title: FP3

city: Zhengzhou

first: chen

last: dong

address: 100 Lianhua Street

institute: Henan University of Technology

data_processing:
We use internal software SOAPnuke by GBI to filter reads
Firstly, mRNA was enriched by oligo(dT) magnetic beads, and then interrupted to short fragments with the size of about 200 bp. The short fragments were used as templates to synthesize the double-strand cDNA by random primers. Additionally, sequencing adaptors were ligated to the fragments, and the required fragments were purified and amplified by PCR reaction. The clean reads were obtained by filtering the low-quality reads, and mapped to the genome of N.nucifera (http://lotus-db.wbgcas.cn/) using HISAT.
All the coding transcripts were merged with reference transcripts to obtain complete reference. Subsequently, the clean reads were mapped to it using Bowtie2. The mRNA expression was determined by RPKM method (Reads per kb per million reads). FDR (false discovery rate)≦0.001 and the absolute valve of log2Ratio≥1 were used as threshold to define the differentially expressed genes (DEGs).
Genome_build: The genome of N.nucifera (http://lotus-db.wbgcas.cn/)
Supplementary_files_format_and_content: RPKM values for each Sample

organism: Nelumbo nucifera

extract_protocol:
Rhizomes of first-year planting  (FP) and consecutive planting  (CP) were harvested, flash frozen on dry ice, and RNA was harvested using Trizol reagent. 1 ug of total RNA was used for the construction of sequencing libraries by BGISEQ-500.
Six rhizomes were randomly collected from both FP and CP groups, with three samples for each group. The total RNA of rhizomes was isolated by Trizol reagent, and the RNA quality was checked by an Agilent 2100 Bioanalyzer (Aglient Technologies, Santa Clara, CA, USA). Firstly, mRNA was enriched by oligo(dT) magnetic beads, and then interrupted to short fragments with the size of about 200 bp. The short fragments were used as templates to synthesize the double-strand cDNA by random primers. Additionally, sequencing adaptors were ligated to the fragments, and the required fragments were purified and amplified by PCR reaction. The library products were scanned for sequencing analysis via BGISEQ-500 (BGI, Wuhan, China).

treatment_protocol: Ten pools were seperated as two groups. One group of N.nucifera was grown in five pools, where N.nucifera had never been planted. For the other group, N.nucifera was consecutively planted in five pools for four years.

tag: cultivar

value: Taikonglian 36

tag: group

value: first-year planting group (FP)

value: rhizome


title: FP1

status: Public on Apr 30 2020

type: SRA

country: China

library_source: transcriptomic

molecule: total RNA

tag: tissue


title: FP2


title: CP1

value: consecutive planting  (CP)


title: DKS14_nL2

city: PACIFICA

first: Hiten

last: Madhani

state: CA

country: USA

institute: UCSF

description:
Affinity purified DNA
processed data file:

library_source: genomic

data_processing:
Illumina adaptor removed from read 3' ends using Cutadapt
Reads were aligned to the  genome with bowtie v 1.1.2
Analysis performed using custom scripts from bedgraph files normalized to wild type (untagged)
Genome_build: C. neoformans CNA3
Supplementary_files_format_and_content: .bedgraph: Signal of ChIP-seq of the sample normalized to the untagged (wild type, K1) sample and smoothed to a 50bp window

molecule: genomic DNA

organism: Cryptococcus neoformans

source_name: yeast

growth_protocol: Samples were inoculated at less than an OD600 of 0.01 from starter cultures in YPAD or YNB at 30°C until reaching OD600 of 1.0, 5.0, or 10.0. An aliquot of the culture as it grew in optical density was taken and harvested by centrifugation at 2000xg for 5 minutes.

extract_protocol:
ChIP DNA was extracted by bead beater with 0.5mm zirconia beads.  ChIP DNA was isolated by antibodies directed to our proteins of interest (10 mg anti-FLAG M2).  DNA was then isolated by incubation with SDS/proteinase K followed by column purification (Macherey-Nagel Nucleospin Gel Cleanup Columns).
library preparation as described in Homer et al. 2016

tag: strain

tag: genotype

value: Liv3-2xFLAG; nrg1_delta

tag: optical density (od600)

tag: media

value: YNB

tag: antibody

value: anti-FLAG

tag: protocol

value: ChIP-seq


title: RET GFP, biological replicate 2

type: RNA

city: Heidelberg

first: Ashik Ahmed

last: Abdul Pari

address: Im neuenheimer Feld 280

country: Germany

institute: German Cabcer Research Institute

hyb_protocol: 15 μg of cRNA was then converted to sensestrand DNA (ss-cDNA). After fragmentation and terminal labeling, 5,5 ug of biotinylated ss-cDNA were hybridized for 17 hr at 45°C on Affymetrix Mouse Gene 2.0 ST Arrays.

scan_protocol: Microarray scanning was done using an Affymetrix GeneChip® Scanner 3000 according to GeneChip® Expression Wash, Stain and Scan Manual for Cartridge Arrays.

data_processing: All intensity data were quarantile normalized.

label: Biotin

organism: Mus musculus

source_name: RET (mouse melanoma cells)

label_protocol: Biotin-labeled ss-cDNA samples for hybridization on GeneChip™ Mouse Gene 2.0 ST Arrays (Affymetrix) were prepared according to Affymetrix's recommended sample labeling procedure as described in the Gene Chip WT Plus Reagent Kit guide. In brief, 100 ng total RNA was used for complementary DNA (cDNA) synthesis and in vitro transcription step to synthesize cRNA according to the Affymetrix® GeneChip® WT Plus Reagent Kit. Quality of cRNA was controlled using the RNA Nano Chip Assay on an Agilent 2100 Bioanalyzer and spectrophotometrically quantified (NanoDrop).

growth_protocol: Control and Angpt2 silenced melanoma cells  were cultured in DMEM high glucose supplemented with 10% FCS, 1% penicillin/streptomycin  and 1% non-essential amino acid

extract_protocol: Total RNA were isolated using  GenElute Mammalian Total RNA Purification Kit (Sigma) according to the manufacturer described protocol.

treatment_protocol: Equal number of control and Angpt2 silenced cells were seeded in a 6 well plate and cultured for 24h. Next, the cells were lysed and RNA was isolated.

tag: genotype/variation

value: non targeting shrna


title: DKS13_qL2_log

value: Liv3-2xFLAG; qsp1_delta


title: DKS14_cN1

value: 3xFLAG-Nrg1; cqs21_delta


title: DKS13_qL1_log


title: DKS13_L2-log


title: DKS13_qN1_OD1

value: 3xFLAG-Nrg1; qsp1_delta


title: DKS13_qN2_OD1


title: DKS13_N2_OD1


title: DKS13_N1_oD1


title: DKS14_C1


title: DKS13_qC2_OD1

value: Cqs2-2xFLAG; qsp1_delta


title: DKS13_C2_OD1


title: CP3


title: Control DKO clone1 - rep3

city: Los Angeles

first: Weiya

middle: Stephanie

last: Ni

state: California

address: 1450 Biggy Street

institute: University of Southern California

data_processing:
RNA-seq results were aligned to Gencode v19 and reads were counted using STAR.
Read count table is generated using STAR as well.
Genome_build: hg19
Supplementary_files_format_and_content: text files include read count for each gene.

organism: Homo sapiens

source_name: Control DKO clone1

growth_protocol: HEK293T cells were grown according to ATCC protocol

extract_protocol:
Total RNA were extracted following the trizol protocol.
Libraries were prepared using Kapa Stranded mRNA kit with beads (Kapa #KK8421) according to provided protocol.

tag: cell line

tag: clone name

value: EGV

tag: grna


title: CP2


title: DKS13_C1_OD1


title: Control DKO clone1 - rep2

description: USC_RNA405_Control_DKO_clone1_EGV_rep2_PE1_read1.fq.gz


title: DKS13_qC1_OD1


title: DKS13_WCE_log

value: mixed

value: all


title: wt_ZFX_FLAG_DKO_clone1_rep2

value: DKO clone1

value: DKO

value: ZFX KO:g1, g2; ZNF711 KO:g1,g2


title: Control DKO clone1 - rep1


title: S2plus: HAP1 SREBF2 +FBS rep2

city: Toronto

first: Jason

last: Moffat

state: ON

address: 686 Bay Street, Room 13.9705

department: Molecular Genetics

country: Canada

institute: University of Toronto

data_processing:
FASTQ files were first filtered using a bespoke Perl script to remove reads shorter than 36bp.
Filtered FASTQ files were aligned to Gencode v25 gene annotations and hg38 genome build using the STAR short-read aligner (v2.6.0a)
Read count files generated by STAR were merged with gene annotations in R
Genome_build: hg38
Supplementary_files_format_and_content: tab-delimited text read counts

source_name: HAP1 cell line

growth_protocol: Human HAP1 cells were maintained in low glucose (10 mM), low glutamine (1 mM) DMEM supplemented with 10% FBS and 1% Penicillin/Streptomycin.

extract_protocol:
RNA was extracted using the RNeasy Kit (QIAGEN) according to manufacturer’s instructions. RNA samples were DNase treated using RNase-free DNase Set (Qiagen, 79254).
1000 ng per sample was processed using the NEBNext Ultra II Directional RNA Library Prep Kit for Illumina (New England Biolabs, E7760L) and included polyA-enrichment using NEBNext Poly(A) mRNA Magnetic Isolation Module (New England Biolabs, E7490L), fragmentation for 15 minutes at 94°C prior to first strand synthesis, and 8 cycles of amplification after adapter ligation.

treatment_protocol: HAP1 WT, C12orf49 or SREBF2 KO cells were cultured in minimal DMEM medium described above for 48h and either control treated or serum-starved for 4 hours as indicated. Each cell line was cultured and processed in three biological replicates.

tag: treatment

value: control


title: S3plus: HAP1 SREBF2 +FBS rep3


title: S2min: HAP1 SREBF2 -FBS rep2

value: serum starvation


title: S3min: HAP1 SREBF2 -FBS rep3


title: C3min: HAP1 C12orf49 -FBS rep3


title: C2min: HAP1 C12orf49 -FBS rep2


title: S1plus: HAP1 SREBF2 +FBS rep1


title: S1min: HAP1 SREBF2 -FBS rep1


title: C1min: HAP1 C12orf49 -FBS rep1


title: DKS13_L1-log


title: C2plus: HAP1 C12orf49 + FBS rep2


title: C1plus: HAP1 C12orf49 + FBS rep1


title: C3plus: HAP1 C12orf49 + FBS rep3


title: W3min: HAP1 WT -FBS rep3

value: WT


title: W2min: HAP1 WT -FBS rep2


title: W1min: HAP1 WT -FBS rep1


title: wt_ZFX_FLAG_DKO_clone1_rep3


title: W3plus: HAP1 WT +FBS rep3


title: wt_ZFX_FLAG_DKO_clone1_rep1


title: W1plus: HAP1 WT +FBS rep1


title: G3.11: Aβ+YW group 1

city: Fukuoka

first: Takashi

last: Ichinose

address: 744 Motooka,Nishi-ku

country: Japan

institute: Kyushu University

hyb_protocol: The cDNAs were subsequently hybridized to a Clariom S Mouse Array (Thermo Fisher Scientific). The arrays were washed and labeled with streptavidin–phycoerythrin using a GeneChip Hybridization, Wash and Stain Kit and Fluidics Station 450 system (Thermo Fisher Scientific).

scan_protocol: Fluorescence was detected using a GeneChip Scanner 3000 7G (Thermo Fisher Scientific).

data_processing: Affymetrix GeneChip Command Console software was used to reduce the array images to the intensity of each probe (CEL files). The CEL files were quantified using the Factor Analysis for Robust Microarray Summarization algorithm (quantile normalization, qFARMS) with the statistical packages R and Bioconductor.

label: biotin

source_name: Murine cerebral corex (Aβ peptides 25–35 injection)

label_protocol: Biotinylated single-stranded complementary DNA (cDNA) was synthesized from 100 ng of total RNA using the GeneChip WT PLUS reagent kit (Thermo Fisher Scientific).

growth_protocol: Male ddY mice at 10 weeks of age (SLC, Inc., Shizuoka, Japan) were housed in a room with controlled lighting (12 h light/12 h dark), and constant temperature (25°C), and provided with moderate fat diet (Oriental Yeast Co., Ltd., Tokyo, Japan) and water ad libitum. Mice were divided into 3 groups based on similar average body weight.

extract_protocol: Total RNA was isolated from each liver sample using TRIzol reagent (Thermo Fisher Scientific, Waltham, MA, USA) and was subsequently purified using an RNeasy Mini Kit (Qiagen, Hilden, Germany) and RNase-Free DNase Set (Qiagen), according to the manufacturer’s protocol.

treatment_protocol: Mice were divided into 3 groups (n=10 per group) as follows: the sham group, which received saline administration and distilled water injection; the amyloid beta (Aβ) group, which received saline administration and Aβ peptides 25–35 injection; and the Aβ+YW group, which received Tyr-Trp administration and Aβ peptides 25–35 injection. Tyr-Trp was orally administered Tyr-Trp (100 mg kg-1 day-1, twice a day), starting 7 days before Aβ peptides injection. The other groups received oral administration of saline (5 mL kg-1). After 7 days of Aβ peptides injection or sham operation, behavioral test was conducted and Cx samples were collected.

