gently rinsed with ice-cold 1×PBS/1 mM EDTA. 1 ml of Trizol reagent was added to the chromatin and incubated for 30 min at 50oC to dissolve it. Nascent RNA was extracted following the manufacturer’s instructions.
For 4C-seq, crosslinked cells were lysed for 15 min on ice in 10 mM Tris-HCl pH 8.0, 10 mM NaCl, and 0.2% NP-40 supplemented with cOmplete protease inhibitor. Nuclei were isolated by centrifugation and by removing the supernatant. Nuclei corresponding to 10 million cells were resuspended in 500 μl of the primary restriction enzyme buffer. SDS was added to a final concentration of 0.3% and samples were incubated for 1 h at 37oC at 1,200 rpm on a thermo-mixer. SDS was quenched by addition of Triton X-100 to a final concentration of 3%. Primary restriction enzyme (400 units) was added and samples were digested for 6 h on a thermo-mixer followed by addition of 400 additional units of the primary restriction enzyme and overnight digestion. The primary restriction enzyme was inactivated by heating to 65oC for 20 min. Samples were diluted to a total volume of 8 ml in ligation buffer (66 mM Tris-HCl pH 7.5, 5 mM MgCl2, 5 mM DTT, 1 mM ATP). Proximity ligation was carried out by adding 4000 units of T4 DNA ligase and incubating at room temperature overnight. After reversal of -crosslinking and RNA removal, DNA was extracted by phenol/chloroform and purified by ethanol precipitation. A secondary digestion was performed overnight in a volume of 500 μl with 200 units of the secondary restriction enzyme. For secondary ligation, following inactivation of the restriction enzyme, each sample was diluted to 14 ml with ligation buffer. T4 DNA ligase (4000 units) was added and samples were incubated overnight at room temperature. DNA was extracted by phenol/chloroform and purified by ethanol precipitation. To remove salts, DNA was further purified using QIAquick PCR Cleanup kit.
For ChIP-seq, libraries were prepared using a KAPA LTP kit following manufacturer's instructions.
For mRNA-seq, libraries were prepared with KAPA stranded mRNA-Seq kits.
For Nascent RNA-seq, libraries were constructed using KAPA Stranded RNA-Seq Kits with RiboErase (HMR).
For 4C-seq libraries construction, reading and non-reading primers were designed for each viewpoint. PCR was performed in sixteen 25-μl PCR reactions using Platinum Taq DNA Polymerase High Fidelity. Reaction conditions were as in the manufacturer’s instructions except 1.4 μM of each primer and 200-400 ng of template were used in each reaction. The PCR program was as follows: (1) 95oC, 5 min; (2) 95oC, 30 s; (3) 55oC, 1 min; (4) 68oC, 3 min; (5) go to (2), 30 cycles; (6) 68oC, 7 min. PCR products were first purified using illustra GFX PCR DNA and Gel Band Purification Kit (GE Healthcare) to remove primer dimers and then further purified using QIAquick PCR Purification Kit to remove salt.

treatment_protocol:
Plasmids expressing shRNA were transfected into mESCs using Lipofectamine 2000 following manufacturer's protocol. Selection with puromycin started 24 h post transfection. After selection for 72 h, cells were harvested for downstream analysis.
For knockdown by siRNA, siRNA was transfected into mESCs using Lipofectamine RNAiMAX following manufacturer's protocol. Cells were harvested and analyzed 72 h post transfection.
For knockdown by shRNA, plasmids expressing shRNA were transfected into mESCs using Lipofectamine 2000 following manufacturer's protocol. Selection with puromycin started 24 h post transfection. After selection for 72 h, cells were harvested for downstream analysis.

value: embryonic stem cells (mESC)

value: shCtrl


title: mRNAseq_shKlf4_Rep2


title: mRNAseq_shKlf4_Rep1


title: Nascent_RNAseq_siCtrl

value: nuclear RNA


title: T47D_Her2pos_populations.Rescue.seq.HER2.mut.Jan10_S110 [S110_wt.DMSO]


title: Control n5

type: RNA

city: Toulouse

first: Philippe

middle: V

last: Rouet

address: 1 Ave Jean Poulhes

department: Cardiology

country: France

institute: INSERM U1048

description: T 5

hyb_protocol: Labeled RNAs were hybridized to human pangenomic glass microarrays from the consortium Reseau National des Genopoles France and Medical Research Council, England, consisting of 25,000 51-mer oligonucleotide probes. The hybridization realized was a standard hybridization.

scan_protocol: The glass array were washed ina Ventana robotized apparatus and scanned using a GenePix 4000 scanner (Axon).

data_processing:
Scanned images were processed by X-dot reader software with operator validation.
Microarray datas for the same gene were summarized to obtain a single value of intensity for every gene. Data normalization was performed in <<R>> (http://www.r-project.org/index.html) and the bioconductor package limma. Two-color GenePix files were normalized within arrays using the loess method and between arrays with Rquantile normalization.Three groups (8 samples for control group, 14 samples for hypertensive group and 14 samples for hypertensive group with left ventricular remodeling) were established and used to calculate p-values by Bayes linear model. Log-ratios were extracted for probes in which the p-value was less than or equal to 0,05. Benjamini & Hochberg test was used for the false discovery rate.

label: Fluorescent labeling (QuickAmp Labeling, Agilent)

source_name: White blood cells_Control

label_protocol: Total RNAs (300 ng) were used for fluorescent labeling (QuickAmp Labeling, Agilent). Fluorescent RNAs were further purified on Rneasy columns. A pooled reference sample was used as a common reference for all hybridizations.

extract_protocol: Total RNA was purified from collected white blood cells using Rneasy kit (Qiagen) in a QIAcube automated sample prep system (Qiagen). Total RNA integrity was checked by capillary electrophoresis (Experion, Bio-Rad). Sample with RNA Quality Indicator ≥ 8,5/10 were selected for analysis. Total RNAs were quantified with RiboGreen on a VictorTM X5 2030multilabel reader (Perkin Elmer).

treatment_protocol: Venous blood samples were collected after overnight fasting in 8 ml BD CPT Vacutainer tubes. White blood cells  separation was processed immediately after blood collection according to the manufacturer's protocol.

