title: IL2RA low_rep1

description:
cd25.trim.lo.bedgraph: Bedgraph file containing log2-fold enrichment data for each gRNA, averaged across the two replicates
cd25.trim.window5.lo.bedgraph: Bedgraph file containing log2-fold enrichment data for a 5-gRNA sliding window, averaged across the two replicates


title: IL2RA high_rep1

status: Public on Apr 26 2017

type: SRA

city: Berkeley

first: Jacob

middle: Ellery

last: Corn

state: CA

address: 2151 Berkeley Way

department: Innovative Genomics Institute

country: USA

institute: University of California, Berkeley

description:
cd25.trim.hi.bedgraph: Bedgraph file containing log2-fold enrichment data for each gRNA, averaged across the two replicates
cd25.trim.window5.hi.bedgraph: Bedgraph file containing log2-fold enrichment data for a 5-gRNA sliding window, averaged across the two replicates

library_source: genomic

data_processing:
Basecalls were performed with Bcl2Fastq version 2.17
Reads failing CASAVA quality filters were excluded.
Reads were then trimmed of the gRNA common sequence using the cutadapt script and the command “cutadapt -a GTTTAAGAGCTAAGCTG”. Ref: Marcel, M. Cutadapt removes adapter sequences from high-throughput sequencing reads. EMBnet.journal 17, 10–12 (2011).
Reads were then aligned against a database of gRNA sequences containing the gRNA name and the gRNA sequence appended with a 5' G.
Alignment was performed using bowtie2-2.2.6 with options -U --norc --no-hd and requiring full length alignment.
gRNA counts were generated by counting the number of times the guide id occurred in the aligned sam file
sgRNAs with fewer than 50 reads in either of the background samples were removed from all downstream analysis
Read counts for each sgRNA in each sample were then normalized to the total number of sgRNA read counts in that sample. A pseudo-count of 1 was added to all normalized guide counts.
sgRNA enrichment was calculated as follows: mean(log2(gene_gate_rep1/gene_background_rep1),log2(gene_gate_rep2/gene_background_rep2)), e.g. mean(log2(CD69_top_rep1/CD69_background_rep1),log2(CD69_top_rep2/CD69_background_rep2))
All non-targeting control sgRNAs and any sgRNAs that map perfectly to multiple sequences within the sgRNA library window were excluded from visualization.
Genome_build: hg19
Supplementary_files_format_and_content: Individual sgRNA enrichment was visualized in .bedgraph format, plotting the log2-fold enrichment (as indicated above) for each sgRNA. The genomic coordinate used for an individual sgRNA is the 3' end of the PAM for that sgRNA.
Supplementary_files_format_and_content: Windowed sgRNA enrichmend was also visualized in .bedgraph format. The average log2-fold enrichment, and the average genomic coordinate, were calculated for a 5-sgRNA sliding window.

molecule: genomic DNA

organism: Homo sapiens

source_name: Jurkat cells

growth_protocol: Jurkat cells were cultured in RPMI 1640 media supplemented with 10% fetal calf serum, 1 mM Sodium Pyruvate (Gibco), 100 U/mL penicillin (Gibco) and 100 μg/mL streptomycin (Gibco). Cell line identity was authenticated by short tandem repeat (STR) analysis and verified mycoplasma free using the MycoAlert Mycoplasma Detection Kit (Lonza). Jurkat cells were passaged every 2-3 days and maintained at 0.1-2.0x10^6 cells/mL of media. Jurkat cells were transduced with a lentiviral dCas9-VP64-2A-GFP expression vector (Addgene 61422). Single GFP+ cells were sorted by FACS into the wells of a 96-well plate, and a clone with bright uniform GFP expression were selected for use in these experiments.