value: ddY

value: male

value: cerebral cortex


title: G2.10: Aβ group 5


title: 2-THP1-T200

status: Public on Apr 29 2020

city: Melbourne

first: Zheng

last: Fan

state: Victoria

address: 305 Grattan Street

country: Australia

institute: Peter MacCallum Cancer Centre

data_processing:
The first 12 bases of each read were clipped using Cutadapt (v1.18) and
Clipped reads were processed by the Tail Tools pipeline (v1.5) using the ENSEMBL Homo sapiens reference (release 93) to generate differential gene expression matrices and perform alternative polyadenylation analysis
For differential gene expression analysis, reads were counted at the level of whole genes, considering reads anywhere within the annotated gene region and up to 2kb down-stream of the annotated TES. Reads that covered a down-stream annotated gene on the same strand were excluded and genes for which no relevant sample had at least 10 reads were removed.
Differential gene expression was tested between experimental groups using the Fitnoise library (https://github.com/pfh/fitnoise) as part of the Tail Tools pipeline, following log transformation and weighting of normalized counts using voom from the limma R package (v3.40.6).
For alternative polyadenylation analysis, peaks were called if at least 10  reads ending in a poly(A) sequence of at least 6 bases were present. Peaks where genomic sequences consisting of 60% or more "A"s could account for the poly(A) sequence were regarded as mis-priming events and excluded.
To assign peaks to genes, peaks called anywhere within an annotated gene region and up to 2kb down-stream of the annotated TES were included. Peaks that covered a down-stream annotated gene on the same strand were excluded.  For genes where more than two peaks were detected, the two peaks with the highest number of reads across all samples were selected.
To compare between two experimental groups (e.g. DMSO-treated), a test was performed for each gene with two peaks to detect an interaction between peak usage and experimental group. This test was performed by the Fitnoise library, following log transformation and weighting of peak counts using voom from the limma R package (v3.40.6).
Bigwig files were also generated by Tail Tools pipeline.
Genome_build: ENSEMBL Homo sapiens reference (release 93) (hg38)
Supplementary_files_format_and_content: Comma-separated values of genewise; peakwise; peakpairwise count file before normalisation. Bigwig file with forward  (positive) and reverse (negitave) strands RNA reads densities. GFF file for all the peaks called by Tailtools.

molecule: polyA RNA

source_name: THP-1 cells

growth_protocol: Cells were cultured in RPMI- 1640 supplemented with 10% v/v heat-inactivated fetal bovine serum, 100U/mL penicillin and 100μg/mL streptomycin 2mM GlutaMAX  and 1mM sodium pyruvate at 37 degree Celsius and 5% carbon dioxide.

extract_protocol:
Cells were lysed in TRIzol, then RNA was extrated using Direct-Zol RNA MiniPrep kit (Zymo Research)
3’ RNA-sequencing, sequencing libraries were prepared from 500ng RNA using the Lexogen QuantSeq 3’ mRNA Seq Library Prep Kit FWD for Illumina. 75 single-end base-pair reads were sequenced using the
Illumina NEXTseq 500.
QuantSeq 3’ mRNA Seq Library Prep Kit FWD for Illumina

treatment_protocol: Cells were treated with 200nM THZ531/DMSO for 6 hours

tag: treatmet

tag: replicate


title: 1-THP1-D

value: DMSO


title: 2-H1106-293-1

data_processing:
The first 12 bases of each read were clipped using Cutadapt (v1.18) and
Clipped reads were processed by the Tail Tools pipeline (v1.5) using the ENSEMBL Homo sapiens reference (release 93) to generate differential gene expression matrices and perform alternative polyadenylation analysis
For differential gene expression analysis, reads were counted at the level of whole genes, considering reads anywhere within the annotated gene region and up to 2kb down-stream of the annotated TES. Reads that covered a down-stream annotated gene on the same strand were excluded and genes for which no relevant sample had at least 10 reads were removed.
Differential gene expression was tested between experimental groups using the Fitnoise library (https://github.com/pfh/fitnoise) as part of the Tail Tools pipeline, following log transformation and weighting of normalized counts using voom from the limma R package (v3.40.6).
For alternative polyadenylation analysis, peaks were called if at least 10  reads ending in a poly(A) sequence of at least 6 bases were present. Peaks where genomic sequences consisting of 60% or more "A"s could account for the poly(A) sequence were regarded as mis-priming events and excluded.
To assign peaks to genes, peaks called anywhere within an annotated gene region and up to 2kb down-stream of the annotated TES were included. Peaks that covered a down-stream annotated gene on the same strand were excluded.  For genes where more than two peaks were detected, the two peaks with the highest number of reads across all samples were selected.
To compare between two experimental groups (e.g. DMSO-treated to THZ531 treated and WT Amr to Pol II mutants), a test was performed for each gene with two peaks to detect an interaction between peak usage and experimental group. This test was performed by the Fitnoise library, following log transformation and weighting of peak counts using voom from the limma R package (v3.40.6).
Bigwig files were also generated by Tail Tools pipeline.
Genome_build: ENSEMBL Homo sapiens reference (release 93) (hg38)
Supplementary_files_format_and_content: Comma-separated values of genewise; peakwise; peakpairwise count file before normalisation. Bigwig file with forward  (positive) and reverse (negitave) strands RNA reads densities. GFF file for all the peaks called by Tailtools.

growth_protocol: Cells were cultured in DMEM supplemented with 10% v/v HI-FBS, 100U/mL penicillin and 100μg/mL streptomycin, 200μg/mL hygromycin, 6.5μg/mL blasticidin at 37 degree Celsius and 10% carbon dioxide.

treatment_protocol: Experiments performed with Flp-In-HEK293 pcDNA5/FRT/TO-Rpb1 cells involved 16-hour incubation with 2μg/mL doxycycline to induce mutant Rpb1 expression followed by 42-hour incubation with 2.5μg/mL a-amanitin to inactivate endogenous RNA polymerase II.  For HEK293T cells, celles were treated with 400nM THZ531/DMSO for 6 hours.

tag: flp in pol ii mutations

value: Rbp1 H1108Y Amr

value: a-amanitin


title: 1-293-D-1


title: 2-THP1-D


title: 2-293-D-2


title: 3-293-T-2


title: G1.3: Sham group 3

source_name: Murine cerebral corex (sham operation)


title: 5-WT-293-2

value: Rpb1 WT Amr


title: G1.1: Sham group 1


title: G3.14: Aβ+YW group 4

source_name: Murine cerebral corex (Aβ peptides 25–35 injection+Tyr-Trp administration)


title: G2.7: Aβ group 2


title: G3.15: Aβ+YW group 5


title: G2.6: Aβ group 1


title: G2.8: Aβ group 3


title: G2.9: Aβ group 4


title: G1.4: Sham group 4


title: G3.13: Aβ+YW group 3


title: G3.12: Aβ+YW group 2


title: G1.2: Sham group 2


title: 1-THP1-T200


title: 8-R749-293-2

value: Rbp1 R749H Amr


title: 3-293-T1


title: W2plus: HAP1 WT +FBS rep2


title: 6-H1106-293-2


title: 4-R749-293-1


title: 1-WT-293-1


title: 11-Input-WTminusT25-PRO

data_processing:
The sequenced reads were demultiplexed using bcl2fastq (v2.17.1.14) and basic QC was performed on the resulting FASTQ files using FASTQC (v0.11.5).
FASTQ reads were mapped to the GRCh37/hg19 genome using Bowtie2 (v2.3.3) using the standard settings.
The resulting SAM files were converted to BAM files using Samtools (v 1.4.1) using the view command, after which the BAM files were sorted, indexed and potential PCR duplicates removed using the rmdup function.
The BAM files were converted into BigWig files (bw) using the BamCoverage function in Deeptools (v3.0.0)  (--normalizeUsing CPM --smoothLength 150 --binSize 50 -e 200 scaleFactor 1).
Genome_build: hg19
Supplementary_files_format_and_content: bigWig files that contains PRO-seq reads coverage across the genome in CPM

source_name: MV4;11 cells

extract_protocol:
Cells were washed in ice cold PBS prior to incubation in Permeabilization Buffer (1mL/10^6 cells; 100μL/10^7 cells; 10mM Tris-HCl pH 7.4, 200mM sucrose, 10mM KCl, 5mM MgCl2, 1mM EDTA, 0.05% v/v Tween20, 0.15% v/v NP-40, 0.5mM DTT, 2U/mL SUPERase•In (ThermoFisher Scientific, AM2694) containing Roche cOmpleteTM protease inhibitors for 5 minutes on ice. The cell pellet was washed twice with Permeabilization Buffer (0.5mL/10^6 cells per wash) and then resuspended in Storage Buffer (10mM Tris-HCl pH 8, 25% glycerol, 5mM MgCl2, 0.1mM EDTA, 5mM DTT). For the nuclear run-on reaction, permeabilized cells were incubated with one volume of 2X Nuclear Run-On reaction mastermix containing Biotin 11-CTP (1.4mM Tris-HCl pH 8, 0.7mM MgCl2, 0.14mM DTT, 42mM KCl, 0.05mM Biotin 11-CTP (Perkin Elmer NEL544001EA), 2.5µM CTP, 0.25mM ATP, 0.25mM GTP, 0.25mM UTP, 40U SUPERase•In, 1% Sarkozyl (Sigma Alrich L5125)) at 37ºC for three minutes with gentle agitation prior to the addition of TRIzol-LS (ThermoFisher 10296010). TRIzol samples were mixed with chloroform and the aqueous phase was isolated after centrifugation. 1μL GlycoBlue and 2.5 volumes ethanol was added to the aqueous phase for RNA precipitation (incubation at room temperature for 10 minutes, 14000g centrifugation at 4ºC for 20 minutes). Precipitated RNA was washed with 70 percent ethanol, resuspended in nuclease free water. RNA was denatured at 65ºC for 40 seconds and then was incubated on ice with 0.2M NaOH for 12.5 minutes prior to the addition of 1 volume 1M Tris-HCl (pH 6.8). Base hydrolyzed RNA was exchanged to 10mM Tris solution using a Micro Bio-Spin 30 Column (BIORAD, 7326250) as per the manufacturer’s instruction. For enrichment of Biotin-incorporated RNA, hydrolyzed RNA was incubated with Dynabeads M-280 Streptavidin (ThermoFisher Scientific, 11206D) in PRO-Seq Binding Buffer (10mM Tris-HCl pH 7.4, 300mM NaCl, 0.1% v/v TritonX-100) at room temperature for 20 minutes with gentle rotation. Streptavidin beads were washed (1 minute with rotation per wash) three times each in High Salt Wash Buffer (50mM Tris-HCl pH 7.4, 2M NaCl, 0.5% v/v TritonX-100), Binding Buffer and Low Salt Wash Buffer (5mM Tris-HCl pH 7.4, 0.1% TritonX-100) and then resuspended in TRIzol. Following incubation at room temperature for 3 minutes, TRIzol samples were mixed with chloroform and the aqueous phase was isolated after centrifugation. TRIzol/chloroform treatment of washed Streptavidin beads was repeated once and beads were incubated with chloroform for 3 minutes at room temperature prior to centrifugation. 1μL GlycoBlue and 900μL ethanol was added to the pooled aqueous phase solution and samples were incubated at room temperature for 10 minutes prior to centrifugation. Precipitated biotin-enriched RNA was washed with 70% ethanol and resuspended in nuclease free water.
PRO-Seq Sequencing libraries were prepared using NEBNext Ultra II Directional RNA Library Prep Kit with Ribo-Depletion
NEBNext® Ultra II Directional RNA Library Prep Kit for Illumina

treatment_protocol: MV4;11 cells treated with DRB for 4 hours, and released for 25 minutes in the presence of 1-NM-PP-1/DMSO.

tag: analog sensetive muntants


title: 10-D2-plus-T25


title: 9-D2-minus-T25


title: 7-13-plus-T25


title: 8-D1-plus-T25


title: 6-12-plus-T25


title: 4-D1-minus-T25


title: 5-WT-plus-T25


title: 3-13-minus-T25


title: 2-12-minus-T25


title: 3-100 (total RNA)

data_processing:
Reads were trimmed to 100bp with Cutadapt (v1.18) to remove adapter sequences.
Reads were aligned to GRCh37/hg19 with HISAT2 (v2.1.0) using settings for reverse-stranded library. Reads with MAPQ < 10 and secondary alignments were filtered.
MISO (v0.5.4) alternative events analysis was performed using the annotation supplied by MISO.
As MISO does not handle replicates, each replicate experiment was analysed independently and events were required to pass the following thresholds in all 3 replicate experiments: --num-inc 1 --num-exc 1 --num-sum-inc-exc 10 --delta-psi 0.20 --bayes-factor 10 --votes 3. As tandem 3′UTR events do not have exclusion reads (the shorter isoform is contained within the longer isoform) the following filter was used for those events: --num-inc 2 --num-sum-inc-exc 10 --delta-psi 0.20 --bayes-factor 10 --votes 3.
For visulisation of the RNA-Seq reads, the reads were trimmed by Cutadapt (Galaxy verdsion1.16.1)
Reads were aligned to hg19 with HISAT2 (Galaxy Version 2.1.0 to GRCh37/hg19 using settings for reverse-stranded library.
bamCoverage (Galaxy Version 3.0.2.0) were used to genarate bigwig file for visulisation usage.
Genome_build: hg19
Supplementary_files_format_and_content: Bigwig files that contains the coverage of RNA-Seq reads in CPM, and Tab-separated values files that contains MISO splicing analysis output for each alternative splicing event types.