tag: subject status

value: control individual

value: white blood cells


title: Control n16

description: T 16


title: Control n12

description: T 12


title: Control n13

description: T 13


title: Control n11

description: T 11


title: Control n6

description: T 6


title: Hypertensive patient with left ventricular remodeling n28

description: R 28

source_name: White blood cells_hypertensive patient with left ventricular remodeling


title: mRNAseq_shCtrl_Rep1


title: T47D_Her2pos_populations.Rescue.seq.HER2.mut.Jan10_S182 [S182_ESR1.DMSO]

description: Parental cell line with ESR1 Y537S over-expression treated with DMSO


title: Control n15

description: T 15


title: Control n3

description: T 3


title: T47D_Her2pos_populations.Rescue.seq.HER2.mut.Jan10_S170 [S170_L869R.DMSO]

description: Parental cell line with HER2 L869R over-expression treated with DMSO

value: HER2 L869R over-expression


title: T47D_Her2pos_populations.Rescue.seq.HER2.mut.Jan10_S218 [S218_D845A.DMSO]


title: Hypertensive patient with left ventricular remodeling n29

description: R 29

value: hypertensive patient with left ventricular remodeling


title: Input_ChIPseq_siEsrrb_Rep2

extract_protocol:
Crosslinked cells were resuspended in swelling buffer (25 mM HEPES pH 7.9, 1.5 mM MgCl2, 10 mM KCl, 0.1% NP-40, and 1X cOmplete protease inhibitor cocktail [Roche]) and incubated for 10 min on ice. Cells were centrifuged and the cell pellet was resuspended in Buffer A (50 mM HEPES-KOH pH 7.9, 140 mM NaCl, 1 mM EDTA, 1% Triton X-100, 0.1% Na-deoxycholate, 0.1% SDS) plus 1X cOmplete protease inhibitor cocktail. Cell sonication was performed on a Qsonica Q800R2 sonicator with 20% amplitude for 20 cycles at 10s each with 30s between cycles at 4oC. Sonicated lysate was pre-cleared by incubating with Dynabeads Protein A/G. Part of the pre-cleared lysate was used as input and the remainder was incubated overnight at 4°C with 2-10 micrograms of antibody. DNA/Protein-antibody conjugates were precipitated using Dynabeads Protein A/G blocked with 5 mg/ml BSA in PBS. Beads were washed twice each with Buffer A, Buffer B (50 mM HEPES-KOH pH 7.9, 500 mM NaCl, 1 mM EDTA, 1% Triton X-100, 0.1% Na-deoxycholate, 0.1% SDS), LiCl buffer (20 mM Tris-HCl pH 8.0, 250 mM LiCl, 1 mM EDTA, 0.5% Na-deoxycholate, 0.5% NP-40) and TE buffer (10 mM Tris-HCl pH 8.0, 1 mM EDTA). DNA was eluted in elution buffer (10 mM Tris-HCl pH 8.0, 1 mM EDTA, 1% SDS). Cross-links were reversed overnight at 65°C. RNA and protein were digested using RNase A and Proteinase K, respectively, and DNA was purified by phenol-chloroform extraction and ethanol precipitation.
Libraries were prepared using KAPA LTP kits.
For ChIP-seq, crosslinked cells were resuspended in swelling buffer (25 mM HEPES pH 7.9, 1.5 mM MgCl2, 10 mM KCl, 0.1% NP-40, and 1X cOmplete protease inhibitor cocktail [Roche]) and incubated for 10 min on ice. Cells were centrifuged and the cell pellet was resuspended in Buffer A (50 mM HEPES-KOH pH 7.9, 140 mM NaCl, 1 mM EDTA, 1% Triton X-100, 0.1% Na-deoxycholate, 0.1% SDS) plus 1X cOmplete protease inhibitor cocktail. Cell sonication was performed on a Qsonica Q800R2 sonicator with 20% amplitude for 20 cycles at 10s each with 30s between cycles at 4oC. Sonicated lysate was pre-cleared by incubating with Dynabeads Protein A/G. Part of the pre-cleared lysate was used as input and the remainder was incubated overnight at 4°C with 2-10 micrograms of antibody. DNA/Protein-antibody conjugates were precipitated using Dynabeads Protein A/G blocked with 5 mg/ml BSA in PBS. Beads were washed twice each with Buffer A, Buffer B (50 mM HEPES-KOH pH 7.9, 500 mM NaCl, 1 mM EDTA, 1% Triton X-100, 0.1% Na-deoxycholate, 0.1% SDS), LiCl buffer (20 mM Tris-HCl pH 8.0, 250 mM LiCl, 1 mM EDTA, 0.5% Na-deoxycholate, 0.5% NP-40) and TE buffer (10 mM Tris-HCl pH 8.0, 1 mM EDTA). DNA was eluted in elution buffer (10 mM Tris-HCl pH 8.0, 1 mM EDTA, 1% SDS). Cross-links were reversed overnight at 65°C. RNA and protein were digested using RNase A and Proteinase K, respectively, and DNA was purified by phenol-chloroform extraction and ethanol precipitation.
For mRNA-seq, total RNA was extracted from mESCs using TRIzol reagent (Invitrogen) according to manufacturer’s instruction. To remove genomic DNA contamination, RNA samples were treated with DNaseI and purified again by phenol/chloroform extraction and ethanol precipitation.
For Nascent RNA-seq, mESCs were harvested, washed with DPBS and lysed in 200 μl of ice-cold lysis buffer (10 mM Tris-HCl pH 7.5, 0.1% NP40, 150 mM NaCl). The cell lysate was gently layered over 500 μl of chilled sucrose cushion (24% RNAse-free sucrose in lysis buffer) in a new Eppendorf tube and centrifuged for 10 min at 4°C, 10,000xg. The supernatant (cytoplasmic fraction) was removed and the pellet (nuclei) was washed once with 200 μl of ice-cold 1 × PBS/1 mM EDTA. The nuclear pellet was resuspended in 100 μl of pre-chilled glycerol buffer (20 mM Tris-HCl pH 7.9, 75 mM NaCl, 0.5 mM EDTA, 0.85 mM DTT, 0.125 mM PMSF, 50% glycerol) by gentle flicking of the tube. An equal volume (100 μl) of cold nuclear lysis buffer (10 mM HEPES pH 7.6, 1 mM DTT, 7.5 mM MgCl2, 0.2 mM EDTA, 0.3 M NaCl, 1 M urea, 1% NP-40) was added. The mix was vortexed vigorously for 2 s. The sample was incubated for 2 min on ice, and then centrifuged for 2 min at 4°C, 10,000xg. The supernatant (nuclear fraction/nucleoplasm) was removed and the pellet (chromatin) was gently rinsed with ice-cold 1×PBS/1 mM EDTA. 1 ml of Trizol reagent was added to the chromatin and incubated for 30 min at 50oC to dissolve it. Nascent RNA was extracted following the manufacturer’s instructions.
For 4C-seq, crosslinked cells were lysed for 15 min on ice in 10 mM Tris-HCl pH 8.0, 10 mM NaCl, and 0.2% NP-40 supplemented with cOmplete protease inhibitor. Nuclei were isolated by centrifugation and by removing the supernatant. Nuclei corresponding to 10 million cells were resuspended in 500 μl of the primary restriction enzyme buffer. SDS was added to a final concentration of 0.3% and samples were incubated for 1 h at 37oC at 1,200 rpm on a thermo-mixer. SDS was quenched by addition of Triton X-100 to a final concentration of 3%. Primary restriction enzyme (400 units) was added and samples were digested for 6 h on a thermo-mixer followed by addition of 400 additional units of the primary restriction enzyme and overnight digestion. The primary restriction enzyme was inactivated by heating to 65oC for 20 min. Samples were diluted to a total volume of 8 ml in ligation buffer (66 mM Tris-HCl pH 7.5, 5 mM MgCl2, 5 mM DTT, 1 mM ATP). Proximity ligation was carried out by adding 4000 units of T4 DNA ligase and incubating at room temperature overnight. After reversal of -crosslinking and RNA removal, DNA was extracted by phenol/chloroform and purified by ethanol precipitation. A secondary digestion was performed overnight in a volume of 500 μl with 200 units of the secondary restriction enzyme. For secondary ligation, following inactivation of the restriction enzyme, each sample was diluted to 14 ml with ligation buffer. T4 DNA ligase (4000 units) was added and samples were incubated overnight at room temperature. DNA was extracted by phenol/chloroform and purified by ethanol precipitation. To remove salts, DNA was further purified using QIAquick PCR Cleanup kit.
For ChIP-seq, libraries were prepared using a KAPA LTP kit following manufacturer's instructions.
For mRNA-seq, libraries were prepared with KAPA stranded mRNA-Seq kits.
For Nascent RNA-seq, libraries were constructed using KAPA Stranded RNA-Seq Kits with RiboErase (HMR).
For 4C-seq libraries construction, reading and non-reading primers were designed for each viewpoint. PCR was performed in sixteen 25-μl PCR reactions using Platinum Taq DNA Polymerase High Fidelity. Reaction conditions were as in the manufacturer’s instructions except 1.4 μM of each primer and 200-400 ng of template were used in each reaction. The PCR program was as follows: (1) 95oC, 5 min; (2) 95oC, 30 s; (3) 55oC, 1 min; (4) 68oC, 3 min; (5) go to (2), 30 cycles; (6) 68oC, 7 min. PCR products were first purified using illustra GFX PCR DNA and Gel Band Purification Kit (GE Healthcare) to remove primer dimers and then further purified using QIAquick PCR Purification Kit to remove salt.