extract_protocol:
Genomic DNA was isolated from sorted cells using NucleoSpin Blood kits (Macherey-Nagel), or by Proteinase K digestion and isopropanol precipitation for samples with fewer than 106 cells.
PCR was used to amplify sgRNA cassettes with Illumina sequencing adapters and indexes as described in Kampmann, et al (Nature Protocol, 9, 1825-1847 (2014). Genomic DNA samples containing less than 10 μg of gDNA were loaded directly into PCR. For genomic DNA samples containing more than 10 μg of DNA, samples were first digested for 18 hours with SbfI-HF (NEB) to liberate a ~500 bp fragment containing the gRNA cassette. The amplified sgRNA cassette was isolated by gel electrophoresis as described in Kampmann, et al., and the isolated DNA was then used for PCR. Briefly, DNA was amplified using Phusion polymerase (NEB) for 23-28 cycles. The custom 5’ PCR primer used for each sample is listed in the sample information section. The custom 3’ PCR primer was caagcagaagacggcatacgacgactcggtgccactttttc, The thermocycling protocol was 98 °C, 30”, followed by 23-28 cycles of 98 °C, 15”, 56 °C, 15”, 72 °C, 15”. Indexed samples were pooled and sequenced on an Illumina HiSeq-2500 with the custom sequencing primer 5’-gtgtgttttgagactataagtatcccttggagaaccaccttgttg-3’. Sequencing libraries were pooled proportional to the number of sorted cells in each sample. The target sequencing depth was 2,000 reads/gRNA in the library for unsorted “background” samples, and 10 reads/cell in sorted samples.
PCR amplicon sequencing.

treatment_protocol: Jurkat cells stably expressing dCas9-VP64 were transduced with the lentiviral sgRNA library. Transduction was performed at a low multiplicity of infection to ensure that only one gRNA was present per cell. Cells were selected in media containing 1.5 μg/mL puromycin for days 2-5 post-infection to remove uninfected cells. After selection, cells were grown in fresh media without puromycin. The number of initially infected cells was at least 500x the number of sgRNAs in the library, and at least this many cells were maintained throughout the course of the experiment. 7-10 days post-infection cells were sorted based on IL2RA/CD25 or CD69 expression. Briefly, cells were resuspended in sterile sort buffer (PBS + 2% FBS) containing either CD25-PE or CD69-PE antibody at a 1:25 dilution. Cells were stained for 30 minutes on ice, washed twice with sort buffer, and passed through a 70 μm mesh. Cells were sorted into 4 bins based on IL2RA/CD25 or CD69 expression using a BD Influx cell sorter. The total number of cells collected was at least 500x the number of sgRNAs in the library. Additional unsorted cells totaling 500x the number of sgRNAs in the library were collected at this time. Duplicate infections and sorts were performed for each library. Collected cells were centrifuged at 500 xg for 5 minutes, and cell pellets were stored at -80 °C until genomic DNA was isolated.

tag: cell type

value: Jurkat E6-1

tag: sgrna library

value: IL2RA sgRNA library

tag: treatment

value: Sorted IL2RA-high

tag: custom 5' pcr primer

tag: illumina truseq index number

tag: illumina truseq index sequence

value: TAGCTT


title: CD69 high_rep2

value: CD69 sgRNA library

value: Sorted CD69-high


title: IL2RA background_rep1

description: il2ra.library.G.fa: Database of gRNA sequences for the IL2RA library containing the gRNA name and the gRNA sequence appended with a 5' G.

value: Unsorted

value: AGTTCC


title: CD69 low_rep2

value: Sorted CD69-low

value: GCCAAT


title: CD69 medium_rep2

value: Sorted CD69-medium


title: IL2RA background_rep2


title: CD69 baseline_rep2

value: Sorted CD69-baseline

value: CTTGTA


title: hCS890

city: Palo Alto

first: Wu

last: Wei

address: 3165 Porter Dr.

institute: Stanford Genome Technology Center

description: human interneurons

library_source: transcriptomic

data_processing:
Base-calling was performed using Illumina BaseSpace pipeline and standard parameters.
Sequence reads were aligned to human reference genome using TopHat v2.0.12 with default parameters
Nubmer of reads per gene were counted by HTSeq, version 0.6.0
Nubmer of reads per gene were normalized by DeSeq2 (1.10.1)
Genome_build: Homo_sapiens_Ensembl_GRCh37
Supplementary_files_format_and_content: tab-delimited text files include normalized expresson level of genes for all cells.

molecule: polyA RNA

source_name: hiPSC-derived 3D culture

growth_protocol: Differentiation of human iPSC into subpallium-like spheroids; 3D cultures were dissociated and FACS was used to isolate eGFP cells

extract_protocol: The single-cell RNA-Seq library preparation was adapted from the original SMART Seq-2 protocol. Single-cells were FACS sorted into a 96-well PCR plate followed by lysis, reverse transcription and PCR amplification according to SMART-seq2 protocol. After checking the quality control of cDNA library, final libraries were prepared using Nextera XT library prep kit (96 index primers, Illumina) protocol

treatment_protocol: Before and after migration in fused pallium-subpallium spheroids

value: hiPSC-derived neural 3D culture

tag: strain

tag: time

value: 4 weeks


title: CD69 background_rep1

description: cd69.library.G.fa: Database of gRNA sequences for the CD69 library containing the gRNA name and the gRNA sequence appended with a 5' G.