extract_protocol:
Cells were lysed in TRIzol, then RNA was extrated using Direct-Zol RNA MiniPrep kit (Zymo Research)
NEBNext® rRNA Depletion Kit (New England BioLabs, Ipswich MA USA, E6310) sequencing libraries were prepared from 500ng RNA using the NEBNext® Ultra II Directional RNA Library Prep Kit for Illumina. 150 paired-end base-pair reads were sequenced using the Illumina NEXTseq 500 (Illumina, San Diego CA USA).
NEBNext® Ultra II Directional RNA Library Prep Kit for Illumina

treatment_protocol: Cells were treated with DMSO or 100/200nM THZ531 for 6 hours


title: 1-WT-minus-T25


title: 2-D (total RNA)


title: 2-100 (total RNA)


title: 3-200 (total RNA)


title: 1-200 (total RNA)


title: 3-D (total RNA)


title: 1-100 (total RNA)


title: ZF-Input-Pooled

data_processing:
The sequenced reads were demultiplexed using bcl2fastq (v2.17.1.14) and basic QC was performed on the resulting FASTQ files using FASTQC (v0.11.5).
FASTQ reads were mapped to the GRCh37/hg19 genome using Bowtie2 (v2.3.3) using the standard settings.
The resulting SAM files were converted to BAM files using Samtools (v 1.4.1) using the view command, after which the BAM files were sorted, indexed and potential PCR duplicates removed using the rmdup function.
Deeptools (v3.0.0) was used to generate occupancy heatmaps and the resulting CPM normalized occupancy matrix was used as input for custom R scripts to generate average profile plots and to calculate processitivy indeces.
Briefly, the BAM files were converted into BigWig files (bw) using the BamCoverage function (--normalizeUsing CPM --smoothLength 150 --binSize 50 -e 200 scaleFactor 1).
Genome_build: GRCh37/hg19
Supplementary_files_format_and_content: bigWig files that contains Pol II ChIP-Seq reads coverage on the genome in CPM.

extract_protocol:
Cells were crossed-linked with formaldehylde solution. Then nuclear extracts were isolated, and sonicated. Chromatin bound to Pol II were isolated using RNA polymerase II, clone CTD4H8, mouse monoclonal antibody.
ChIP-Seq libraries were prepared using the NEBNext Ultra II DNA Library Prep Kit for Illumina (New England Biolabs, E7645). Fragments between 200 and 500 base pairs were selected using a Pippin Prep 1.5% agarose cassette (Sage Science, Beverly MA US), pooled and 75 single-end base pair reads were sequenced using the Illumina NEXTseq 500.
NEBNext Ultra II DNA Library Prep Kit for Illumina (New England Biolabs, E7645)

value: DMSO/100nM/200nM THZ531


title: 13-T45-D

extract_protocol:
Cells were washed in ice cold PBS prior to incubation in Permeabilization Buffer (1mL/10^6 cells; 100μL/10^7 cells; 10mM Tris-HCl pH 7.4, 200mM sucrose, 10mM KCl, 5mM MgCl2, 1mM EDTA, 0.05% v/v Tween20, 0.15% v/v NP-40, 0.5mM DTT, 2U/mL SUPERase•In (ThermoFisher Scientific, AM2694) containing Roche cOmpleteTM protease inhibitors for 5 minutes on ice. The cell pellet was washed twice with Permeabilization Buffer (0.5mL/10^6 cells per wash) and then resuspended in Storage Buffer (10mM Tris-HCl pH 8, 25% glycerol, 5mM MgCl2, 0.1mM EDTA, 5mM DTT). For the nuclear run-on reaction, permeabilized cells were incubated with one volume of 2X Nuclear Run-On reaction mastermix containing Biotin 11-CTP (1.4mM Tris-HCl pH 8, 0.7mM MgCl2, 0.14mM DTT, 42mM KCl, 0.05mM Biotin 11-CTP (Perkin Elmer NEL544001EA), 2.5µM CTP, 0.25mM ATP, 0.25mM GTP, 0.25mM UTP, 40U SUPERase•In, 1% Sarkozyl (Sigma Alrich L5125)) at 37ºC for three minutes with gentle agitation prior to the addition of TRIzol-LS (ThermoFisher 10296010). TRIzol samples were mixed with chloroform and the aqueous phase was isolated after centrifugation. 1μL GlycoBlue and 2.5 volumes ethanol was added to the aqueous phase for RNA precipitation (incubation at room temperature for 10 minutes, 14000g centrifugation at 4ºC for 20 minutes). Precipitated RNA was washed with 70 percent ethanol, resuspended in nuclease free water. RNA was denatured at 65ºC for 40 seconds and then was incubated on ice with 0.2M NaOH for 12.5 minutes prior to the addition of 1 volume 1M Tris-HCl (pH 6.8). Base hydrolyzed RNA was exchanged to 10mM Tris solution using a Micro Bio-Spin 30 Column (BIORAD, 7326250) as per the manufacturer’s instruction. For enrichment of Biotin-incorporated RNA, hydrolyzed RNA was incubated with Dynabeads M-280 Streptavidin (ThermoFisher Scientific, 11206D) in PRO-Seq Binding Buffer (10mM Tris-HCl pH 7.4, 300mM NaCl, 0.1% v/v TritonX-100) at room temperature for 20 minutes with gentle rotation. Streptavidin beads were washed (1 minute with rotation per wash) three times each in High Salt Wash Buffer (50mM Tris-HCl pH 7.4, 2M NaCl, 0.5% v/v TritonX-100), Binding Buffer and Low Salt Wash Buffer (5mM Tris-HCl pH 7.4, 0.1% TritonX-100) and then resuspended in TRIzol. Following incubation at room temperature for 3 minutes, TRIzol samples were mixed with chloroform and the aqueous phase was isolated after centrifugation. TRIzol/chloroform treatment of washed Streptavidin beads was repeated once and beads were incubated with chloroform for 3 minutes at room temperature prior to centrifugation. 1μL GlycoBlue and 900μL ethanol was added to the pooled aqueous phase solution and samples were incubated at room temperature for 10 minutes prior to centrifugation. Precipitated biotin-enriched RNA was washed with 70% ethanol and resuspended in nuclease free water.
PRO-Seq Sequencing libraries were prepared using NEBNext Ultra II Directional RNA Library Prep Kit.
NEBNext® Ultra II Directional RNA Library Prep Kit for Illumina

treatment_protocol: Cells were treated with a co-treatment that consistent of  DMSO/100nM /200nM THZ531 and DRB for 4 hours. Control samples were treated with DMSO/100nM/200nM THZ531 without DRB for 4 hours

value: DRB+DMSO

tag: release time

value: 45 minute


title: 14-T45-T1

value: DRB+100nM THZ531


title: ZF-200-P

value: RNA polymerase II, clone CTD4H8, mouse monoclonal


title: 2-200 (total RNA)


title: ZF-100-P


title: Input

value: Pooled


title: 15-T45-T2

value: DRB+200nM THZ531


title: ZF-DMSO-P


title: 12-T25-T2

value: 25 miunte


title: 9-T125-T2

value: 12.5 minute


title: 10-T25-D


title: 2-T1


title: 3-T2


title: 1-D (DRB-PRO-Seq)


title: 4-T0-D

value: 0 minute


title: 7-T125-D


title: 8-T125-T1


title: 6-T0-T2


title: 1-D

data_processing:
5bp from 5' side and 20bp from 3' side of the reads were trimmed by Cutadapt (1.14)
Trimmed reads were mapped to hg19 using HISAT2 (2.1.0)
The resulting SAM files were converted to BAM files using Samtools (1.4.1) using the view command, after which the BAM files were sorted, indexed
Bam files were converted to tdf files using IGVtools (2.3.95)
Genome_build: hg19
Supplementary_files_format_and_content: tdf files that contains the coverage of RNA-Seq on the genome.


title: 1-THZ100


title: 1-THZ200


title: 22-Parental-DMSO2

data_processing:
The first 12 bases of each read were clipped using Cutadapt (v1.18)
Clipped reads were processed by the Tail Tools pipeline (v1.5) using the ENSEMBL Homo sapiens reference (release 93) to generate differential gene expression matrices and perform alternative polyadenylation analysis
For differential gene expression analysis, reads were counted at the level of whole genes, considering reads anywhere within the annotated gene region and up to 2kb down-stream of the annotated TES. Reads that covered a down-stream annotated gene on the same strand were excluded and genes for which no relevant sample had at least 10 reads were removed.
Differential gene expression was tested between experimental groups using the Fitnoise library (https://github.com/pfh/fitnoise) as part of the Tail Tools pipeline, following log transformation and weighting of normalized counts using voom from the limma R package (v3.40.6).
For alternative polyadenylation analysis, peaks were called if at least 10  reads ending in a poly(A) sequence of at least 6 bases were present. Peaks where genomic sequences consisting of 60% or more "A"s could account for the poly(A) sequence were regarded as mis-priming events and excluded.
To assign peaks to genes, peaks called anywhere within an annotated gene region and up to 2kb down-stream of the annotated TES were included. Peaks that covered a down-stream annotated gene on the same strand were excluded.  For genes where more than two peaks were detected, the two peaks with the highest number of reads across all samples were selected.
To compare between two experimental groups (e.g. DMSO-treated and 1-NM-PP1-treated), a test was performed for each gene with two peaks to detect an interaction between peak usage and experimental group. This test was performed by the Fitnoise library, following log transformation and weighting of peak counts using voom from the limma R package (v3.40.6).
Bigwig files were also generated by Tail Tools pipeline.
Genome_build: ENSEMBL Homo sapiens reference (release 93) (hg38)
Supplementary_files_format_and_content: Comma-separated values of genewise; peakwise; peakpairwise count file before normalisation. Bigwig file with forward  (positive) and reverse (negitave) strands RNA reads densities. GFF file for all the peaks called by Tailtools.

extract_protocol:
RNA was extracted using Trizol/Chloroform extraction, DTT was added to protect 4sU to be converted. RNA pellet was treated with IAA to convert the 4sU, then cleaned up using ethanol.
3’ RNA-sequencing, sequencing libraries were prepared from 500ng RNA using the Lexogen QuantSeq 3’ mRNA Seq Library Prep Kit FWD for Illumina. 75 single-end base-pair reads were sequenced using the
Illumina NEXTseq 500.
QuantSeq 3’ mRNA Seq Library Prep Kit FWD for Illumina

treatment_protocol: Cells were treated with 10 μM of  1-NM-PP1/DMSO for 4 hours, or 200nM THZ531/DMSO for 6 hours at tast 30 minutes treatment, 500μM 4-thiouridine was added.

value: Parental MV4;11 cells


title: 5-T0-T1


title: 21-Parental-DMSO1


title: 6-S2-200-DM

data_processing:
The sequenced reads were demultiplexed using bcl2fastq (v2.17.1.14) and basic QC was performed on the resulting FASTQ files using FASTQC (v0.11.5).
FASTQ reads were mapped to combined hg19/dm3 genome using Bowtie2 (v2.3.3) using the standard settings.
The resulting SAM files were converted to BAM files using Samtools (v 1.4.1) using the view command, after which the BAM files were sorted, indexed and potential PCR duplicates removed using the rmdup function.
Reads were mapped to a combined hg19/dm3 genome after which the reads mapping to fly and human genomes were calculated using FeatureCounts from the Subread package (v1.5.0).
The scaleFactor subquently determined by calculating the reads mapping to the fly genome relative to total mapped reads (combined hg19/dm3 reference) and applied in the BamCoverage command (--normalizeUsing CPM --smoothLength 150 --binSize 50 -e 200 scaleFactor (as calculated)).
Genome_build: hg19 & dm3
Supplementary_files_format_and_content: Bigwig files with normalised Pol II Ser2 ChIP-Seq reads coverage acrossed the genome in CPM.

organism:
Drosophila melanogaster
Homo sapiens

source_name: MV4;11 cells & Schneider 2 cells

extract_protocol:
Cells were crossed-linked with formaldehylde solution. Then nuclear extracts were isolated, and sonicated. Chromatin bound to phospho Ser2 Pol II were isolated using RNA polymerase II (phospho CTD Ser-2), clone 3E10, rat monoclonal.
ChIP-Seq libraries were prepared using the NEBNext Ultra II DNA Library Prep Kit for Illumina (New England Biolabs, E7645). Fragments between 200 and 500 base pairs were selected using a Pippin Prep 1.5% agarose cassette (Sage Science, Beverly MA US), pooled and 75 single-end base pair reads were sequenced using the Illumina NEXTseq 500.
NEBNext Ultra II DNA Library Prep Kit for Illumina (New England Biolabs, E7645)


title: 11-T25-T1


title: LN308-untreated-R1 (RNA-Seq)

first: Grischa

last: Toedt

address: Meyerhofstr. 1

department: Computational Biology

institute: EMBL

data_processing:
Illumina Casava1.8+ software used for basecalling.
Sequenced reads were trimmed for adaptor sequence, then mapped to GRCh38 using STAR v2.5.4b with gencode.v26.annotation.  Parameters for Ribo-Seq: --outFilterType BySJout --outFilterMultimapNmax 20 --outFilterMultimapScoreRange 1 --outFilterScoreMinOverLread 0.66 --outFilterMatchNmin 18 --outFilterMatchNminOverLread 0.66 --outFilterMismatchNmax 999 --outFilterMismatchNoverLmax 0.04 --alignIntronMin 20 --alignIntronMax 1000000 --alignMatesGapMax 1000000 --alignSJoverhangMin 8 --alignSJDBoverhangMin 1 --alignEndsType EndToEnd --alignSoftClipAtReferenceEnds No --chimSegmentMin 20 --chimSegmentReadGapMax 0 10 --best --strata  Parameters for RNA-Seq: --outFilterMultimapNmax 20 --outFilterMultimapScoreRange 1 --outFilterMismatchNmax 999 --outFilterMismatchNoverLmax 0.04 --outFilterMatchNmin 16 --outFilterMatchNminOverLread 0.66 --outFilterScoreMinOverLread 0.66 --outFilterType BySJout --alignIntronMin 20 --alignIntronMax 1000000 --alignMatesGapMax 1000000 --alignSJoverhangMin 8 --alignSJDBoverhangMin 1 --alignEndsType Local --alignSoftClipAtReferenceEnds No --chimSegmentMin 20 --chimSegmentReadGapMax 0 --outMultimapperOrder Random --outSAMattributes Standard --outSAMmode Full --outSAMstrandField intronMotif --outReadsUnmapped Fastx --outSAMtype BAM SortedByCoordinate --outSAMmultNmax -1
Gene level expression were quantified using HTSeq (Anders et al. 2015) and raw RNA-Sequencing count data normalized and fold changes computed using the DESeq2 library (Love et al. 2014). Translation efficiencies of ribosomal profiling data was computed using RiboDiff (Zhong et al. 2017).
Genome_build: GRCh38
Supplementary_files_format_and_content: tab delimited data files include ensembl gene ids and counts per sample.