value: siEsrrb


title: mRNAseq_siCtrl_Rep1

value: siCtrl


title: Hypertensive patient with left ventricular remodeling n26

description: R 26


title: T47D_Her2pos_populations.Rescue.seq.HER2.mut.Jan10_S206 [S206_wt.DMSO]


title: Hypertensive patient with left ventricular remodeling n20

description: R 20


title: Input_ChIPseq_siCtrl_Rep2


title: Input_ChIPseq_siCtrl_Rep1


title: mRNAseq_siCtrl_Rep2


title: mRNAseq_siEsrrb_Rep2


title: mRNAseq_siEsrrb_Rep1


title: Hypertensive patient with left ventricular remodeling n23

description: R 23


title: T47D_Her2pos_populations.Rescue.seq.HER2.mut.Jan10_S145 [S145_L755S.DMSO]


title: Input_ChIPseq_WT

treatment_protocol:
none
For knockdown by siRNA, siRNA was transfected into mESCs using Lipofectamine RNAiMAX following manufacturer's protocol. Cells were harvested and analyzed 72 h post transfection.
For knockdown by shRNA, plasmids expressing shRNA were transfected into mESCs using Lipofectamine 2000 following manufacturer's protocol. Selection with puromycin started 24 h post transfection. After selection for 72 h, cells were harvested for downstream analysis.

value: wild type


title: T47D_Her2pos_populations.Rescue.seq.HER2.mut.Jan10_S25 [S25_D845A.DMSO]

value: HER2 D845A over-expression


title: Smc1_ChIPseq_WT

tag: chip antibody

value: Smc1 (Bethyl, A300-055A)


title: Med26_ChIPseq_siCtrl_Rep2

value: Med26 (Santa Cruz, sc-1521)


title: Hypertensive patient with left ventricular remodeling n22

description: R 22


title: TAF2_ChIPseq_siCtrl_Rep1

value: TAF2 (Novus, NBP1-21371)


title: TAF2_ChIPseq_siCtrl_Rep2


title: Hypertensive patient with left ventricular remodeling n18

description: R 18


title: T47D_Her2pos_populations.Rescue.seq.HER2.mut.Jan10_S181 [S181_ESR1.DMSO]

value: ESR1 Y537S over-expression

value: DMSO


title: Esrrb_ChIPseq_siEsrrb_Rep2

value: Esrrb (Perseus Proteomics,PP-H6705-00)


title: Esrrb_ChIPseq_siEsrrb_Rep1

value: embryonic stem cells (mESC)


title: Esrrb_ChIPseq_siCtrl_Rep2


title: T47D_Her2pos_populations.Rescue.seq.HER2.mut.Jan10_S277 [S277_ESR1.DMSO]


title: Esrrb_ChIPseq_siCtrl_Rep1


title: TFIIB_ChIPseq_siEsrrb_Rep1

value: Carey Lab


title: TFIIB_ChIPseq_siEsrrb_Rep2


title: T47D_Her2pos_populations.Rescue.seq.HER2.mut.Jan10_S217 [S217_D845A.DMSO]


title: Med26_ChIPseq_siCtrl_Rep1


title: TFIIB_ChIPseq_siCtrl_Rep2


title: T47D_Her2pos_populations.Rescue.seq.HER2.mut.Jan10_S157 [S157_V777L.DMSO]

value: HER2 V777L over-expression


title: T47D_Her2pos_populations.Rescue.seq.HER2.mut.Jan10_S133 [S133_S653C.DMSO]

description: Parental cell line with HER2 S653C over-expression treated with DMSO

value: HER2 S653C over-expression


title: TFIIB_ChIPseq_siCtrl_Rep1


title: Hypertensive patient with left ventricular remodeling n11

description: R 11

value: white blood cells


title: Pol II_ChIPseq_siEsrrb_Rep2

source_name: Pol II_ChIPseq_siEsrrb

value: Pol II (QED Bioscience, 70101)


title: Hypertensive patient with left ventricular remodeling n15

description: R 15

value: hypertensive patient with left ventricular remodeling


title: T47D_Her2pos_populations.Rescue.seq.HER2.mut.Jan10_S241 [S241_L755S.DMSO]

value: HER2 L755S over-expression


title: Pol II_ChIPseq_siCtrl_Rep1

source_name: Pol II_ChIPseq_siCtrl

value: siCtrl


title: Pol II_ChIPseq_siEsrrb_Rep1

value: siEsrrb


title: T47D_Her2pos_populations.Rescue.seq.HER2.mut.Jan10_S73 [S73_L869R.DMSO]