title: CD69 medium_rep1

description:
cd69.trim.medium.bedgraph: Bedgraph file containing log2-fold enrichment data for each gRNA, averaged across the two replicates
cd69.trim.window5.medium.bedgraph: Bedgraph file containing log2-fold enrichment data for a 5-gRNA sliding window, averaged across the two replicates


title: CD69 baseline_rep1

description:
cd69.trim.baseline.bedgraph: Bedgraph file containing log2-fold enrichment data for each gRNA, averaged across the two replicates
cd69.trim.window5.baseline.bedgraph: Bedgraph file containing log2-fold enrichment data for a 5-gRNA sliding window, averaged across the two replicates


title: hCS889

description: human interneurons


title: CD69 high_rep1

description:
cd69.trim.high.bedgraph: Bedgraph file containing log2-fold enrichment data for each gRNA, averaged across the two replicates
cd69.trim.window5.high.bedgraph: Bedgraph file containing log2-fold enrichment data for a 5-gRNA sliding window, averaged across the two replicates


title: CD69 low_rep1

description:
cd69.trim.low.bedgraph: Bedgraph file containing log2-fold enrichment data for each gRNA, averaged across the two replicates
cd69.trim.window5.low.bedgraph: Bedgraph file containing log2-fold enrichment data for a 5-gRNA sliding window, averaged across the two replicates


title: hCS875

description: human interneurons


title: IL2RA neg_rep1

description:
cd25.trim.neg.bedgraph: Bedgraph file containing log2-fold enrichment data for each gRNA, averaged across the two replicates
cd25.trim.window5.neg.bedgraph: Bedgraph file containing log2-fold enrichment data for a 5-gRNA sliding window, averaged across the two replicates

value: Sorted IL2RA-negative


title: hCS891

description: human interneurons


title: IL2RA mid_rep2

value: Sorted IL2RA-mid


title: hCS887

description: human interneurons


title: hCS876

description: human interneurons


title: hCS866

description: human interneurons


title: hCS872

description: human interneurons


title: hCS860

description: human interneurons


title: IL2RA neg_rep2

value: Jurkat E6-1


title: hCS863

description: human interneurons


title: hCS858

description: human interneurons


title: hCS874

description: human interneurons


title: IL2RA mid_rep1

description:
cd25.trim.mid.bedgraph: Bedgraph file containing log2-fold enrichment data for each gRNA, averaged across the two replicates
cd25.trim.window5.mid.bedgraph: Bedgraph file containing log2-fold enrichment data for a 5-gRNA sliding window, averaged across the two replicates