source_name: LN308 cell line

growth_protocol: LN308 cells were cultured in DMEM supplemented with 10% FBS, 100 units/ml penicillin and 100µg/ml streptomycin at 37°C and at 5% CO2.

extract_protocol:
Ribo-Seq  TH- or TM-stimulated cells were treated with 0.1mg/ml cycloheximide for 5min.  After washing with ice-cold PBS (supplemented with cycloheximide), cells were harvested and lysed for 15min on ice in a buffer containing 5mM Tris pH7.4, 1.5mM KCl, 5mM MgCl2, 0.5% Triton-X, 0.5% sodium deoxycholate, 0.1mg/ml cycloheximide, RNasin and 1x cOmplete protease inhibitor cocktail (Roche).  The cell lysate was cleared by centrifugation and digested with 0.2U RNaseI (Ambion) per 1µg RNA for 15min.  To stop the reaction, 1U SUPERaseIn RNase Inhibitor (Invitrogen) was added per unit of RNaseI.  Ribosomal complexes were separated on 12ml 10-50% sucrose gradients (in a buffer containing 20mM Tris pH7.4, 75 mM NaCl, 5mM MgCl2, 0.1mg/ml cycloheximide, and 1mM DTT) by centrifugation at 4°C and 35,000rpm for 3h in a SW41 rotor.  Fractions were collected at 0.5ml/min with continuous monitoring of conductivity and UV absorption at 254nm.  Fractions containing 80S monosomes were diluted with an equal amount of RNase-free water followed by organic extraction with Phenol:Chloroform:Isoamylalkohol 25:24:1 (Roth) and ethanol precipitation.  RNAs were then separated by denaturing Urea-PAGE and visualized by SYBR Gold staining.  RNAs with a length of 27-33nt were excised from the gel and eluted overnight in a buffer containing 300mM NaOAc pH 5.5, 1mM EDTA, and 0.25% SDS followed by ethanol precipitation.
RNA-Seq  Total cellular RNA was extracted using TRIzol reagent according to the manufacturer’s instructions.
Ribo-Seq  After dephosphorylation for 1h at 37°C with 20U of T4 Polynucleotide Kinase (NEB), RNAs were ligated to a universal miRNA cloning linker (NEB) using truncated T4 RNA ligase 2.  After gelpurification reverse transcription was performed using SuperscriptIII reverse transcriptase and a primer complementary to the ligated adapter, followed by alkaline hydrolysis of the RNA strand.  The resulting cDNAs were gelpurified and contaminating sequences originating from rRNA were depleted by substractive hybridization to a mixture of biotinylated DNA oligonucleotides and Streptavidin magnetic beads (New England Biolabs).  The recovered RNAs were then circularized with CircLigase II ssDNA Ligase (Epicentre) according to the manufacturer’s instructions and subjected to PCR amplification to generate amplicons suitable for Illumina sequencing.  After gelpurification, DNA concentration was determined using a Qubit Fluorometer (Qubit 2.0, Thermo Fisher Scientific).  Bioanalyzer or TapeStation (Agilent) analyses were performed to assess the quality of the samples followed by deep sequencing on the Illumina platform.
RNA-Seq  RNA integrity was checked using the RNA Nano 6000 Assay Kit of the Bioanalyzer 2100 system (Agilent Technologies, Santa Clara, CA), and concentration was measured with Qubit® RNA Assay Kit in Qubit® 2.0 Flurometer (Life Technologies).  Starting with ~300ng of total RNA as input, ribosomal RNA was removed by NEBNext rRNA Depletion Kit (Human/Mouse/Rat) (New England Biolabs).  Subsequently, stranded total RNA-seq libraries were prepared from rRNA free RNA sample using the NEBNext® Ultra™ II Directional RNA Library Prep Kit for Illumina® (New England Biolabs) according to the protocol.  RNA-seq barcoded libraries that passed the QC step, which was assessed on the Agilent Bioanalyzer system, were then pooled in equimolar amounts;  1.8 pM solution of this pool was loaded on the Illumina sequencer NextSeq 500 and sequenced bi-directionally.

treatment_protocol: To induce ER stress the medium was supplemented with 200nM Thapsigargin or 2.5µg/ml Tunicamycin (dissolved in DMSO); DMSO only served as a control.

tag: cell type

value: Astrocytoma-derived p53-deficient glial cells

tag: time

value: 0 h

value: untreated


title: 25-Parental-THZ200-1


title: 4-S2-D-DM

value: DMSO

value: RNA polymerase II (phospho CTD Ser-2), clone 3E10, rat monoclonal


title: 5-S2-100-DM


title: 19-Dual2-N1

value: #2CDK12AS/NULL;CDK13AS/AS Clone


title: 3-Input-S2-200-DM


title: 2-Input-S2-100-DM


title: 1-Input-S2-D-DM


title: 18-Dual1-N2

value: #1CDK12AS/NULL;CDK13AS/NULL Clone


title: 17-Dual1-N1


title: 15-13-N1

value: CDK13AS/AS Clone


title: 16-13-N2


title: R2-NEW-WT-NMP

description: 4sU labeled RNA

data_processing:
The sequenced reads were demultiplexed using bcl2fastq (v2.17.1.14) and basic QC was performed on the resulting FASTQ files using FASTQC (v0.11.5).
FASTQ reads were mapped to combined hg19/dm3 genome using Bowtie2 (v2.3.3) using the standard settings.
The resulting SAM files were converted to BAM files using Samtools (v 1.4.1) using the view command, after which the BAM files were sorted, indexed and potential PCR duplicates removed using the rmdup function.
Reads were mapped to a combined hg19/dm3 genome after which the reads mapping to fly and human genomes were calculated using FeatureCounts from the Subread package (v1.5.0).
The scaleFactor subquently determined by calculating the reads mapping to the fly genome relative to total mapped reads (combined hg19/dm3 reference) and applied in the BamCoverage command (--normalizeUsing CPM --smoothLength 150 --binSize 50 -e 200 scaleFactor (as calculated)).
Genome_build: hg19 & dm3
Supplementary_files_format_and_content: bigWig files that contains normalised TT-seq reads coverage across the genome in CPM

extract_protocol:
Cells were lysed in TRIZOL. Then, chloroform (1/5th volume) was added and samples were vortexed and centrifuged at 4 degree Celsius (13000g, 15 minutes). The aqueous phase was isolated and total RNA was extracted by isopropanol precipitation at room temperature (10 minutes) followed by centrifugation at 4 degree Celsius (13000g, 10 minutes). The isolated RNA pellet was washed with 70 percent ethanol followed by centrifugation at 4 degree Celsius (13000g, 15 minutes). The RNA pellet was dissolved in 100 L nuclease-free H2O and denatured at 65ºC for 10 minutes. 150μg total RNA per treatment condition (plus 15μg D.melanogaster S2 RNA spike-in) in 100μL final volume were sonicated in microTUBE AFA Fiber screw cap tubes (6 x 16mm, Covaris, 520096) using the Covaris S220 Focused Ultrasonicator at maximum power for 15 seconds. Thiol-specific biotinylation of fragmented RNA was performed by adding 750μL volume nuclease free water followed by incubatation with 10mM TRIS (pH 7.4), 1mM EDTA, 200 μg/mL dimethylformamide (DMF; Sigma Aldrich, 227056) and 300μg EZ-LinkTM HPDP-Biotin (ThermoFisher Scientific, 21341) at room temperature for 1.5 hours with constant rotation (1500μL final volume – ‘biotin labelling reaction’). One volume chloroform was added to the biotin-labelling reaction and the reaction was vortexed and centrifuged at 4ºC (20000g, 5 minutes). The aqueous phase was isolated and the chloroform isolation was repeated twice. NaCl (to 0.5M) and one volume isopropanol was added to the aqueous phase and fragmented RNA was isolated by centrifugation at 4ºC (20000g, 20 minutes). The RNA pellet was washed with 75 percent ethanol, centrifuged at 4ºC (20000g, 20 minutes) and dissolved in nuclease-free water (1μL per 1μg input RNA). Fragmented RNA was denatured at 65 degrees Celcius for 10 minutes prior to incubation with streptavidin beads (μMACs Streptavidin Kit, Miltenyi Biotec, Bergisch Gladbach Germany, 130-074-101) at room temperature for 15 minutes with constant rotation. Biotinylated RNA was isolated using μMACs columns that had been pre-equillibrated with room temperature wash buffer (100mM Tris-HCl pH 7.4, 10mM EDTA, 1M NaCl, 0.1% v/v Tween20). Following incubation with biotinylated RNA / streptavidin beads the columns were washed with 5 times each with 0.9mL pre-warmed (65ºC) and room-temperature wash buffer. Biotinylated RNA was eluted through two additions of 100μL 100mM dithiothreitol (DTT) 3 minutes apart and collected into 700μL RLT buffer from the RNeasy MinElute Cleanup Kit (Qiagen, Hilden Germany, 74204). Biotinylated RNA was isolated using the Qiagen RNeasy MinElute Cleanup Kit using 12μL elution buffer to elute RNA.
TT-SeqSequencing libraries were prepared using NEBNext Ultra II Directional RNA Library Prep Kit.
NEBNext® Ultra II Directional RNA Library Prep Kit for Illumina

treatment_protocol: MV4;11 analog senstive cells were treated with 10 μM of  1-NM-PP1/DMSO for 20 minutes, MV4;11 WT clone cells were treated with a CDK9 inhibtior AZ'5576 for 20 minutes, then 4sU was added for 5 minutes, Schneider 2 cells were labelled with 4sU for 5 minutes.

value: WT


title: 26-Parental-THZ200-2


title: 10-Dual2-D2


title: 12-WT-N2

value: WT Clone


title: 20-Dual2-N2


title: 14-12-N2

value: CDK12AS/NULL Clone


title: 13-12-N1


title: R2-NEW-13-DMSO

description: 4sU labeled RNA


title: R2-NEW-WT-AZ9

description: 4sU labeled RNA


title: R2-NEW-12-NMP

description: 4sU labeled RNA


title: R2-NEW-12-DMSO

description: 4sU labeled RNA


title: 6-13-D2


title: 5-D2-InputPooled

data_processing:
The sequenced reads were demultiplexed using bcl2fastq (v2.17.1.14) and basic QC was performed on the resulting FASTQ files using FASTQC (v0.11.5).
FASTQ reads were mapped to combined hg19/dm3 genome using Bowtie2 (v2.3.3) using the standard settings.
The resulting SAM files were converted to BAM files using Samtools (v 1.4.1) using the view command, after which the BAM files were sorted, indexed and potential PCR duplicates removed using the rmdup function.
Reads were mapped to a combined hg19/dm3 genome after which the reads mapping to fly and human genomes were calculated using FeatureCounts from the Subread package (v1.5.0).
The scaleFactor subquently determined by calculating the reads mapping to the fly genome relative to total mapped reads (combined hg19/dm3 reference) and applied in the BamCoverage command (--normalizeUsing CPM --smoothLength 150 --binSize 50 -e 200 scaleFactor (as calculated)).
Genome_build: hg19 & dm3
Supplementary_files_format_and_content: bigWig files of Pol II and normalised PolII Ser2 reads coverage across the genome in CPM.