value: HER2 L869R over-expression


title: Hypertensive patient with left ventricular remodeling n5

description: R 5


title: Pol II_ChIPseq_siCtrl_Rep2


title: Hypertensive patient with left ventricular remodeling n14

description: R 14


title: Hypertensive patient with left ventricular remodeling n12

description: R 12


title: Hypertensive patient with left ventricular remodeling n4

description: R 4


title: Hypertensive patient with left ventricular remodeling n2

description: R 2


title: T47D_Her2pos_populations.Rescue.seq.HER2.mut.Jan10_S121 [S121_D845A.DMSO]

value: HER2 D845A over-expression


title: T47D_Her2pos_populations.Rescue.seq.HER2.mut.Jan10_S265 [S265_L869R.DMSO]


title: Med12_ChIPseq_siEsrrb_Rep1

value: Med12 (Abcam, Ab70842)


title: Med12_ChIPseq_siCtrl_Rep2


title: Hypertensive patient with normal left ventricular size n29

description: H 29

source_name: White blood cells_hypertensive patient

value: hypertensive patient with normal left ventricular size


title: Hypertensive patient with normal left ventricular size n31

description: H 31


title: Hypertensive patient with normal left ventricular size n30

description: H 30


title: Med12_ChIPseq_siCtrl_Rep1


title: T47D_Her2pos_populations.Rescue.seq.HER2.mut.Jan10_S13 [S13_wt.DMSO]

value: HER2 wt over-expression


title: Med1_ChIPseq_siEsrrb_Rep1

value: Med1 (Santa Cruz, sc-5334)


title: Med1_ChIPseq_siEsrrb_Rep2


title: Med12_ChIPseq_siEsrrb_Rep2


title: T47D_Her2pos_populations.Rescue.seq.HER2.mut.Jan10_S253 [S253_V777L.DMSO]

value: DMSO


title: Med1_ChIPseq_siCtrl_Rep2

value: embryonic stem cells (mESC)


title: Hypertensive patient with normal left ventricular size n27

description: H 27


title: T47D_Her2pos_populations.Rescue.seq.HER2.mut.Jan10_S37 [S37_S653C.DMSO]


title: Day 1 CLP 2 [17477]

city: Iowa City

first: Isaac

middle: J

last: Jensen

state: IA

institute: University of Iowa

data_processing:
The sequencing quality of RNA-Seq libraries was assessed by FastQC v0.10.1 (http://www.bioinformatics.babraham.ac.uk/projects/fastqc/)
RNA-Seq libraries were mapped to mouse genome using Tophat (v2.1.0, Trapnell et al., Bioinformatics). UCSC genes for mouse mm9 from iGenome (m9 from iGenome (http://support.illumina.com/sequencing/sequencing_software/igenome.html) were used for gene annotation.
Gene expressions were calculated with featureCounts, a summarization program suitable for counting reads generated from RNASeq, Expression is given as DESeq2 values with muliple test correction of FDR to identify differentially expressed genes
Genome_build: mm9
Supplementary_files_format_and_content: Excel file includes DESeq2 values for each sample

source_name: Day 1 CLP_NK cells

extract_protocol:
NK cells were sort-purified and total RNA was extracted using standard Qiagen Rnease columns
RNA libraries were prepared for sequencing using standard Illumina protocol

tag: mouse group

value: cecal ligation and puncture (CLP) surgery

tag: day post-surgery

value: Splenic NK cells

tag: cell phenotype


title: Day 2 CLP 2 [17481]

source_name: Day 2 CLP_NK cells

tag: strain


title: Day 1 CLP 1 [17476]


title: Day 2 CLP 1 [17480]


title: Hypertensive patient with normal left ventricular size n24

description: H 24


title: Day 2 SHAM 1 [17474]

source_name: Day 2 SHAM_NK cells

value: Sham (control)


title: Med1_ChIPseq_siCtrl_Rep1


title: T47D_Her2pos_populations.Rescue.seq.HER2.mut.Jan10_S49 [S49_L755S.DMSO]


title: Day 2 SHAM 2 [17475]


title: T47D_Her2pos_populations.Rescue.seq.HER2.mut.Jan10_S229 [S229_S653C.DMSO]

value: HER2 S653C over-expression


title: T47D_Her2pos_populations.Rescue.seq.HER2.mut.Jan10_S85 [S85_ESR1.DMSO]

value: ESR1 Y537S over-expression


title: T47D_Her2pos_populations.Rescue.seq.HER2.mut.Jan10_S205 [S205_wt.DMSO]


title: T47D_Her2pos_populations.Rescue.seq.HER2.mut.Jan10_S169 [S169_L869R.DMSO]


title: Hypertensive patient with normal left ventricular size n22

description: H 22

value: white blood cells


title: T47D_Her2pos_populations.Rescue.seq.HER2.mut.Jan10_S97 [S97_GFP.DMSO]

value: GFP over-expression


title: Hypertensive patient with normal left ventricular size n20

description: H 20


title: T47D_Her2pos_populations.Rescue.seq.HER2.mut.Jan10_S193 [S193_GFP.DMSO]


title: T47D_Her2pos_populations.Rescue.seq.HER2.mut.Jan10_S61 [S61_V777L.DMSO]

value: HER2 V777L over-expression


title: Hypertensive patient with normal left ventricular size n9

description: H 9


title: Hypertensive patient with normal left ventricular size n18

description: H 18


title: Hypertensive patient with normal left ventricular size n11

description: H 11


title: Hypertensive patient with normal left ventricular size n5

description: H 5


title: Hypertensive patient with normal left ventricular size n10

description: H 10


title: NSP2 injected mammary tissue, in-vivo 12 weeks, biological rep2

city: Memphis

first: Evan

middle: M.

last: Savage

state: TN

address: 1910 Nonconnah Blvd., Suite

institute: Genome Explorations

description: Gene expression data from NSP2-injected mammary tissue after 12 weeks

hyb_protocol: Fragmented cDNA was hybridized for 17hr at 45oC to GeneChip® MoGene 1.0 ST Arrays (Affymetrix, Santa Clara CA). GeneChips were washed and stained in Affymetrix Fuildics Station 450 instruments according to manufacturer's protocol

scan_protocol: GeneChips were scanned using an Affymetrix GeneChipScanner 3000 7G high-resolution confocal laser scanner

data_processing: Expression data was processed using background correction, quantile normalization and probe set signal summarization by Robust Multi-Array algorithm in Affymetrix Expression Console according to manufacturer's protocol

label: biotin

source_name: in-vitro NSP2 injected mammary tissue after 12 weeks

label_protocol: Total RNA was processed by the random primed RT-IVT-RT method using the GeneChip WT cDNA Synthesis and Amplification Kit (Affymetrix, Santa Clara CA), according to the manufacturer’s instructions. Amplified cDNA was fragmented and labeled with biotin using the GeneChip WT Terminal Labeling Kit (Affymetrix, Santa Clara CA), according to the manufacturer’s instructions.