value: IL2RA sgRNA library


title: hCS856

description: human interneurons


title: hCS850

description: human interneurons


title: hCS853

description: human interneurons


title: hCS854

description: human interneurons


title: CD69 background_rep2


title: hCS848

description: human interneurons


title: hCS841

description: human interneurons


title: hCS840

description: human interneurons


title: hCS829

description: human interneurons


title: hCS836

description: human interneurons


title: hCS828

description: human interneurons


title: hCS824

description: human interneurons


title: hCS837

description: human interneurons


title: hCS821

description: human interneurons


title: hCS816

description: human interneurons


title: hCS809

description: human interneurons


title: hCS814

description: human interneurons


title: hCS820

description: human interneurons


title: hCS808

description: human interneurons


title: hCS807

description: human interneurons


title: hCS806

description: human interneurons


title: hCS802

description: human interneurons


title: hCS801

description: human interneurons


title: hCS788

description: human interneurons


title: hCS803

description: human interneurons


title: hCS790

description: human interneurons


title: hCS787

description: human interneurons


title: hCS792

description: human interneurons


title: hCS847

description: human interneurons


title: hCS780

description: human interneurons


title: hCS781

description: human interneurons


title: hCS783

description: human interneurons


title: hCS775

description: human interneurons


title: hCS774

description: human interneurons


title: hCS831

description: human interneurons


title: hCS823

description: human interneurons


title: hCS771

description: human interneurons


title: hCS769

description: human interneurons


title: hCS765

description: human interneurons


title: hCS764

description: human interneurons


title: hCS766

description: human interneurons


title: hCS767

description: human interneurons


title: hCS758

description: human interneurons


title: hCS768

description: human interneurons


title: hCS756

description: human interneurons


title: hCS755

description: human interneurons


title: hCS759

description: human interneurons


title: hCS760

description: human interneurons


title: hCS763

description: human interneurons


title: hCS762

description: human interneurons


title: hCS751

description: human interneurons


title: hCS750

description: human interneurons


title: hCS782

description: human interneurons


title: hCS749

description: human interneurons


title: hCS757

description: human interneurons


title: hCS746

description: human interneurons


title: hCS753

description: human interneurons


title: hCS744

description: human interneurons


title: hCS742

description: human interneurons


title: hCS752

description: human interneurons


title: hCS740

description: human interneurons


title: hCS745

description: human interneurons


title: hCS737

description: human interneurons


title: hCS736

description: human interneurons


title: hCS733

description: human interneurons


title: hCS732

description: human interneurons


title: hCS731

description: human interneurons


title: hCS729

description: human interneurons


title: hCS730

description: human interneurons


title: hCS726

description: human interneurons


title: hCS728

description: human interneurons


title: hCS725

description: human interneurons


title: hCS724

description: human interneurons


title: hCS716

description: human interneurons


title: hCS712

description: human interneurons


title: hCS717

description: human interneurons


title: hCS705

description: human interneurons


title: hCS702

description: human interneurons


title: hCS701

description: human interneurons


title: hCS713

description: human interneurons


title: hSS693

description: human interneurons


title: hSS692

description: human interneurons


title: hCS703

description: human interneurons


title: hSS690

description: human interneurons


title: hSS687

description: human interneurons


title: hSS682

description: human interneurons


title: hSS683

description: human interneurons


title: hSS678

description: human interneurons


title: hSS681

description: human interneurons


title: hSS680

description: human interneurons


title: hSS675

description: human interneurons


title: hSS677

description: human interneurons


title: hSS673

description: human interneurons


title: hSS672

description: human interneurons


title: hSS671

description: human interneurons


title: hSS665

description: human interneurons


title: hSS670

description: human interneurons


title: hSS666

description: human interneurons


title: hSS657

description: human interneurons


title: hSS655

description: human interneurons


title: hCS720

description: human interneurons


title: hSS649

description: human interneurons


title: hSS648

description: human interneurons


title: hSS643

description: human interneurons


title: hSS647

description: human interneurons


title: hSS650

description: human interneurons


title: hSS646

description: human interneurons


title: hSS658

description: human interneurons


title: hSS638

description: human interneurons


title: hSS633

description: human interneurons


title: hSS634

description: human interneurons


title: hSS645

description: human interneurons


title: hSS679

description: human interneurons


title: hSS630

description: human interneurons


title: hSS627

description: human interneurons


title: hSS623

description: human interneurons


title: hSS624

description: human interneurons


title: hSS621

description: human interneurons


title: hSS622

description: human interneurons


title: hSS625