extract_protocol:
Cells were crossed-linked with formaldehylde solution. Then nuclear extracts were isolated, and sonicated. Chromatin bound to phospho Ser2 Pol II was isolated using RNA polymerase II (phospho CTD Ser-2), clone 3E10, rat monoclonal, and chromatin bound to total Pol II was isolated uisng RNA polymerase II, clone CTD4H8, mouse monoclonal.
ChIP-Seq libraries were prepared using the NEBNext Ultra II DNA Library Prep Kit for Illumina (New England Biolabs, E7645). Fragments between 200 and 500 base pairs were selected using a Pippin Prep 1.5% agarose cassette (Sage Science, Beverly MA US), pooled and 75 single-end base pair reads were sequenced using the Illumina NEXTseq 500.
NEBNext Ultra II DNA Library Prep Kit for Illumina (New England Biolabs, E7645)

treatment_protocol: Cells were treated with 10 μM of  1-NM-PP1/DMSO for 4 hours,


title: 3-12-D1


title: 4-12-D2


title: R2-NEW-D2-DMSO

description: 4sU labeled RNA


title: R2-NEW-D2-NMP

description: 4sU labeled RNA


title: R2-NEW-13-NMP

description: 4sU labeled RNA


title: 11-WT-N1


title: R2-NEW-WT-DMSO

description: 4sU labeled RNA


title: 8-Dual1-D2


title: R2-NEW-D1-NMP

description: 4sU labeled RNA


title: 5-13-D1


title: 7-Dual1-D1


title: 9-Dual2-D1


title: 1-WT-D1


title: 4-D2plusS2


title: 3-D2minusS2


title: 1-D2minusP

value: RNA polymerase II, clone CTD4H8, mouse monoclonal


title: 16-DplusS2


title: 13-13minusS2


title: 10-WTplusS2


title: 2-WT-D2


title: R2-NEW-D1-DMSO

description: 4sU labeled RNA


title: 2-D2plusP


title: 9-WTminusS2


title: 14-13plusS2


title: 11-12minusS2


title: 5-13minusP


title: 12-12plusS2


title: 15-DminusS2


title: 7-DminusP


title: 8-DplusP


title: 3-12minusP


title: 6-13plusP


title: Ovule (mix)

city: Yangzhou

first: Li

last: Wang

address: Wenhui east road no. 48

institute: Yangzhou University

data_processing:
the NGS QC Toolkit v2.3.3 was used to filiter the reads with adapter sequences, reads with an N (unknown bases in a read＞5%) and low-quality reads (reads with a base quality ≤ 20).
Sequence alignments were performed using TopHat v2.0.12 with v2.2.3 for mapping to the reference genome (http://gigadb.org/dataset/100209, Guan et al.  2016).
The Cufflinks v2.1.1 Reference Annotation Based Transcript (RABT) assembly method was used to construct and identify both known and novel transcripts from TopHat alignment results.
FPKM (expected number of Fragments Per Kilobase of transcript sequence per Millions base pairs sequenced) of each gene was calculated based on the length of the gene and reads count mapped to this gene
sRNA libraries were sequenced using the Illumina HiSeq 4000. Raw data of fastq format were firstly processed through custom perl and python scripts.
The small RNA tags were mapped to reference sequence by Bowtie (Langmead et al., 2009) without mismatch to analyze the expression and distribution on the reference.miRBase22.0 was used as reference, and modified software mirdeep2 and srna-tools-cli were used to obtain the potential miRNA and draw the secondary structures. Custom scripts were used to obtain the miRNA counts.The mapped sRNA tags were also mapped to Rfam, RepeatMasker, to annotate the tags and remove those originating from proteincoding genes, repeat sequences, ribosomal RNAs (rRNAs), transfer RNAs (tRNAs), small nuclear RNAs (snRNAs), and small nucleolar RNAs (snoRNAs). The remaining unannotated tags were used to predict novel miRNAs by exploring secondary structures, Dicer cleavage sites, and minimum free energies using the miREvo and miRDeep2 software packages.
RNA fragments with a poly (A) tail were isolated from the total RNA of mixed samples. A series of steps, including ligation of 5′RNA adaptors, digestion, ligation of double-stranded DNA adaptor, purification, and amplification were conducted and sequencing was done on an Illumina HiSeq 2500. The raw sequencing reads were processed by Illumina’s Pipeline v1.5 to obtain clean reads, which were then used to identify potentially cleaved targets.
Genome_build: GigaScience GigaDB   (http://gigadb.org/dataset/100209, Guan et al. 2016).
Supplementary_files_format_and_content: Text files include gene.FPKM (gene_id, PD1.FPKM, PD2.FPKM, PN1.FPKM, PN1.FPKM), miRNA.readcount_TPM (miRNA.readcount, Ov1.readcount, Ov2.readcount, Ov1.tpm, Ov2.tpm) and miRNA.targets (miRNA_id, target_id)

organism: Ginkgo biloba

source_name: ovule_mix of two development stages

growth_protocol: Ovules were obtained from 3-5 healthy G. biloba trees (~20-year-old; Fozhi cultivar) growing under natural conditions in Yangzhou University, Yangzhou, China (32°20ʹ N, 119°30ʹ E).

extract_protocol:
Samples were collected in early April 2017. Total RNAs of ovules were extracted using MiniBEST Plant RNA Extraction Kit (Takara, Dalian, Liaoning, China), according to the manufacturer’s protocol.
RNA libraries were prepared for sequencing using standard Illumina protocols

tag: tissue

value: ovule_mix of two development stages


title: 1-WTminusP


title: 2-WTplusP


title: 4-12plusP


title: ColonCancer 084

status: Public on Apr 24 2020

city: Kobe

first: Yuichiro

last: Yoshida

address: 4-4-4 Takatsukadai Nishi-ku

department: central research laboratories

institute: Sysmex corporation

description: Frozen sections with tumor burden > 60% were prepared from each specimen and subjected to DNA microarray measurement

hyb_protocol: standard Affymetrix procedures

scan_protocol: standard Affymetrix procedures

data_processing: Data was normalized with Microarray Suite (MAS) 5.0 in the Expression console (Affymetrix).

source_name: Frozen tissue of primary colon cancer

label_protocol: Approximately 100ng of total RNA was processed to produce biotinylated cRNA targets using 3’IVT express kit (Affymetrix)

extract_protocol: Total RNA was extracted from cancer tissue using RNeasy Mini Kit (QIAGEN).

tag: Stage

tag: chemotherapy

value: Y

tag: budding (p)

tag: budding signature score

value: N

value: MSS

value: colon cancer


title: ColonCancer 082


title: ColonCancer 077


title: ColonCancer 083


title: ColonCancer 085


title: ColonCancer 073


title: ColonCancer 079


title: ColonCancer 080


title: ColonCancer 078


title: ColonCancer 074


title: ColonCancer 075


title: ColonCancer 081


title: ColonCancer 066


title: ColonCancer 069


title: ColonCancer 067


title: ColonCancer 063LCM-F

description: The tumor center regions dissected from frozen whole tissue sections were prepared from each specimen and subjected to DNA microarray measurement


title: ColonCancer 071


title: ColonCancer 076


title: ColonCancer 065


title: ColonCancer 064


title: ColonCancer 063LCM-B

description: The invasive front regions of the tumor dissected from frozen whole tissue sections were prepared from each specimen and subjected to DNA microarray measurement


title: ColonCancer 063


title: ColonCancer 061


title: ColonCancer 068


title: ColonCancer 059


title: ColonCancer 058


title: ColonCancer 070


title: ColonCancer 062


title: ColonCancer 072


title: ColonCancer 060


title: ColonCancer 053LCM-F


title: ColonCancer 055


title: ColonCancer 054


title: ColonCancer 045


title: ColonCancer 042


title: ColonCancer 053LCM-B


title: ColonCancer 044


title: ColonCancer 052


title: ColonCancer 050


title: ColonCancer 049


title: ColonCancer 053


title: ColonCancer 051


title: ColonCancer 057


title: ColonCancer 048


title: ColonCancer 043


title: ColonCancer 056


title: ColonCancer 038


title: ColonCancer 039


title: ColonCancer 037


title: ColonCancer 047


title: ColonCancer 036


title: ColonCancer 040


title: ColonCancer 035LCM-B


title: ColonCancer 035LCM-F


title: ColonCancer 041


title: ColonCancer 033


title: ColonCancer 030


title: ColonCancer 029


title: ColonCancer 028


title: ColonCancer 034


title: ColonCancer 046


title: ColonCancer 035


title: ColonCancer 020


title: ColonCancer 031


title: ColonCancer 026


title: ColonCancer 021


title: ColonCancer 019


title: ColonCancer 032


title: ColonCancer 023


title: ColonCancer 025


title: ColonCancer 027


title: ColonCancer 022


title: ColonCancer 024


title: Control-ZCY miRNA-Seq

status: Public on Apr 23 2020

city: Nantong

first: Yunzhao

last: Xu

state: China

address: Qixiu Road

department: Prenatal Diagnosis Center

institute: Affiliated Hospital of Nantong University

description: All.counts.smallRNA.txt

data_processing:
mRNA
We applied the reads filtering towards the raw reads after sequencing to achieve the clean data following the criteria: a)10% base quality <15 b) 13% base quality <20.
Hisat2 2.0.4 mapping to reference genome
count counting
miRNA
We applied the reads filtering towards the raw reads after sequencing to achieve the clean data following the criteria: a) 30% base quality <13 b) read length < 17bp c) adaptor sequence.
Utilizing the BWA software, we mapped the clean data to miRBase 22.0 human miRNA database and piRNAclusterDB+NCBI human piRNA database and RNAcentral human rRNA database and GtRNAdb human tRNA database and MINTBase+tRFdb human tsRNA database.
count counting
Genome_build: GRCh_38
Supplementary_files_format_and_content: TXT file contain counts for each sample

source_name: normal control testis

extract_protocol:
prefrontal cortex were rharvested , flash frozen on dry ice, and RNA was harvested using Trizol reagent.
RNA libraries were prepared for sequencing using standard Illumina protocols

value: normal control testis

tag: disease

value: testicular biopsy with testicular contusion and laceration


title: Control-ZJC miRNA-Seq

description: All.counts.smallRNA.txt


title: Control-QZY miRNA-Seq

description: All.counts.smallRNA.txt

value: testicular biopsy with testicular necrosis


title: ColonCancer 018


title: XCY miRNA-Seq

description: All.counts.smallRNA.txt

source_name: cryptorchidism

value: cryptorchidism

value: testicular biopsy with unilateralis cryptorchidism


title: YXH miRNA-Seq

description: All.counts.smallRNA.txt


title: ColonCancer 016


title: ColonCancer 014


title: GCY miRNA-Seq

description: All.counts.smallRNA.txt


title: CBB miRNA-Seq

description: All.counts.smallRNA.txt


title: ColonCancer 005


title: ColonCancer 004


title: ColonCancer 001


title: ColonCancer 003


title: Control-QZY RNA-seq

description: All.counts.transcriptome.txt


title: ColonCancer 012


title: ColonCancer 017


title: ColonCancer 015


title: ColonCancer 009


title: Control-ZJC RNA-seq

description: All.counts.transcriptome.txt


title: ColonCancer 010


title: ColonCancer 006


title: ColonCancer 007


title: ColonCancer 008


title: ColonCancer 011


title: XZY miRNA-Seq

description: All.counts.smallRNA.txt

value: testicular biopsy with bilateral cryptorchidism


title: YXH RNA-seq

description: All.counts.transcriptome.txt


title: XZY RNA-seq

description: All.counts.transcriptome.txt


title: DZX miRNA-Seq

description: All.counts.smallRNA.txt


title: GCY RNA-seq

description: All.counts.transcriptome.txt


title: CBB RNA-seq

description: All.counts.transcriptome.txt


title: DZX RNA-seq

description: All.counts.transcriptome.txt


title: GM03620_HTT_SNP_chr4_3471519

status: Public on Apr 07 2020

city: Lausanne

first: Gustavo

middle: Agustín

last: Ruiz Buendía

state: Vaud

address: University of Lausanne

department: Center for Integrative Genomics

country: Switzerland

institute: University of Lausanne

description: Sequencing data from GM03620 (HD-B) of a PCR amplicon that contains a potential SNP in position chr4: 3471519

data_processing:
The reads were aligned to the GRCh38 human reference genome using bowtie 2 (Langmead & Salzberg, 2012) and the sequencing depth for each nucleotide was calculated using samtools.
The sequencing data from the genomic regions in unaffected and HD/DM1 patient cell lines was used to validate biallelic SNPs identified from the corresponding 4C-seq data.
Genome_build: hg38
Supplementary_files_format_and_content: VCF files that contain the sequencing depth for each position in the PCR-amplified regions that contained the putative SNPs identified from 4C-seq data.

growth_protocol: All LCLs were obtained from the Coriell Institute for Medical Research Cell Repository. They were grown at 37ºC and 5% CO2 in RPMI 1640 medium supplemented with 15% FBS, 2 mM L-glutamine, and 1% penicillin-streptomycin. Cells were counted and passaged every 3-4 days depending on cell density.