extract_protocol:
RNA Isolation/extraction from cultured cells: Cells were washed in PBS, then lysed with buffer RLT and 10 μl/ml β-mercaptoethanol, using 350 μl per T25 flask or 600 μl per 10 cm plate. The lysate was collected with a cell scraper, placed into an RNase-free tube, and homogenized by applying it to a QIAshredder spin column (Qiagen 79654), and spinning for 2 min at 13,000 rpm. The homogenized lysate could be stored at -80°C, or used for the RNA isolation. The RNeasy Mini protocol was followed, including on-column incubation with RNase-free DNase (Qiagen 79254). After the appropriate washes, RNA was eluted with 30 μl RNase-free water and the concentration was determined using a Nanodrop spectrophotometer. RNA was stored at -80°C. Each sample was isolated and used individually, in triplicate.
RNA Isolation/extraction from tissue: RNAlater-ICE (Ambion AM7030) was chilled in polypropylene tubes at -80°C or on dry ice. Each mouse was individually euthanized, and sprayed with RNase Zap before quickly removing mammary tissue and placing it directly into liquid nitrogen. Frozen tissues were then soaked in chilled RNAlater-ICE at -20°C for at least 16 hr to enable complete penetration. Tissues were weighed and cut to ≤100 mg. RNA was isolated according to the RNeasy Lipid Tissue Kit Mini protocol (Qiagen 74804), including on-column DNase treatment. RNA was eluted with 30 μl RNase-free water and the concentration was determined using a Nanodrop spectrophotometer. RNA was stored at -80°C.  Each sample was isolated and used individually, in triplicate.

treatment_protocol: sample prep protocol: CDβ, SP3, NSP2 and normal Balb/C MEC were cultured in identical tissue culture medium to provide in-vitro cell samples. 20,000 cells from each of in vitro cell culture was suspended in 10ml of DMEM without serum and injected into the epithelium-cleared inguinal mammary fat pads of Nu/Nu 3-week-old females. After injection into Nu/Nu mammary fat-pads, three replicate in-vivo samples were collected from each culture (CDb, SP3, NSP2 and normal Balb/C) after 5 and 12 weeks, respectively.

value: NSP2-injected mammary tissue

tag: vivo/vitro

value: in-vivo

tag: weeks


title: NSP2 injected mammary tissue, in-vivo 12 weeks, biological rep3


title: Hypertensive patient with normal left ventricular size n2

description: H 2


title: T47D_Her2pos_populations.Rescue.seq.HER2.mut.Jan10_S109 [S109_wt.DMSO]

tag: genotype/variation


title: CommaD cultered cells, in-vivo 12 weeks, biological rep1

description: Gene expression data from CommaD-injected mammary tissue after 12 weeks

source_name: in-vitro CommaD injected mammary tissue after 12 weeks

value: CommaD

tag: tissue

value: CommaD-injected mammary tissue


title: SP3 injected mammary tissue, in-vivo 5 weeks, biological rep3

description: Gene expression data from SP3-injected mammary tissue after 5 weeks

source_name: in-vitro SP3 injected mammary tissue after 5 weeks

tag: cell line

value: SP3-injected mammary tissue


title: Hypertensive patient with normal left ventricular size n3

description: H 3

value: hypertensive patient with normal left ventricular size


title: BalbC injected mammary tissue, in-vivo 12 weeks, biological rep2

description: Gene expression data from BalbC-injected mammary tissue after 12 weeks

source_name: in-vitro Balbc injected mammary tissue after 12 weeks

value: normal BalbC MEC

value: BalbC-injected mammary tissue


title: T47D_Her2pos_populations.Rescue.seq.HER2.mut.Jan10_S1 [S1_GFP.DMSO]

source_name: Breast Cancer

tag: agent

value: DMSO


title: NSP2 injected mammary tissue, in-vivo 12 weeks, biological rep1


title: SP3 injected mammary tissue, in-vivo 12 weeks, biological rep1

description: Gene expression data from SP3-injected mammary tissue after 12 weeks

source_name: in-vitro SP3 injected mammary tissue after 12 weeks


title: SP3 injected mammary tissue, in-vivo 12 weeks, biological rep3


title: CommaD cultered cells, in-vivo 12 weeks, biological rep3


title: BalbC injected mammary tissue, in-vivo 12 weeks, biological rep1


title: BalbC injected mammary tissue, in-vivo 12 weeks, biological rep3


title: SP3 injected mammary tissue, in-vivo 5 weeks, biological rep2


title: SP3 injected mammary tissue, in-vivo 5 weeks, biological rep1


title: NSP2 injected mammary tissue, in-vivo 5 weeks, biological rep3

description: Gene expression data from NSP2-injected mammary tissue after 5 weeks

source_name: in-vitro NSP2 injected mammary tissue after 5 weeks


title: NSP2 injected mammary tissue, in-vivo 5 weeks, biological rep2


title: SP3 injected mammary tissue, in-vivo 12 weeks, biological rep2


title: CommaD cultered cells, in-vivo 12 weeks, biological rep2


title: CommaD cultered cells, in-vivo 5 weeks, biological rep2

description: Gene expression data from CommaD-injected mammary tissue after 5 weeks

source_name: in-vitro CommaD injected mammary tissue after 5 weeks


title: NSP2 injected mammary tissue, in-vivo 5 weeks, biological rep1


title: BalbC injected mammary tissue, in-vivo 5 weeks, biological rep3

description: Gene expression data from BalbC-injected mammary tissue after 5 weeks

source_name: in-vitro Balbc injected mammary tissue after 5 weeks


title: KO_3

city: Vienna

first: Nadine

middle: Elpida

last: Tatto

address: Muthgasse 18

department: Department of Biotechnology

country: Austria

institute: University of Natural Resources and Life Sciences

data_processing:
Conversion BAM → FASTQ with SamToFastq (picard v2.17.3-3-g3bf7fdc-SNAPSHOT).
Quality Control with FastQC v0.11.2 with UGENE workflow designer (Unipro UGENE IDE v1.9.8).
Read trimming with TrimGalore v0.4.2.
Read quantification with kallisto v0.44.0 and samtools v1.5 (kallisto index based on K. pastoris (aka K. phaffii) CBS7435 transcript sequences as reference).
Differential expression analysis with R (DESeq2).
Processed RNASeq data with gene annotations and statistically significant fold-changes are provided as "Supplementary File 2" with the manuscript.
Genome_build: ASM170810v1
Supplementary_files_format_and_content: *.tsv: Tab-separated text files include raw read counts.
Supplementary_files_format_and_content: *_normalizedCounts.csv: Comma-separated text files include normalized counts (DESeq2).

organism: Komagataella phaffii

source_name: Δgal4-like cells

growth_protocol: All shake flask experiments were performed in 200 ml wide neck shake flasks including 20 mL M2 medium containing (g L-1) 5 glucose, 3.15 (NH4)2HPO4; 0.49 MgSO4.7H2O; 0.8 KCl; 0.0268 CaCl2.2H2O; 22 citric acid monohydrate; 1.47 mL PTM1; 2 mL biotin (0.2 g L-1). For the sampling of the RNASeq analysis, cells grown in YP medium including 25 μg mL-1 Zeocin for 24-26 h at 25°C were washed and transferred into M2 medium. Samples were collected during the exponential growth phase for the characterization of the strains. Cultivations were done in triplicates.