description: human interneurons


title: hSS617

description: human interneurons


title: hSS664

description: human interneurons


title: hSS613

description: human interneurons


title: hSS608

description: human interneurons


title: hSS612

description: human interneurons


title: hSS610

description: human interneurons


title: hSS604

description: human interneurons


title: hSS601

description: human interneurons


title: hSS590

description: human interneurons


title: hSS584

description: human interneurons


title: hSS586

description: human interneurons


title: hSS576

description: human interneurons


title: hSS568

description: human interneurons


title: hSS566

description: human interneurons


title: hSS632

description: human interneurons


title: hSS573

description: human interneurons


title: hSS558

description: human interneurons


title: hSS575

description: human interneurons


title: hSS564

description: human interneurons


title: hSS561

description: human interneurons


title: hSS565

description: human interneurons


title: hSS556

description: human interneurons


title: hSS555

description: human interneurons


title: hSS551

description: human interneurons


title: hSS547

description: human interneurons


title: hSS544

description: human interneurons


title: hSS538

description: human interneurons


title: hSS552

description: human interneurons


title: hSS536

description: human interneurons


title: hSS534

description: human interneurons


title: hSS611

description: human interneurons


title: hSS532

description: human interneurons


title: hSS528

description: human interneurons


title: hSS530

description: human interneurons


title: hSS526

description: human interneurons


title: hSS524

description: human interneurons


title: hSS525

description: human interneurons


title: hSS523

description: human interneurons


title: hSS522

description: human interneurons


title: hSS518

description: human interneurons


title: hSS515

description: human interneurons


title: hSS516

description: human interneurons


title: hSS510

description: human interneurons


title: hSS514

description: human interneurons


title: hSS512

description: human interneurons


title: hSS507

description: human interneurons


title: hSS503

description: human interneurons


title: hSS504

description: human interneurons


title: hSS543

description: human interneurons


title: hSS501

description: human interneurons


title: hSS479

description: human interneurons

source_name: After migration in fused pallium-subpallium spheroids


title: hSS485

description: human interneurons


title: hSS490

description: human interneurons


title: hSS489

description: human interneurons


title: hSS477

description: human interneurons


title: hSS474

description: human interneurons


title: hSS475

description: human interneurons


title: hSS483

description: human interneurons


title: hSS473

description: human interneurons


title: hSS471

description: human interneurons


title: hSS472

description: human interneurons


title: hSS467

description: human interneurons


title: hSS466

description: human interneurons


title: hSS469

description: human interneurons


title: hSS495

description: human interneurons


title: hSS455

description: human interneurons


title: hSS460

description: human interneurons


title: hSS463

description: human interneurons


title: hSS453

description: human interneurons


title: hSS451

description: human interneurons


title: hSS452

description: human interneurons


title: hSS481

description: human interneurons


title: hSS444

description: human interneurons


title: hSS450

description: human interneurons


title: hSS470

description: human interneurons


title: hSS448

description: human interneurons


title: hSS478

description: human interneurons


title: hSS443

description: human interneurons


title: hSS454

description: human interneurons


title: hSS439

description: human interneurons


title: hSS449

description: human interneurons


title: hSS436

description: human interneurons


title: hSS438

description: human interneurons


title: hSS441

description: human interneurons


title: hSS442

description: human interneurons


title: hSS432

description: human interneurons


title: hSS430

description: human interneurons


title: hSS435

description: human interneurons


title: hSS429

description: human interneurons


title: hSS423

description: human interneurons


title: hSS425

description: human interneurons


title: hSS424

description: human interneurons


title: hSS421

description: human interneurons


title: hSS417

description: human interneurons


title: hSS420

description: human interneurons


title: hSS419

description: human interneurons


title: hSS422

description: human interneurons


title: hSS414

description: human interneurons


title: hSS413

description: human interneurons


title: hSS412

description: human interneurons


title: hSS406

description: human interneurons


title: hSS407

description: human interneurons


title: hSS431

description: human interneurons


title: hSS411

description: human interneurons


title: hSS427

description: human interneurons


title: hSS408

description: human interneurons


title: hSS410

description: human interneurons


title: hSS403

description: human interneurons


title: hCS390

description: human interneurons

source_name: Before migration in fused pallium-subpallium spheroids


title: hSS402

description: human interneurons


title: hCS392

description: human interneurons


title: hSS405

description: human interneurons


title: hCS388

description: human interneurons


title: hCS389

description: human interneurons


title: hCS391

description: human interneurons


title: hCS387

description: human interneurons