extract_protocol:
Genomic DNA was isolated from the UN-B, UN-C, HD-A, HD-B, and DM1-A cell lines. We used it for PCR amplification of the genomic region encompassing the variants in the parental and offspring LCLs.
2 x 250 bp paired-end sequencing was performed with Illumina Miseq nano

value: Lymphoblastoid Cell Line

value: HD


title: ColonCancer 013


title: GM03620_HTT_SNP_chr4_3367372

description: Sequencing data from GM03620 (HD-B) of a PCR amplicon that contains a potential SNP in position chr4: 3367372


title: GM03620_HTT_SNP_chr4_3366537

description: Sequencing data from GM03620 (HD-B) of a PCR amplicon that contains a potential SNP in position chr4: 3366537


title: GM03620_HTT_SNP_chr4_3239422

description: Sequencing data from GM03620 (HD-B) of a PCR amplicon that contains a potential SNP in position chr4: 3239422


title: Control-ZCY RNA-seq

description: All.counts.transcriptome.txt


title: GM03620_HTT_SNP_chr4_3238432

description: Sequencing data from GM03620 (HD-B) of a PCR amplicon that contains a potential SNP in position chr4: 3238432


title: GM03620_HTT_SNP_chr4_3212105

description: Sequencing data from GM03620 (HD-B) of a PCR amplicon that contains a potential SNP in position chr4: 3212105


title: GM03620_HTT_SNP_chr4_3195217

description: Sequencing data from GM03620 (HD-B) of a PCR amplicon that contains a potential SNP in position chr4: 3195217


title: ColonCancer 002


title: GM03620_HTT_SNP_chr4_2980555

description: Sequencing data from GM03620 (HD-B) of a PCR amplicon that contains a potential SNP in position chr4: 2980555


title: GM03620_HTT_SNP_chr4_2831799

description: Sequencing data from GM03620 (HD-B) of a PCR amplicon that contains a potential SNP in position chr4: 2831799


title: GM03620_HTT_SNP_chr4_3182737

description: Sequencing data from GM03620 (HD-B) of a PCR amplicon that contains a potential SNP in position chr4: 3182737


title: GM03620_HTT_SNP_chr4_2886895

description: Sequencing data from GM03620 (HD-B) of a PCR amplicon that contains a potential SNP in position chr4: 2886895


title: GM03620_HTT_SNP_chr4_3160329

description: Sequencing data from GM03620 (HD-B) of a PCR amplicon that contains a potential SNP in position chr4: 3160329


title: GM03620_HTT_SNP_chr4_2820562

description: Sequencing data from GM03620 (HD-B) of a PCR amplicon that contains a potential SNP in position chr4: 2820562


title: GM03620_HTT_SNP_chr4_2817336

description: Sequencing data from GM03620 (HD-B) of a PCR amplicon that contains a potential SNP in position chr4: 2817336


title: XCY RNA-seq

description: All.counts.transcriptome.txt


title: GM03620_ACTA1_SNP_chr1_229431049

description: Sequencing data from GM03620 (HD-B) of a PCR amplicon that contains a potential SNP in position


title: GM03620_ACTA1_SNP_chr1_229409333

description: Sequencing data from GM03620 (HD-B) of a PCR amplicon that contains a potential SNP in position chr1: 229409333


title: GM03620_ACTA1_SNP_chr1_229431047

description: Sequencing data from GM03620 (HD-B) of a PCR amplicon that contains a potential SNP in position chr1: 229431047


title: GM03620_ACTA1_SNP_chr1_229286446

description: Sequencing data from GM03620 (HD-B) of a PCR amplicon that contains a potential SNP in position chr1: 229286446


title: GM03620_ACTA1_SNP_chr1_229324609

description: Sequencing data from GM03620 (HD-B) of a PCR amplicon that contains a potential SNP in position chr1: 229324609


title: GM03620_ACTA1_SNP_chr1_229276495

description: Sequencing data from GM03620 (HD-B) of a PCR amplicon that contains a potential SNP in position chr1: 229276495


title: GM03620_HTT_SNP_chr4_2797269

description: Sequencing data from GM03620 (HD-B) of a PCR amplicon that contains a potential SNP in position chr4: 2797269


title: GM02164_HTT_SNP_chr4_3238432

description: Sequencing data from GM02164 (HD-A) of a PCR amplicon that contains a potential SNP in position chr4: 3238432


title: GM02164_HTT_SNP_chr4_3367372

description: Sequencing data from GM02164 (HD-A) of a PCR amplicon that contains a potential SNP in position chr4: 3367372


title: GM02164_HTT_SNP_chr4_3366605

description: Sequencing data from GM02164 (HD-A) of a PCR amplicon that contains a potential SNP in position chr4: 3366605


title: GM02164_HTT_SNP_chr4_3366537

description: Sequencing data from GM02164 (HD-A) of a PCR amplicon that contains a potential SNP in position chr4: 3366537


title: GM02164_HTT_SNP_chr4_3239422

description: Sequencing data from GM02164 (HD-A) of a PCR amplicon that contains a potential SNP in position chr4: 3239422


title: GM03620_ACTA1_SNP_chr1_229425528

description: Sequencing data from GM03620 (HD-B) of a PCR amplicon that contains a potential SNP in position chr1: 229425528


title: GM03620_HTT_SNP_chr4_3366605

description: Sequencing data from GM03620 (HD-B) of a PCR amplicon that contains a potential SNP in position chr4: 3366605


title: GM03620_ACTA1_SNP_chr1_229335452

description: Sequencing data from GM03620 (HD-B) of a PCR amplicon that contains a potential SNP in position chr1: 229335452


title: GM02164_HTT_SNP_chr4_3195217

description: Sequencing data from GM02164 (HD-A) of a PCR amplicon that contains a potential SNP in position chr4: 3195217


title: GM02164_HTT_SNP_chr4_3182737

description: Sequencing data from GM02164 (HD-A) of a PCR amplicon that contains a potential SNP in position chr4: 3182737


title: GM02164_HTT_SNP_chr4_3212105

description: Sequencing data from GM02164 (HD-A) of a PCR amplicon that contains a potential SNP in position chr4: 3212105


title: GM02164_HTT_SNP_chr4_3160329

description: Sequencing data from GM02164 (HD-A) of a PCR amplicon that contains a potential SNP in position chr4: 3160329


title: GM02164_HTT_SNP_chr4_3471519

description: Sequencing data from GM02164 (HD-A) of a PCR amplicon that contains a potential SNP in position chr4: 3471519


title: GM02164_HTT_SNP_chr4_2831799

description: Sequencing data from GM02164 (HD-A) of a PCR amplicon that contains a potential SNP in position chr4: 2831799


title: GM02164_HTT_SNP_chr4_2980555

description: Sequencing data from GM02164 (HD-A) of a PCR amplicon that contains a potential SNP in position chr4: 2980555


title: GM02164_HTT_SNP_chr4_2817336

description: Sequencing data from GM02164 (HD-A) of a PCR amplicon that contains a potential SNP in position chr4: 2817336


title: GM02164_HTT_SNP_chr4_2820562

description: Sequencing data from GM02164 (HD-A) of a PCR amplicon that contains a potential SNP in position chr4: 2820562


title: GM02164_ACTA1_SNP_chr1_229431047

description: Sequencing data from GM02164 (HD-A) of a PCR amplicon that contains a potential SNP in position chr1: 229431047


title: GM03620_ACTA1_SNP_chr1_229253648

description: Sequencing data from GM03620 (HD-B) of a PCR amplicon that contains a potential SNP in position chr1: 229253648


title: GM03620_ACTA1_SNP_chr1_229205066

description: Sequencing data from GM03620 (HD-B) of a PCR amplicon that contains a potential SNP in position chr1: 229205066


title: GM02164_ACTA1_SNP_chr1_229409333

description: Sequencing data from GM02164 (HD-A) of a PCR amplicon that contains a potential SNP in position chr1: 229409333


title: GM02164_ACTA1_SNP_chr1_229324609

description: Sequencing data from GM02164 (HD-A) of a PCR amplicon that contains a potential SNP in position chr1: 229324609


title: GM02164_ACTA1_SNP_chr1_229425528

description: Sequencing data from GM02164 (HD-A) of a PCR amplicon that contains a potential SNP in position chr1: 229425528


title: GM02164_ACTA1_SNP_chr1_229286446

description: Sequencing data from GM02164 (HD-A) of a PCR amplicon that contains a potential SNP in position chr1: 229286446


title: GM02164_ACTA1_SNP_chr1_229335452

description: Sequencing data from GM02164 (HD-A) of a PCR amplicon that contains a potential SNP in position chr1: 229335452


title: GM02164_ACTA1_SNP_chr1_229276495

description: Sequencing data from GM02164 (HD-A) of a PCR amplicon that contains a potential SNP in position chr1: 229276495


title: GM02164_ACTA1_SNP_chr1_229253648

description: Sequencing data from GM02164 (HD-A) of a PCR amplicon that contains a potential SNP in position chr1: 229253648


title: GM02164_ACTA1_SNP_chr1_229205066

description: Sequencing data from GM02164 (HD-A) of a PCR amplicon that contains a potential SNP in position chr1: 229205066


title: GM02180_HTT_SNP_chr4_3366537

description: Sequencing data from GM02180 (UN-C) of a PCR amplicon that contains a potential SNP in position chr4: 3366537

value: Unaffected


title: GM02180_HTT_SNP_chr4_3238432

description: Sequencing data from GM02180 (UN-C) of a PCR amplicon that contains a potential SNP in position chr4: 3238432


title: GM02180_HTT_SNP_chr4_3212105

description: Sequencing data from GM02180 (UN-C) of a PCR amplicon that contains a potential SNP in position chr4: 3212105


title: GM02180_HTT_SNP_chr4_3239422

description: Sequencing data from GM02180 (UN-C) of a PCR amplicon that contains a potential SNP in position chr4: 3239422


title: GM02180_HTT_SNP_chr4_3182737

description: Sequencing data from GM02180 (UN-C) of a PCR amplicon that contains a potential SNP in position chr4: 3182737


title: GM02180_HTT_SNP_chr4_3160329

description: Sequencing data from GM02180 (UN-C) of a PCR amplicon that contains a potential SNP in position chr4: 3160329


title: GM02180_HTT_SNP_chr4_3195217

description: Sequencing data from GM02180 (UN-C) of a PCR amplicon that contains a potential SNP in position chr4: 3195217


title: GM02164_HTT_SNP_chr4_2886895

description: Sequencing data from GM02164 (HD-A) of a PCR amplicon that contains a potential SNP in position chr4: 2886895


title: GM02180_HTT_SNP_chr4_2831799

description: Sequencing data from GM02180 (UN-C) of a PCR amplicon that contains a potential SNP in position chr4: 2831799


title: GM02164_HTT_SNP_chr4_2797269

description: Sequencing data from GM02164 (HD-A) of a PCR amplicon that contains a potential SNP in position chr4: 2797269


title: GM02180_HTT_SNP_chr4_2886895

description: Sequencing data from GM02180 (UN-C) of a PCR amplicon that contains a potential SNP in position chr4: 2886895


title: GM02180_HTT_SNP_chr4_2817336

description: Sequencing data from GM02180 (UN-C) of a PCR amplicon that contains a potential SNP in position chr4: 2817336


title: GM02164_ACTA1_SNP_chr1_229431049

description: Sequencing data from GM02164 (HD-A) of a PCR amplicon that contains a potential SNP in position chr1: 229431049


title: GM02180_HTT_SNP_chr4_2820562

description: Sequencing data from GM02180 (UN-C) of a PCR amplicon that contains a potential SNP in position chr4: 2820562


title: GM02180_ACTA1_SNP_chr1_229431049

description: Sequencing data from GM02180 (UN-C) of a PCR amplicon that contains a potential SNP in position chr1: 229431049


title: GM02180_ACTA1_SNP_chr1_229431047

description: Sequencing data from GM02180 (UN-C) of a PCR amplicon that contains a potential SNP in position chr1: 229431047


title: GM02180_HTT_SNP_chr4_2980555

description: Sequencing data from GM02180 (UN-C) of a PCR amplicon that contains a potential SNP in position chr4: 2980555


title: GM02180_ACTA1_SNP_chr1_229425528

description: Sequencing data from GM02180 (UN-C) of a PCR amplicon that contains a potential SNP in position chr1: 229425528


title: GM02180_ACTA1_SNP_chr1_229409333

description: Sequencing data from GM02180 (UN-C) of a PCR amplicon that contains a potential SNP in position chr1: 229409333


title: GM02180_ACTA1_SNP_chr1_229335452

description: Sequencing data from GM02180 (UN-C) of a PCR amplicon that contains a potential SNP in position chr1: 229335452


title: GM02180_ACTA1_SNP_chr1_229286446

description: Sequencing data from GM02180 (UN-C) of a PCR amplicon that contains a potential SNP in position chr1: 229286446


title: GM02180_ACTA1_SNP_chr1_229324609

description: Sequencing data from GM02180 (UN-C) of a PCR amplicon that contains a potential SNP in position chr1: 229324609


title: GM02180_HTT_SNP_chr4_3367372

description: Sequencing data from GM02180 (UN-C) of a PCR amplicon that contains a potential SNP in position chr4: 3367372


title: GM02180_HTT_SNP_chr4_3366605

description: Sequencing data from GM02180 (UN-C) of a PCR amplicon that contains a potential SNP in position chr4: 3366605


title: GM02180_HTT_SNP_chr4_3471519

description: Sequencing data from GM02180 (UN-C) of a PCR amplicon that contains a potential SNP in position chr4: 3471519


title: GM02180_ACTA1_SNP_chr1_229253648

description: Sequencing data from GM02180 (UN-C) of a PCR amplicon that contains a potential SNP in position chr1: 229253648


title: GM02180_ACTA1_SNP_chr1_229205066

description: Sequencing data from GM02180 (UN-C) of a PCR amplicon that contains a potential SNP in position chr1: 229205066


title: GM04604_ACTA1_SNP_chr1_229431052

description: Sequencing data from GM04604 (UN-B) of a PCR amplicon that contains a potential SNP in position chr1: 229431052


title: GM02180_HTT_SNP_chr4_2797269

description: Sequencing data from GM02180 (UN-C) of a PCR amplicon that contains a potential SNP in position chr4: 2797269


title: GM04604_DMPK_SNP_chr19_45938336

description: Sequencing data from GM04604 (UN-B) of a PCR amplicon that contains a potential SNP in position chr19: 45938336


title: GM04604_ACTA1_SNP_chr1_229376916

description: Sequencing data from GM04604 (UN-B) of a PCR amplicon that contains a potential SNP in position chr1: 229376916


title: GM04604_DMPK_SNP_chr19_45759028

description: Sequencing data from GM04604 (UN-B) of a PCR amplicon that contains a potential SNP in position chr19: 45759028


title: GM04604_DMPK_SNP_chr19_45824451

description: Sequencing data from GM04604 (UN-B) of a PCR amplicon that contains a potential SNP in position chr19: 45824451


title: GM04604_DMPK_SNP_chr19_45838566

description: Sequencing data from GM04604 (UN-B) of a PCR amplicon that contains a potential SNP in position chr19: 45838566