extract_protocol:
Pellets from the samples collected during the exponential growth phase were resuspended in 1 mL TRI Reagent (Sigma, USA). Cells were disrupted by glass beads using a ribolyser (MP Biomedicals, USA) for 40 sec at 5.5 ms-1. RNA was extracted by chloroform and precipitated with isopropanol. After washing twice with 75% ethanol, RNA was dissolved in nuclease-free water. After DNase treatment (Invitrogen, USA), concentration of RNA was measured by Nanodrop and quality of the samples was verified with the Agilent Bioanalyzer 2100 using the RNA 6000 Nano Assay kit (Agilent Technologies, California).
RNASeq analysis was performed with poly(A)-enriched mRNAs at Vienna Biocenter Core Facilities GmbH (AT). cDNA libraries were constructed from these poly(A)-enriched mRNAs and sequenced by Illumina HiSeq v4.

tag: strain/background

tag: medium

value: glucose


title: NSP2 cultured cells, in-vitro, biological rep2

description: Gene expression data from NSP2 cell line

source_name: in-vitro cultured NSP2 cells

value: cultured cell line

value: in-vitro


title: SP3 cultured cells, in-vitro, biological rep1

description: Gene expression data from SP3 cell line

source_name: in-vitro cultured SP3 cells


title: BalbC cultured cells, in-vitro, biological rep2

description: Gene expression data from BalbC cell line

source_name: in-vitro cultured Balbc cells


title: BalbC injected mammary tissue, in-vivo 5 weeks, biological rep2


title: NSP2 cultured cells, in-vitro, biological rep3


title: SP3 cultured cells, in-vitro, biological rep3

description: Gene expression data from SP3 cell line


title: NSP2 cultured cells, in-vitro, biological rep1


title: CommaD cultered cells, in-vivo 5 weeks, biological rep1


title: CommaD cultered cells, in-vitro, biological rep1

description: Gene expression data from CommaD cell line

source_name: in-vitro cultured CommaD cells


title: CommaD cultered cells, in-vitro, biological rep2


title: SP3 cultured cells, in-vitro, biological rep2

description: Gene expression data from SP3 cell line


title: 14-053O7_LN

city: Seattle

first: Ilsa

last: Coleman

state: WA

address: 1100 Fairview Ave N, E2-112

department: Human Biology

institute: Fred Hutchinson Cancer Center

description: 13-101H1 CRPC lymph node (LN) metastasis

data_processing:
Image analysis and base calling were performed using Illumina's Real Time Analysis v1.18.X software, followed by 'demultiplexing' of indexed reads and generation of FASTQ files, using Illumina's bcl2fastq Conversion Software v1.8.4
Reads were mapped to the hg19 human genome build using TopHat v2.X and bowtie2 v2.2.X.
Mean and standard deviation of fragment size were calculated with picard-tools v2.18.1
Transcript abundance was determined from the TopHat alignments in R using the Genomic Alignments Bioconductor package v1.12.2 using the summarizeOverlaps function with mode=IntersectionStrict, counting reads mapping to the exonic regions of genes.

source_name: CRPC lymph node (LN) metastasis

extract_protocol:
Total RNA was isolated from sections of frozen tissue using RNA STAT-60 (Tel-Test) while incorporating Phase Lock Gel Heavy tubes (5 Prime) into the protocol to better facilitate phase separation.   RNA yield and purity was obtained on a NanoDrop 2000 (Thermo Fisher Scientific).  RNA Integrity was evaluated on a 2100 Bioanalyzer (Agilent Technologies).
RNA-seq libraries were constructed from 1 ug total RNA using the Illumina TruSeq Stranded mRNA LT Sample Prep Kit according to the manufacturer’s protocol.
Barcoded libraries were pooled and sequenced six per lane on the Illumina HiSeq 2500 generating 50 bp paired end reads.

tag: gender

value: male

value: Castration Resistant Prostate Cancer (CRPC)

tag: tumor type

value: CRPC metastasis

tag: tumor site

value: lymph node

tag: patient


title: Input_ChIPseq_siEsrrb_Rep1

tag: cell type

value: embryonic stem cells (mESC)

value: siEsrrb


title: DoxOFF_day4 rep2

city: Kamakura

first: Satoshi

last: Kondo

state: Kanagawa

address: Tebiro 6-10-1

department: New Projects Development Division

country: Japan

institute: Toray Industries,Inc.

hyb_protocol: Hybridized for 16 h at 37 C with rotary shake (250 rpm). Hybridization buffer and washing protocol was followed by the protocol supplied by TORAY Industries, Inc..

scan_protocol: 3D-Gene Scanner ((Toray Industries Inc., Tokyo, Japan) was used for scanning. Images were quantified using Extraction(Toray Industries Inc., Tokyo, Japan).

data_processing: The raw data of each spot was normalized by substitution with a mean intensity of the background signal determined by all blank spots’ signal intensities of 95% confidence intervals.

source_name: embryoid body

label_protocol: Amino Allyl aRNA was synthesis by Amino Allyl MessageAmp II aRNA Amplification Kit (Ambion#1753). CyeDye Coupling and fragmentation were performed as the protocol supplied by TORAY Industries, Inc..

growth_protocol: The cells were suspended in modified serum-free differentiation (SFD) medium without any additional growth factors for 2 days.  Subsequently, the EBs were dissociated and the cells were reaggregated in the SFD medium for 2days. For the transcription factor-based cardiac differentiation, Dox was added into the SFD medium as indicated.

extract_protocol: RNA samples were extracted from cells using RNesy mini kit (Quiagen).

value: embryoid body


title: KO_2


title: DoxOFF_day2 rep2


title: DoxOFF_day4 rep1


title: KO_1


title: DoxOn_day4 rep1


title: DoxOFF_day2 rep1


title: BalbC injected mammary tissue, in-vivo 5 weeks, biological rep1

value: in-vivo


title: CommaD cultered cells, in-vitro, biological rep3

value: CommaD


title: DoxOn_day2 rep1


title: BalbC cultured cells, in-vitro, biological rep3


title: BalbC cultured cells, in-vitro, biological rep1


title: DoxOn_day2 rep2


title: DoxOn_day4 rep2


title: OE_3

source_name: Gal4-like overexpression cells

value: Gal4-like overexpression


title: OE_2


title: OE_1


title: Control_2

source_name: Control cells

value: Control


title: input DNA ccq1, rep2

city: Planegg

first: Sigurd

last: Braun

address: Großhaderner Str. 9

department: BMC

country: Germany

institute: LMU

data_processing:
Base calling with RTA1.18.64.
Sequenced reads were trimmed for the adaptor sequence, then mapped to the S. pombe genome with Novoalign (http://www.novocraft.com) randomly assigned.
Reads mapping with two or fewer mismatches were retained.
Enrichments were calculated with our house Perl scripts.
Normalisation to reads per million.
Genome_build: Schizosaccharomyces_pombe.ASM294v1.18 + pNSU70
Supplementary_files_format_and_content: *.igv.tdf files can be viewed with IGV browser: http://www.broad.mit.edu/igv

organism: Schizosaccharomyces pombe

source_name: ccq1D input

growth_protocol: Strains were grown in logarithmic phase in standard pombe media (YE+5Supplements, 3% glucose).