title: hSS426

description: human interneurons


title: hSS415

description: human interneurons


title: hCS383

description: human interneurons


title: hCS385

description: human interneurons


title: hCS386

description: human interneurons


title: hCS378

description: human interneurons


title: hCS380

description: human interneurons


title: hCS377

description: human interneurons


title: hSS404

description: human interneurons


title: hCS372

description: human interneurons


title: hCS371

description: human interneurons


title: hCS368

description: human interneurons


title: hCS363

description: human interneurons


title: hCS357

description: human interneurons


title: hCS351

description: human interneurons


title: hCS360

description: human interneurons


title: hCS348

description: human interneurons


title: hCS358

description: human interneurons


title: hCS349

description: human interneurons


title: hCS345

description: human interneurons


title: hCS344

description: human interneurons


title: hCS343

description: human interneurons


title: hCS341

description: human interneurons


title: hCS339

description: human interneurons


title: hCS340

description: human interneurons


title: hCS382

description: human interneurons


title: hCS375

description: human interneurons


title: hCS331

description: human interneurons


title: hCS332

description: human interneurons


title: hCS334

description: human interneurons


title: hCS335

description: human interneurons


title: hCS333

description: human interneurons


title: hCS362

description: human interneurons


title: hCS327

description: human interneurons


title: hCS326

description: human interneurons


title: hCS359

description: human interneurons


title: hCS324

description: human interneurons


title: hCS321

description: human interneurons


title: hCS325

description: human interneurons


title: hCS318

description: human interneurons


title: hCS306

description: human interneurons


title: hCS311

description: human interneurons


title: hCS305

description: human interneurons


title: hCS310

description: human interneurons


title: hCS309

description: human interneurons


title: hCS301

description: human interneurons


title: hCS336

description: human interneurons


title: hCS307

description: human interneurons


title: hCS302

description: human interneurons


title: hCS308

description: human interneurons


title: hCS319

description: human interneurons


title: hSS77

description: human interneurons


title: hSS76

description: human interneurons


title: hCS330

description: human interneurons


title: hCS303

description: human interneurons


title: hSS75

description: human interneurons


title: hSS74

description: human interneurons


title: hCS320

description: human interneurons


title: hSS73

description: human interneurons


title: hSS72

description: human interneurons


title: hSS65

description: human interneurons


title: hSS70

description: human interneurons


title: hSS62

description: human interneurons


title: hSS68

description: human interneurons


title: hSS64

description: human interneurons


title: hSS60

description: human interneurons


title: hCS315

description: human interneurons


title: hSS63

description: human interneurons


title: hSS58

description: human interneurons


title: hSS57

description: human interneurons


title: hSS55

description: human interneurons


title: hSS51

description: human interneurons


title: hSS48

description: human interneurons


title: hSS52

description: human interneurons


title: hSS49

description: human interneurons


title: hSS54

description: human interneurons


title: hSS46

description: human interneurons


title: hSS45

description: human interneurons


title: hSS71

description: human interneurons


title: hSS44

description: human interneurons


title: hSS43

description: human interneurons


title: hSS67

description: human interneurons


title: hSS42

description: human interneurons


title: hSS39

description: human interneurons


title: hSS40

description: human interneurons


title: hSS38

description: human interneurons


title: hSS36

description: human interneurons


title: hSS33

description: human interneurons


title: hSS34

description: human interneurons


title: hSS27

description: human interneurons


title: hSS37

description: human interneurons


title: hSS29

description: human interneurons


title: hSS59

description: human interneurons


title: hSS26

description: human interneurons


title: hSS22

description: human interneurons


title: hSS21

description: human interneurons


title: hSS24

description: human interneurons


title: hSS50

description: human interneurons


title: hSS20

description: human interneurons


title: hSS19

description: human interneurons


title: hSS30

description: human interneurons


title: hSS13

description: human interneurons


title: hSS15

description: human interneurons


title: hSS11

description: human interneurons


title: hSS10

description: human interneurons


title: hSS08

description: human interneurons


title: hSS05

description: human interneurons


title: hSS09

description: human interneurons


title: hSS06

description: human interneurons


title: hSS04

description: human interneurons


title: hCS84

description: human interneurons


title: hCS82

description: human interneurons


title: hSS03

description: human interneurons


title: hCS83

description: human interneurons


title: hCS75

description: human interneurons


title: hCS72

description: human interneurons


title: hCS68

description: human interneurons


title: hCS74

description: human interneurons


title: hCS71

description: human interneurons


title: hCS67

description: human interneurons


title: hCS64

description: human interneurons