title: GM04604_DMPK_SNP_chr19_45816157

description: Sequencing data from GM04604 (UN-B) of a PCR amplicon that contains a potential SNP in position chr19: 45816157


title: GM04604_DMPK_SNP_chr19_45819221

description: Sequencing data from GM04604 (UN-B) of a PCR amplicon that contains a potential SNP in position chr19: 45819221


title: GM04604_DMPK_SNP_chr19_45766055

description: Sequencing data from GM04604 (UN-B) of a PCR amplicon that contains a potential SNP in position chr19: 45766055


title: GM04604_DMPK_SNP_chr19_45793475

description: Sequencing data from GM04604 (UN-B) of a PCR amplicon that contains a potential SNP in position chr19: 45793475


title: GM04604_DMPK_SNP_chr19_45777111

description: Sequencing data from GM04604 (UN-B) of a PCR amplicon that contains a potential SNP in position chr19: 45777111


title: GM04604_DMPK_SNP_chr19_45824412

description: Sequencing data from GM04604 (UN-B) of a PCR amplicon that contains a potential SNP in position chr19: 45824412


title: GM04604_DMPK_SNP_chr19_45688664

description: Sequencing data from GM04604 (UN-B) of a PCR amplicon that contains a potential SNP in position chr19: 45688664


title: GM02180_ACTA1_SNP_chr1_229276495

description: Sequencing data from GM02180 (UN-C) of a PCR amplicon that contains a potential SNP in position chr1: 229276495


title: GM04604_ACTA1_SNP_chr1_229431048

description: Sequencing data from GM04604 (UN-B) of a PCR amplicon that contains a potential SNP in position chr1: 229431048


title: GM04604_ACTA1_SNP_chr1_229280321

description: Sequencing data from GM04604 (UN-B) of a PCR amplicon that contains a potential SNP in position chr1: 229280321


title: GM04604_DMPK_SNP_chr19_45528896

description: Sequencing data from GM04604 (UN-B) of a PCR amplicon that contains a potential SNP in position chr19: 45528896


title: GM04604_ACTA1_SNP_chr1_229431047

description: Sequencing data from GM04604 (UN-B) of a PCR amplicon that contains a potential SNP in position chr1: 229431047


title: GM04604_ACTA1_SNP_chr1_229276465

description: Sequencing data from GM04604 (UN-B) of a PCR amplicon that contains a potential SNP in position chr1: 229276465


title: GM04604_ACTA1_SNP_chr1_229128358

description: Sequencing data from GM04604 (UN-B) of a PCR amplicon that contains a potential SNP in position chr1: 229128358


title: GM04604_ACTA1_SNP_chr1_229627320

description: Sequencing data from GM04604 (UN-B) of a PCR amplicon that contains a potential SNP in position chr1: 229627320


title: GM06077_ACTA1_SNP_chr1_229431052

description: Sequencing data from GM06077 (DM1-A) of a PCR amplicon that contains a potential SNP in position chr1: 229431052


title: GM04604_ACTA1_SNP_chr1_229276461

description: Sequencing data from GM04604 (UN-B) of a PCR amplicon that contains a potential SNP in position chr1: 229276461


title: GM06077_ACTA1_SNP_chr1_229399146

description: Sequencing data from GM06077 (DM1-A) of a PCR amplicon that contains a potential SNP in position chr1: 229399146


title: GM06077_ACTA1_SNP_chr1_229376916

description: Sequencing data from GM06077 (DM1-A) of a PCR amplicon that contains a potential SNP in position chr1: 229376916


title: GM04604_ACTA1_SNP_chr1_229399146

description: Sequencing data from GM04604 (UN-B) of a PCR amplicon that contains a potential SNP in position chr1: 229399146


title: GM06077_DMPK_SNP_chr19_45938336

description: Sequencing data from GM06077 (DM1-A) of a PCR amplicon that contains a potential SNP in position chr19: 45938336


title: GM06077_DMPK_SNP_chr19_45759028

description: Sequencing data from GM06077 (DM1-A) of a PCR amplicon that contains a potential SNP in position chr19: 45759028


title: GM06077_DMPK_SNP_chr19_45838566

description: Sequencing data from GM06077 (DM1-A) of a PCR amplicon that contains a potential SNP in position chr19: 45838566


title: GM04604_DMPK_SNP_chr19_45782080

description: Sequencing data from GM04604 (UN-B) of a PCR amplicon that contains a potential SNP in position chr19: 45782080


title: GM06077_DMPK_SNP_chr19_45824451

description: Sequencing data from GM06077 (DM1-A) of a PCR amplicon that contains a potential SNP in position chr19: 45824451


title: GM06077_DMPK_SNP_chr19_45824412

description: Sequencing data from GM06077 (DM1-A) of a PCR amplicon that contains a potential SNP in position chr19: 45824412


title: GM06077_DMPK_SNP_chr19_45819221

description: Sequencing data from GM06077 (DM1-A) of a PCR amplicon that contains a potential SNP in position chr19: 45819221


title: GM06077_DMPK_SNP_chr19_45816157

description: Sequencing data from GM06077 (DM1-A) of a PCR amplicon that contains a potential SNP in position chr19: 45816157


title: GM04604_DMPK_SNP_chr19_45531422

description: Sequencing data from GM04604 (UN-B) of a PCR amplicon that contains a potential SNP in position chr19: 45531422


title: GM06077_DMPK_SNP_chr19_45782080

description: Sequencing data from GM06077 (DM1-A) of a PCR amplicon that contains a potential SNP in position chr19: 45782080


title: GM06077_DMPK_SNP_chr19_45766055

description: Sequencing data from GM06077 (DM1-A) of a PCR amplicon that contains a potential SNP in position chr19: 45766055


title: GM06077_DMPK_SNP_chr19_45777111

description: Sequencing data from GM06077 (DM1-A) of a PCR amplicon that contains a potential SNP in position chr19: 45777111


title: GM06077_ACTA1_SNP_chr1_229431047

description: Sequencing data from GM06077 (DM1-A) of a PCR amplicon that contains a potential SNP in position chr1: 229431047


title: GM06077_DMPK_SNP_chr19_45531422

description: Sequencing data from GM06077 (DM1-A) of a PCR amplicon that contains a potential SNP in position chr19: 45531422


title: GM06077_DMPK_SNP_chr19_45528896

description: Sequencing data from GM06077 (DM1-A) of a PCR amplicon that contains a potential SNP in position chr19: 45528896


title: GM06077_ACTA1_SNP_chr1_229627320

description: Sequencing data from GM06077 (DM1-A) of a PCR amplicon that contains a potential SNP in position chr1: 229627320


title: GM06077_ACTA1_SNP_chr1_229431048

description: Sequencing data from GM06077 (DM1-A) of a PCR amplicon that contains a potential SNP in position chr1: 229431048


title: GM06077_ACTA1_SNP_chr1_229280321

description: Sequencing data from GM06077 (DM1-A) of a PCR amplicon that contains a potential SNP in position chr1: 229280321


title: GM06077_DMPK_SNP_chr19_45688664

description: Sequencing data from GM06077 (DM1-A) of a PCR amplicon that contains a potential SNP in position chr19: 45688664


title: GM06077_ACTA1_SNP_chr1_229276465

description: Sequencing data from GM06077 (DM1-A) of a PCR amplicon that contains a potential SNP in position chr1: 229276465


title: GM06077_ACTA1_SNP_chr1_229276461

description: Sequencing data from GM06077 (DM1-A) of a PCR amplicon that contains a potential SNP in position chr1: 229276461


title: GM04604_input

description: GM04604 input

data_processing:
Data analysis was performed according to the ENCODE Transcription Factor ChIP-seq processing pipeline (phase-3)
ChIP-seq reads were aligned to the GRCh38 human reference genome using bowtie2 (v 2.3.4.3) with the configurations provided by ENCODE pipeline3.
Data were filtered using to remove unmapped reads, reads with no primary alignment, multimapped reads, and duplicate reads.
Peaks were called using SPP (v 1.16.0) with the setting provide by ENCODE pipeline3.
Replicate handling was performed using the Irreproducibility Discovery Rate framework (IDR).
Genome_build: GRCh38 (GCA_000001405.15)
Supplementary_files_format_and_content: bigwig files were generated using deeptools bamCoverage v3.3.0 ; Score represent read counts, extend by 200 with a binsize of 1bp

extract_protocol:
Chromatin immunoprecipitation was performed according to the Diagenode AutoiDeal ChIP-qPCR kit (Diagenode) standard protocol. Samples of 6 x 106 cells were sonicated using the Diagenode Bioruptor Pico (Diagenode), with 10 cycles of 30 sec “on” and 30 sec “off”. Correct DNA fragmentation was verified by agarose gel electrophoresis. Immunoprecipitation was performed with 4 x 10^6 cells using a CTCF antibody (Diagenode).
CTCF ChIP-seq libraries were prepared in triplicate using the Illumina TruSeq ChIP Sample Preparation kit. We performed 1 x 50 bp single-end sequencing with Illumina Hiseq 4000.

tag: chip antibody

value: Diagenode, C15410210


title: Bisulf_seq_downstream_CTCF_GM06077

description: Bisulfite sequencing of an amplicon that contains the CTCF binding site directly downstream of the CTG repeats of DMPK in GM06077 (DM1-A) cells.

data_processing:
Sequencing reads were preprocessed using TrimGalore (github.com/FelixKrueger/TrimGalore) with the following parameters: -q 20 --length 20 –paired. Reads were aligned using QuasR (Gaidatzis et al., 2015) to the GRCh38 human reference DNA sequences corresponding to the amplified PCR products. DNA methylation calls for each CpG were extracted using the qMeth() function in QuasR. DNA methylation frequencies were calculated as methylated CpG reads / total number of reads covering the respective CpG x 100.
Genome_build: hg38
Supplementary_files_format_and_content: The rows of the tabl-separated files represent the CpGs in the respective amplicon. The columns are: seqnames (CpG residue number), start (position of C relative to the amplicon), end (position of G relative to the amplicon), …_T (number of all WGBS reads covering the respective CpG residues), ..._M (number of methylated WGBS reads covering the respective CpG residues).

extract_protocol:
Bisulfite sequencing was performed according to a previously described method (Barbé et al., 2017). Bisulfite conversion was performed with the EZ DNA Methylation kit (Zymo Research) using the standard protocol. 200 ng of genomic DNA were used for bisulfite conversion at 50ºC for 12 hours. Bisulfite-converted DNA was desulphonated, eluted, and immediately used for nested and hemi-nested PCR amplification of the upstream and downstream CTCF binding sites respectively using previously described primers (Barbé et al., 2017).
Libraries were constructed by using 50 ng of bisulfite-converted DNA for the first PCR, and 3 µL of the products were used for the second PCR. The final amplicons contained sequencing adapters and were purified with the NucleoSpin PCR Clean-up kit (Macherey-Nagel) and used for 2 x 250 bp paired-end sequencing on Illumina Miseq.


title: Bisulf_seq_upstream_CTCF_GM04648

description: Bisulfite sequencing of an amplicon that contains the CTCF binding site directly upstream of the CTG repeats of DMPK in GM04648 (DM1-B) cells.


title: Bisulf_seq_downstream_CTCF_GM04604

description: Bisulfite sequencing of an amplicon that contains the CTCF binding site directly downstream of the CTG repeats of DMPK in GM04604 (UN-B) cells.


title: Bisulf_seq_upstream_CTCF_GM06077

description: Bisulfite sequencing of an amplicon that contains the CTCF binding site directly upstream of the CTG repeats of DMPK in GM06077 (DM1-A) cells.


title: GM04648_input

description: GM04648 input


title: Bisulf_seq_downstream_CTCF_GM04648

description: Bisulfite sequencing of an amplicon that contains the CTCF binding site directly downstream of the CTG repeats of DMPK in GM04648 (DM1-B) cells.


title: GM06077_DMPK_SNP_chr19_45793475

description: Sequencing data from GM06077 (DM1-A) of a PCR amplicon that contains a potential SNP in position chr19: 45793475


title: Bisulf_seq_upstream_CTCF_GM04604

description: Bisulfite sequencing of an amplicon that contains the CTCF binding site directly upstream of the CTG repeats of DMPK in GM04604 (UN-B) cells.


title: GM06077_ACTA1_SNP_chr1_229128358

description: Sequencing data from GM06077 (DM1-A) of a PCR amplicon that contains a potential SNP in position chr1: 229128358


title: GM04604_CTCF_IP

description: GM04604 CTCF ChIP


title: GM04648_CTCF_IP

description: GM04648 CTCF ChIP


title: GM06077_input

description: GM06077 input


title: GM06077_CTCF_IP

description: GM06077 CTCF ChIP


title: RNAseq_GM06077_rep1

description: RNA-seq library from GM06077 (DM1-A) replicate 1

data_processing:
RNA-seq reads were demultiplexed using the bcl2fastq Illumina software (v 2.20.0). Purity-filtered reads were trimmed with Cutadapt (v 1.3) (Martin, 2011) and filtered for low complexity with seq_crumbs (v 0.1.8).
Reads were aligned against the GRCh38 human transcriptome (GRCh38.82) using STAR (v 2.4.2a) (Dobin, et al., 2013). Read counts per gene were summarized with htseq-count (v 0.6.1) (Anders, et al., 2015).
Genes differentially expressed in HD and DM1 LCLs compared to unaffected LCLs were determined using DESeq2 (v 1.20.0) (Love, et al., 2014) while accounting for batch effects including cell passage number, collection date, and sequencing runs. P-values were adjusted using the Benjamini-Hochberg multiple testing correction. Significantly differentially expressed genes were defined as those with adjusted p-values less than 0.05 and log2 fold-change greater than 0.5.
Genome_build: hg38
Supplementary_files_format_and_content: Gene count files for each replicate.