extract_protocol:
Flash frozen cell pellets (approx. 1.2 billion cells) were homogenized (Precellys 24, Peqlab), sonicated (Q800R, Qsonica, 30 min), supernatant was incubated O/N with antibody, incubated with beads (ProtG/Dynabeads, Invitrogen). After washes (2x Lysis buffer (50mM HEPES/KOH pH7.5, 140 mM NaCl, 1 mM EDTA, 1% TritonX-100, 0.1NA-DeoXycholate), 2x high salt buffer (50mM HEPES/KOH pH7.5, 500 mM NaCl, 1 mM EDTA, 1% TritonX-100, 0.1NA-DeoXycholate), 2x wash buffer (10 mM Tris/HCl pH 8, 250 mM LiCl , 1 mM EDTA, 0.5% TritonX-100, 0.5 Na-DeoXycholate)), samples were decrosslinked (4h, 65C), treated with 10ul ProtK (4mg/ml) and purified (zymo Research D5201).
Libraries were prepared according to manufacturer's instruction (NEB E7645S) with pooled samples of three independent experiments.

treatment_protocol: Samples were crosslinked (1%) formaldehyde for 10 min at RT.

tag: chip antibody


title: input DNA ccq1, rep1


title: input DNA wt, rep2

source_name: Wild type input

value: Wild type


title: H3_ChIPseq_rik1

source_name: rik1D H3 ChIP

value: H3-Antibody (Active Motif 61475)


title: input DNA wt, rep1


title: H3_ChIPseq_mit1

source_name: mit1D H3 ChIP


title: input DNA rik1

source_name: rik1D input


title: H3_ChIPseq_ccq1, rep2

source_name: ccq1D H3 ChIP


title: H3_ChIPseq_ccq1, rep1


title: H3_ChIPseq_wt, rep2

source_name: Wild type H3 ChIP


title: Control_3


title: H3_ChIPseq_wt, rep1


title: input DNA mit1

source_name: mit1D input


title: Control_1


title: 12-021J3_PLEURAL

description: 16-071H10 CRPC pleural metastasis

source_name: CRPC pleural metastasis

value: pleural


title: huoxue_muscle_3

city: harbin

first: Xiaoting

last: Li

address: haerbindajie

country: China

institute: Biotechnology

description: Formula of SHENQI compound

hyb_protocol: Each Slide was hybridized with 1.65g Cy3-labeled cRNA using Gene Expression Hybridization Kit (Cat#5188-5242, Agilent technologies, Santa Clara, CA, US) in Hybridization Oven (Cat#G2545A, Agilent technologies, Santa Clara, CA, US), according to the manufacturers instructions.

scan_protocol: Slides were scanned by Agilent Microarray Scanner (Cat#G2565CA, Agilent technologies, Santa Clara, CA, US) with default settings, Dye channel: Green, Scan resolution=5m, PMT 100%, 10%, 16bit.

data_processing: Data were extracted with Feature Extraction software 10.7 (Agilent technologies, Santa Clara, CA, US). Raw data were normalized by Quantile algorithm, Gene Spring Software 11.0 (Agilent technologies, Santa Clara, CA, US).

label_protocol: Low Input Quick Amp Labeling Kit

extract_protocol: Total RNA was amplified and labeled by Low Input Quick Amp Labeling Kit, One-Color (Cat#5190-2305, Agilent technologies, Santa Clara, CA, US), following the manufacturers instructions. Labeled cRNA were purified by RNeasy mini kit (Cat#74106, QIAGEN, GmBH, Germany).

value: Skeletal muscle


title: rosiglitazone_aorta_3

value: aorta


title: huoxue_muscle_2

description: Formula of SHENQI compound


title: rosiglitazone_aorta_2


title: rosiglitazone_muscle_2


title: rosiglitazone_muscle_1


title: Slide US45102948_256525610010_S03.7

city: STOCKHOLM

first: Sulman

last: Shafeeq

address: nobel vag 16

department: MTC

country: Sweden

institute: Karolinska Institutet

description: Sample 1

hyb_protocol: The protocol was performed as described in Kloosterman et al., 2006 and Shafeeq et al., 2011.

scan_protocol: Scanning was done using the Genepix 4200AL laser scanner.

data_processing: Dual-channel array images were analyzed with Genepix 6.  Spots were screened visually to identify those of low quality. Slide data were processed with MicroPreP software. Prior to analysis, automatically and manually flagged spots and spots with very low background subtracted signal intensity (5% of the weakest spots [sum of Cy3 and Cy5 net signals]) were filtered out of all data sets. Net signal intensities were calculated by grid-based background subtraction. A grid-based Lowess transformation was performed for slide normalization, negative and empty values were removed, and outliers were removed by the deviation test. Expression ratios  were calculated. Further analysis was performed with a Cyber-T Student T-test for paired data.

organism: Streptococcus pneumoniae D39

source_name: F635 Mean

label_protocol: Total RNA was incubated for 16 h at 42°C in the presence of 400 U Superscript III RNase H reverse transcriptase (Invitrogen), 0.2 mM aminoallyl dUTP (Amersham), 0.5 mM dATP, 0.5 mM dCTP, 0.5 mM dGTP, 0.3 mM dTTP, and 3.2 g random nonamers. The synthesized cDNA was then labeled by coupling either Cy3 or Cy5 to the dUTP aminoallyl reactive group (CyScribe postlabeling kit; Amersham Biosciences) and was purified using a GFX PCR, DNA, and gel band purification kit (Amersham Biosciences).

growth_protocol: Cells were grown in CDM supplemented either with 55mM mannose. Cells were grown until early exponential phase. This corresponded to OD600 of 0.3.

extract_protocol: RNA was isolated and purified using the High Pure RNA isolation kit (Roche diagnostics) as described (Kloosterman et al., 2006). Contaminating genomic DNA was removed by treatment with RNase-free DNase I (Roche diagnostics). RNA was isolated from three replicate cultures.

tag: genotype

value: wild-type

source_name: F532 Mean

value: spd_0939 mutant


title: Slide US45102948_256525610010_S03.8

description: Sample 2


title: rosiglitazone_muscle_3


title: huoxue_aorta_3

description: Formula of SHENQI compound


title: rosiglitazone_aorta_1


title: huoxue_muscle_1

description: Formula of SHENQI compound


title: Slide US45102948_256525610010_S02.7

description: Sample 1

growth_protocol: Cells were grown in CDM supplemented either with 55mM galactose. Cells were grown until early exponential phase. This corresponded to OD600 of 0.3.