title: hCS59

description: human interneurons


title: hCS63

description: human interneurons


title: hSS16

description: human interneurons


title: hSS17

description: human interneurons


title: hSS18

description: human interneurons


title: hCS57

description: human interneurons


title: hCS62

description: human interneurons


title: hCS50

description: human interneurons


title: hCS54

description: human interneurons


title: hCS53

description: human interneurons


title: hCS49

description: human interneurons


title: hCS42

description: human interneurons


title: hCS45

description: human interneurons


title: hCS36

description: human interneurons


title: hCS39

description: human interneurons


title: hCS41

description: human interneurons


title: hCS38

description: human interneurons


title: hCS31

description: human interneurons


title: hCS33

description: human interneurons


title: hCS35

description: human interneurons


title: hCS19

description: human interneurons


title: hCS23

description: human interneurons


title: hCS24

description: human interneurons


title: hCS26

description: human interneurons


title: hCS18

description: human interneurons


title: hCS17

description: human interneurons


title: hCS16

description: human interneurons


title: hCS08

description: human interneurons


title: hCS09

description: human interneurons


title: hCS51

description: human interneurons


title: hCS14

description: human interneurons


title: hCS10

description: human interneurons


title: hCS11

description: human interneurons


title: hCS12

description: human interneurons


title: hCS13

description: human interneurons


title: hCS07

description: human interneurons


title: hCS06

description: human interneurons


title: hCS03

description: human interneurons


title: hSS23

description: human interneurons


title: hCS02

description: human interneurons


title: hCS04

description: human interneurons


title: hCS01

description: human interneurons


title: Fe-deficient 3

city: Philadelphia

first: Rebecca

middle: A

last: Simmons

state: PA

address: 3400 Civic Center Blvd

department: Genetics

institute: University of Pennsylvania

description:
Company: Charles River
Whole-genome Bisulfite Seq

data_processing:
FASTQ files generated using bcl2fastq.
Adapter sequence trimmed using in-house software.
Trimmed single or paired reads aligned to rat genome using bs_seeker version 1.
Modified C bases tallied from bs_seeker output using in-house software.
Genome_build: rn4 (Baylor 3.4)
Supplementary_files_format_and_content: *.tab: Tab-delimited text files include location of C, number of reads covering, and number of modified reads covering each C in CG context. chr-nucleotide-nucleotide-methylation % - coverage (reads).

organism: Rattus norvegicus

source_name: hippocampus, iron-deficient diet

growth_protocol: Pregnancy

extract_protocol:
Hippocampi were dissected and stored as described in doi:10.3945/jn.115.227561
Bisulfite conversion with Imprint DNA modification kit, ligated with Illumina methylated paired end adapters.

treatment_protocol: Iron-deficient rats were fed 4ppm iron, and control/iron-sufficient rats were fed 200ppm iron.

tag: strain/background

value: Sprague-Dawley

tag: tissue

value: hippocampus

value: Iron-deficient diet


title: Fe-deficient 1


title: Fe-sufficient 3

source_name: hippocampus, iron-sufficient diet

value: Iron-sufficient diet


title: Fe-sufficient 2


title: H3K27ac-rSC-2

city: Cincinnati

first: Richard

last: Lu

state: OH

address: 3333 Burnet Ave

department: CBDI

institute: Cincinnati Children's Hospital Medical Center

data_processing:
Illumina Casava software used for basecalling.
Sequenced reads were trimmed for adaptor sequence.
RNA-seq: Reads mapped to mm10 whole genome by Tophat with default options
ChIP-seq: Reads mapped to rn5 whole genome by Bowtie2 with default options
RNA-seq: Fragment Per Kilobase of exon per Megabase of library size (FPKM) were calculated using Cufflinks with default options. Outputs from Cufflinks are provided as supplementary files as a tab-delimited text file.
ChIP-seq: Peak calling was performed using MACS1.4 with a default p value cut off.
Genome_build: mm10
Genome_build: rn5
Supplementary_files_format_and_content: format:fpkm_traking ; Contain the estimated expression values produced by Cufflinks.
Supplementary_files_format_and_content: wig files were generated by MACS14

source_name: rat Schwann cell

growth_protocol: Rat Schwann cells from sciatic nerves of newborn rats (1–2 days-old) were isolated. Schwann cells were grown routinely in DMEM/10% FBS (Life Technologies), supplemented with 10 ng/ml Neuregulin 1 (Nrg1; R&D Systems), and 5 μM forskolin (Sigma), plus L-glutamine and penicillin/streptomycin, hereafter denoted as SC proliferation medium. Cells between passages 2 and 6 were used in all experiments. >95% SC purity was achieved, assessed by positive SOX10 and S100β immunoreactivity.

extract_protocol:
RNA from control and Gnas or TazYap mutant cerebella were extracted using TRIZOL (Life Technologies) followed by purification using an RNeasy Mini Kit (Qiagen).
Purified rat SCs (∼20 million cells) were fixed for 10 min at room temperature with 1% formaldehyde-containing medium. Nuclei were isolated and sonicated in sonication buffer (10 mM Tris-HCl [pH 8.0], 1 mM EDTA, 0.5 mM EGTA, and protease inhibitor cocktail). Sonicated chromatin (∼300 μg) was used for immunoprecipitation by incubation with rabbit IgG oranti-Taz, anti-H3K27ac antibody (4 μg) overnight at 4°C.
RNA-seq libraries were prepared using Illumina RNA-Seq Preparation Kit.
The ChIP-seq libraries were prepared using NEBNext ChIP-seq Library Prep Master Mix Set for Illumina (NEB catalogue number E6240L) and then run on the Illumina sequencer HiSeq 2000.