extract_protocol:
Total RNA from 5 million cells was extracted using the NucleoSpin RNA kit (Macherey-Nagel) and its concentration was measured by Nanodrop (NanoPhotometer NP80, Implen). RNA quality was assessed on a Fragment Analyzer (Agilent Technologies) and all of the RNA samples had a RQN between 8.7 and 10.
RNA-seq libraries were prepared using 500 ng of total RNA and the Illumina TruSeq Stranded Total RNA Library Prep Kit with Ribo-Zero Gold (Illumina) according to the manufacturer’s protocol. Multiplexed samples were pooled in equimolar amounts and 2 x 100 bp paired-end sequencing was performed on Illumina HiSeq 2500.

tag: passage


title: RNAseq_GM04648_rep2

description: RNA-seq library from GM04648 (DM1-B) replicate 2


title: RNAseq_GM04648_rep1

description: RNA-seq library from GM04648 (DM1-B) replicate 1


title: RNAseq_GM06077_rep2

description: RNA-seq library from GM06077 (DM1-A) replicate 2


title: RNAseq_GM04604_rep1

description: RNA-seq library from GM04604 (UN-B) replicate 1


title: RNAseq_GM02180_rep2

description: RNA-seq library from GM02180 (UN-C) replicate 2


title: RNAseq_GM03620_rep1

description: RNA-seq library from GM03620 (HD-B) replicate 1


title: RNAseq_GM02164_rep2

description: RNA-seq library from GM02164 (HD-B) replicate 2


title: RNAseq_GM03620_rep2

description: RNA-seq library from GM03620 (HD-B) replicate 2


title: RNAseq_GM14044_rep2

description: RNA-seq library from GM14044 (HD-C) replicate 2


title: RNAseq_GM02180_rep1

description: RNA-seq library from GM02180 (UN-C) replicate 1


title: RNAseq_GM14044_rep1

description: RNA-seq library from GM14044 (HD-C) replicate 1


title: RNAseq_GM02164_rep1

description: RNA-seq library from GM02164 (HD-B) replicate 1


title: 4C_HTT_d85_GM14044_rep2

description: GM14044 (HD-C) HTT_d85 4C viewpoint replicate 2

data_processing:
Demultiplexing, trimming, and mapping were performed using the BBCF HTSstation (David et al., 2014). Reads were trimmed to keep the first 40 bp. The 4C fragments surrounding the viewpoints (±2.5 kb) were excluded from the rest of the analysis. The demultiplexed reads were mapped to the GRCh38 human reference genome using bowtie2 (v 2.2) (Langmead & Salzberg, 2012).
Fragment read counts were obtained using FourCSeq (v 1.18.0) (Klein et al., 2015). Fragment counts were normalized (reads per million) and smoothed with a running mean (window size = 5 fragments).
Significant chromatin interactions were identified with two 4C-seq data analysis packages: FourCSeq (v 1.18.0) (Klein et al., 2015) and 4C-ker (v 0.0.0.9000) (Raviram et al., 2016).
For the FourCSeq analysis, we defined significant interactions as fragments with a z-score equal to or greater than 1.96 and a false-discovery rate of 0.1, using the following parameters: minCount=20 and fitFun="distFitMonotone" in getZScores function; zScoreThresh=1.96, fdrThresh = 0.1 in addPeaks function.
For the 4C-ker analysis, we used k=5 in the nearBaitAnalysis function, and plotted the high-interacting regions.
Differential interactions were identified with the differentialAnalysis function of 4C-ker, which is based on the DESeq2 (Love et al., 2014) framework, using default parameters (including a p-value threshold of 0.5). The UN-B cell line was used as the reference condition for all comparisons except for Fig. 1 and Fig. S2 where the reference condition was the UN-A cell line.
For plotting the data, the smoothed and normalized fragment counts were averaged among replicates of the same 4C library samples and visualized with gFeatBrowser (http://www.gfeatbrowser.com).
Genome_build: hg38
Supplementary_files_format_and_content: Smoothed and normalized 4C fragment count data (bedGraph) of the cis chromosome, one per viewpoint and per replicate.

growth_protocol: All LCLs were obtained from the Coriell Institute for Medical Research Cell Repository. They were grown at 37ºC and 5% CO2 in RPMI 1640 medium supplemented with 15% FBS, 2 mM L-glutamine, and 1% penicillin-streptomycin. Cells were counted and passaged every 3-4 days depending on cell density. The GFP(CAG)270 and GFP(CAG)15 cell lines were maintained at 37°C and 5% CO2 in DMEM with Glutamax, 1% penicillin-streptomycin, 15 μg mL−1 blasticidine, and 150 μg/mL hygromycin. Transcription of the GFP mini-gene was activated by culturing GFP(CAG)n cells with doxycycline at a final concentration of 2 µg/mL for five days.

extract_protocol:
Samples of 10 million cells were crosslinked in 2% formaldehyde for 10 minutes at room temperature and quenched with glycine to a final concentration 0.13 M. Cross-linked samples were rinsed once with PBS and were either used immediately or flash-frozen in liquid nitrogen and stored at -80ºC for later use. Cells were lysed for 15 minutes on ice in lysis buffer (50 mM Tris-HCl pH 7.5, 150 mM NaCl, 5 mM EDTA, 0.5% NP-40, 1% Triton X-100, 1x protease inhibitors). The first digestion was performed with 200 U DpnII (NEB) and incubating for 4 hours at 37ºC, then 200 U DpnII and overnight incubation at 37ºC, and finally 200 U DpnII and incubating 4 hours at 37ºC. DpnII digestion efficiency was assessed by agarose gel electrophoresis and DpnII was subsequently heat inactivated. The first ligation was performed at 16ºC overnight with 50 U T4 DNA ligase (Thermo Scientific) in 7 mL. Ligated samples were de-crosslinked with 30 µL Proteinase K (10 mg/mL) at 65ºC overnight followed by RNAse A treatment and phenol-chloroform purification. The second digestion was performed with 50 U of BfaI (NEB) at 37ºC overnight followed by BfaI heat inactivation. The second ligation was performed at 16ºC overnight with 100 U T4 DNA ligase in 14 mL. The resulting 4C template samples were precipitated and purified with the QIAquick PCR purification kit.
4C libraries were generated by amplifying 1 µg total of purified 4C template using the Expand Long Template PCR System (Roche) with 4C viewpoint primers with Illumina sequencing adapters that permitted multiplexing (in 50 µL PCRs), pooling reactions and purifying the PCR products with AMPure XP beads to exclude products less than 130 bp. Single-end sequencing of pooled 4C libraries was performed on Illumina HiSeq 2500.


title: 4C_HTT_d85_GM14044_rep3

description: GM14044 (HD-C) HTT_d85 4C viewpoint replicate 3


title: 4C_HTT_d85_GM14044_rep1

description: GM14044 (HD-C) HTT_d85 4C viewpoint replicate 1


title: 4C_HTT_d85_GM04604_rep3

description: GM04604 (UN-B) HTT_d85 4C viewpoint replicate 3


title: 4C_HTT_d85_GM04604_rep2

description: GM04604 (UN-B) HTT_d85 4C viewpoint replicate 2


title: 4C_HTT_d85_GM03620_rep1

description: GM03620 (HD-B) HTT_d85 4C viewpoint replicate 1


title: 4C_HTT_d85_GM04604_rep1

description: GM04604 (UN-B) HTT_d85 4C viewpoint replicate 1


title: 4C_HTT_d85_GM02180_rep3

description: GM02180 (UN-C) HTT_d85 4C viewpoint replicate 3


title: 4C_HTT_d85_GM03620_rep3

description: GM03620 (HD-B) HTT_d85 4C viewpoint replicate 3


title: 4C_HTT_d85_GM03620_rep2

description: GM03620 (HD-B) HTT_d85 4C viewpoint replicate 2


title: RNAseq_GM04604_rep2

description: RNA-seq library from GM04604 (UN-B) replicate 2


title: 4C_HTT_d85_GM02164_rep2

description: GM02164 (HD-A) HTT_d85 4C viewpoint replicate 2


title: 4C_HTT_d85_GM02164_rep3

description: GM02164 (HD-A) HTT_d85 4C viewpoint replicate 3


title: 4C_HTT_d85_GM02164_rep1

description: GM02164 (HD-A) HTT_d85 4C viewpoint replicate 1


title: 4C_HTT_d1_GM14044_rep2

description: GM14044 (HD-C) HTT_d1 4C viewpoint replicate 2


title: 4C_HTT_d1_GM14044_rep3

description: GM14044 (HD-C) HTT_d1 4C viewpoint replicate 3


title: 4C_HTT_d1_GM04604_rep1

description: GM04604 (UN-B) HTT_d1 4C viewpoint replicate 1


title: 4C_HTT_d1_GM04604_rep3

description: GM04604 (UN-B) HTT_d1 4C viewpoint replicate 3


title: 4C_HTT_d1_GM04604_rep2

description: GM04604 (UN-B) HTT_d1 4C viewpoint replicate 2


title: 4C_HTT_d1_GM02180_rep3

description: GM02180 (UN-C) HTT_d1 4C viewpoint replicate 3


title: 4C_HTT_d1_GM03620_rep1

description: GM03620 (HD-B) HTT_d1 4C viewpoint replicate 1


title: 4C_HTT_d1_GM02180_rep2

description: GM02180 (UN-C) HTT_d1 4C viewpoint replicate 2


title: 4C_HTT_d1_GM02180_rep1

description: GM02180 (UN-C) HTT_d1 4C viewpoint replicate 1


title: 4C_HTT_d1_GM03620_rep3

description: GM03620 (HD-B) HTT_d1 4C viewpoint replicate 3


title: 4C_HTT_d1_GM03620_rep2

description: GM03620 (HD-B) HTT_d1 4C viewpoint replicate 2


title: 4C_HTT_d1_GM02164_rep3

description: GM02164 (HD-A) HTT_d1 4C viewpoint replicate 3


title: 4C_HTT_d1_GM02164_rep2

description: GM02164 (HD-A) HTT_d1 4C viewpoint replicate 2


title: 4C_HTT_d1_GM02164_rep1

description: GM02164 (HD-A) HTT_d1 4C viewpoint replicate 1


title: 4C_HTT_d85_GM02180_rep1

description: GM02180 (UN-C) HTT_d85 4C viewpoint replicate 1


title: 4C_HTT_d85_GM02180_rep2

description: GM02180 (UN-C) HTT_d85 4C viewpoint replicate 2


title: 4C_FMR1_u1_GM09237_rep3

description: GM09237 (FXS) FMR1_u1 4C viewpoint replicate 3

value: FXS


title: 4C_FMR1_u1_GM09237_rep2

description: GM09237 (FXS) FMR1_u1 4C viewpoint replicate 2


title: 4C_FMR1_u1_GM09237_rep1

description: GM09237 (FXS) FMR1_u1 4C viewpoint replicate 1


title: 4C_FMR1_u1_GM09236_rep3

description: GM09236 (UN-A) FMR1_u1 4C viewpoint replicate 3


title: 4C_FMR1_u195_GM09237_rep2

description: GM09237 (FXS) FMR1_u195 4C viewpoint replicate 2


title: 4C_FMR1_u1_GM09236_rep2

description: GM09236 (UN-A) FMR1_u1 4C viewpoint replicate 2


title: 4C_FMR1_u1_GM09236_rep1

description: GM09236 (UN-A) FMR1_u1 4C viewpoint replicate 1


title: 4C_FMR1_u195_GM09236_rep3

description: GM09236 (UN-A) FMR1_u195 4C viewpoint replicate 3


title: 4C_FMR1_u195_GM09237_rep1

description: GM09237 (FXS) FMR1_u195 4C viewpoint replicate 1


title: 4C_FMR1_u195_GM09237_rep3

description: GM09237 (FXS) FMR1_u195 4C viewpoint replicate 3


title: 4C_FMR1_u195_GM09236_rep2

description: GM09236 (UN-A) FMR1_u195 4C viewpoint replicate 2


title: 4C_HTT_d1_GM14044_rep1

description: GM14044 (HD-C) HTT_d1 4C viewpoint replicate 1


title: 4C_DMPK_u65_GM06077_rep2

description: GM06077 (DM1-A) DMPK_u65 4C viewpoint replicate 2


title: 4C_DMPK_u65_GM06077_rep1

description: GM06077 (DM1-A) DMPK_u65 4C viewpoint replicate 1


title: 4C_DMPK_u65_GM04604_rep2

description: GM04604 (UN-B) DMPK_u65 4C viewpoint replicate 2


title: 4C_DMPK_u65_GM04604_rep1

description: GM04604 (UN-B) DMPK_u65 4C viewpoint replicate 1


title: 4C_DMPK_u65_GM04648_rep1

description: GM04648 (DM1-B) DMPK_u65 4C viewpoint replicate 1


title: 4C_DMPK_u16_GM06077_rep1

description: GM06077 (DM1-A) DMPK_u16 4C viewpoint replicate 1


title: 4C_DMPK_u16_GM06077_rep2

description: GM06077 (DM1-A) DMPK_u16 4C viewpoint replicate 2


title: 4C_DMPK_u65_GM02180_rep2

description: GM02180 (UN-C) DMPK_u65 4C viewpoint replicate 2