title: Slide US45102948_256525610010_S02.8

description: Sample 2


title: huoxue_aorta_2

description: Formula of SHENQI compound


title: Slide US45102948_256525610010_S03.6

description: Sample 2

growth_protocol: Cells were grown in CDM supplemented  with 55 mM mannose. Cells were grown until early exponential phase. This corresponded to OD600 of 0.3.

value: spd_0144 mutant


title: Slide US45102948_256525610010_S02.6

description: Sample 2

growth_protocol: Cells were grown in CDM supplemented  with 55 mM Galactose. Cells were grown until early exponential phase. This corresponded to OD600 of 0.3.


title: Slide US45102948_256525610010_S02.5

description: Sample 1


title: yangyinyiqi_muscle_3

description: Formula of SHENQI compound


title: huoxue_aorta_1

description: Formula of SHENQI compound


title: Donor 6 NAC treated

city: Rozzano, Milan

first: Emilia Maria Cristina

last: Mazza

address: Via Alessandro Manzoni, 56

country: Italy

institute: Humanitas Clinical and Research Center

data_processing:
Basecall was performed using CASAVA version 1.8
RNA-seq reads were aligned to the GRCh38 genome assembly using STAR version 2.5.1b
Raw gene counts were generated from STAR output and normalized in TPM counts
Differential Gene Expression analysis was performed using EdgeR 3.22.1 R version 3.3.3 . False discovery rate < 0.05
Genome_build: GRCh38
Supplementary_files_format_and_content: raw_counts

source_name: human CD8 T cells_NAC treated

growth_protocol: human naive T cells from human healty donors were put in culture for 8 days in presence of culture medium and interleukins 2 and 12 with CD3/CD28 antibody-conjugated  beads

extract_protocol:
Total RNA extraction with RNeasy Micro Kit (Qiagen), 2 ng of total RNA were udes for the construction of sequencing libraries.
RNA libraries were prepared for sequencing using Nextera XT DNA Library Preparation Kit

tag: donor id

value: healty donor 6

value: human CD8 T cells

tag: treated with

value: N-acetyl cysteine (NAC)


title: Donor 5 NAC treated

value: healty donor 5


title: yangyinyiqi_aorta_1

description: Formula of SHENQI compound


title: yangyinyiqi_muscle_1

description: Formula of SHENQI compound


title: yangyinyiqi_muscle_2

description: Formula of SHENQI compound


title: Donor 4 NAC treated

value: healty donor 4


title: DNA_Soil_Riparian_wet 1

type: genomic

city: Princeton

first: Bess

middle: B

last: Ward

state: NJ

address: Washington Road

department: Geosciences

institute: Princeton University

description: environmental DNA extracted from agricultural soil

hyb_protocol: Samples were hybridized to duplicate arrays by overnight incubation at 64°C and washed.

scan_protocol: The arrays were scanned with a laser scanner (Molecular Devices 4300) Images were analyzed with Gene Pix Pro 6.0 software (Molecular Devices)

data_processing: Quality control was employed to remove noise by eliminating features in which the Cy3 signal was not at least twice the value of the control probes.

organism: synthetic construct

source_name: 70-mer probe

label_protocol: The genomic DNA was amplified with PCR primers NOSZ1R and NOSZ1F, and labeled with amino-allyl-dUTP (Ambion) during linear amplification using random octamers and a Klenow polymerase (Applied Biosystems). The reaction contained 3.96 mM d(AGC)TP, 0.44 mM dTTP, and 4.84 mM dUaa and was amplified for 3 h at 37°C. The Klenow product was purified and conjugated with Cy3. The Cy3-labelled target (200 ng) was combined with 2x hybridization buffer (1x final concentration; Agilent) and 0.25 pmol of a Cy5-labelled complementary 20-mer standard oligonucleotide and incubated at 95°C for 5 min before being cooled to room temperature.

extract_protocol: DNA was extracted from the filters using the PureGene DNA kit (Gentra, Minneapolis, MN) or the AllPrep DNA/RNA Mini Kit (Qiagen Sciences, Maryland, USA) with slight modifications (as in Ward 2008).

tag: sample type

value: 20-mer DNA oligo

organism: soil metagenome

source_name: PCR fragment amplified from total DNA extract

tag: location

value: agricultural soil, Mississippi, USA


title: DNA_Soil_Lake_Wetland 1


title: DNA_Soil_Riparian_wet 2


title: SHENQI_muscle_3


title: BM-MG 3

city: Stockholm

first: Ewoud

last: Ewing

address: CMM L8:05, Karolinska University Hospital

department: Clinical Neuroscience

hyb_protocol: standard Illumina protocol

scan_protocol: standard Illumina protocol

data_processing: Basically we only read the conserved probes between mouse and human, resulting in 18400 probes being read. This pipeline is an adaptation of the ChAMP pipeline. As for normalization, this mEPIC pipeline, as described in the paper https://www.ncbi.nlm.nih.gov/pubmed/29141580

label: Cy5, Cy3

source_name: Repopulated CNS engrafting macrophage (F4/80hi) in Cx3cr1CreER/+R26DTA/+ mice, 7 weeks after Tamoxifen

label_protocol: standard Illumina protocol

extract_protocol: DNA was extracted using QIAamp DNA micro kit from Qiagen.

treatment_protocol: Naive cells were sorted from completely naive mice or TAM treated Cx3cr1CreER/+ control mice. Cx3cr1CreER/+R26DTA/+ mice were treated with Tamoxifen (TAM) on three consecutive days and cells sorted from the CNS on the indicated weeks.

tag: strain

tag: treatment

value: Tamoxifen

value: Brain

tag: sorting strategy


title: DNA_Soil_Soybean_wet 2


title: Donor 3 NAC treated

value: healty donor 3


title: BM-MG 2

tag: tissue


title: DNA_Soil_Soybean_wet 1


title: BM-MG 1


title: SHENQI_aorta_2


title: BM-MG 1 rep


title: SHENQI_aorta_3


title: SHENQI_muscle_2


title: H44

source_name: Repopulated CNS engrafting macrophage (F4/80hi) in Cx3cr1CreER/+R26DTA/+ mice, 4 weeks after Tamoxifen


title: SHENQI_muscle_1


title: Donor 2 NAC treated

value: healty donor 2


title: yangyinyiqi_aorta_2

description: Formula of SHENQI compound


title: DNA_Soil_Drainage_wet 2


title: H43


title: DNA_Soil_Drainage_wet 1


title: Donor 6 CTRL

source_name: human CD8 T cells_control

value: none (control)


title: Donor 1 NAC treated

value: healty donor 1