value: primary cultured

tag: antibody


title: H3K27ac-rSC-1


title: Taz-rSC-2

value: oranti-Taz


title: Stimulated rep2

data_processing:
Basecalling was performed using the innate HiSeq Control Software as part of the primary run processing.
Demultiplexing was performed with Bcl2Fastq version 2.17
HISAT2 was used to align reads to the IL2RA region (hg38 chr10:6009378-6073794) using arguments --no-mixed --no-discordant --rdg 6,5 --rfg 6,5 --score-min L,-.6,-.4
kallisto was used to generate an index from the Ensembl release 79 transcriptome (Homo_sapiens.GRCh38.rel79.cdna.all.fa) using argument -k 31. Abundances were then quantified relative to that index using arguments -b 30 -l 350 -s 100
Genome_build: GRCh38
Supplementary_files_format_and_content: kallisto abundance.tsv files give transcript-level abundances in TPM

molecule: total RNA

source_name: HuT78 cells

growth_protocol: HuT78 cells were cultured in RPMI 1640 media supplemented with 10% fetal calf serum, 1 mM Sodium Pyruvate (Gibco), 100 U/mL penicillin (Gibco) and 100 μg/mL streptomycin (Gibco). Cell line identity was authenticated by short tandem repeat (STR) analysis and verified mycoplasma free using the MycoAlert Mycoplasma Detection Kit (Lonza). HuT78 cells were passaged every 2-3 days and maintained at 0.1-1.0x10^6 cells/mL of media. HuT78 cells were transduced with a lentiviral dCas9-VP64-2A-GFP expression vector (Addgene 61422). Single GFP+ cells were sorted by FACS into the wells of a 96-well plate, and a clone with bright uniform GFP expression were selected for use in these experiments.

extract_protocol:
Cells were harvested and total RNA was isolated from samples using the RNeasy Mini Kit (QIAGEN, Cat. #74104) according to the manufacturer’s instructions with the following options: Cells were pelleted and re-suspended in RLT buffer with β-mercaptoethanol and homogenized using QIAshredder (QIAGEN, Cat. #79654). On-column DNase digestion was performed with RNase-Free DNase Set (QIAGEN, Cat. #79254). RNA samples were analyzed with a NanoDrop spectrophotometer and all samples had a 260/280 and 260/230 ratio of 1.80 or higher. RNA integrity was measured with the AATI Fragment Analyzer (Advanced Analytical Technologies, Ankeny, Iowa), and all samples had an RNA Quality score (RQN) of 10.0. RNA concentration using the Qubit RNA BR Assay (ThermoFisher Scientific, Cat. #Q10210).
RNA-Seq libraries from biological duplicate samples were prepared TruSeq RNA Library Preparation Kit v2, Set A (Illumina, Cat. # RS-122-2001) and Set B (Cat. # RS-122-2002), following the Illumina TruSeq sample Preparation v2 Guide: Low Sample (LS) Protocol. The samples were pooled and sequenced with the Illumina HiSeq4000.

treatment_protocol:
For “PBS” and “Stimulated” samples, HuT78 cells were used that stably expressed dCas9-VP64 but no sgRNA. The wells of a flat-bottomed 96-well plate were coated overnight with anti-CD3 (clone UCHT1, 10 μg/ml, TONBO Biosciences) and anti-CD28 (clone CD28.2, 10 μg/ml, TONBO Biosciences) in PBS, or PBS only. Coated wells were washed twice with PBS. Cells were plated at 0.5x10^6 cells/mL in 200 μL of growth media per well in a 96-well plate. Cells were cultured for 48 hours and then harvested for RNA isolation.
Treatment protocols were specific to subsets of samples and are listed in the SAMPLES section.

tag: sgrna sequence


title: IgG

value: IgG


title: Taz-rSC-1


title: Stimulated rep1


title: PBS rep1

value: PBS


title: IL2RA CaRE4 sgRNA 2 rep1

treatment_protocol:
HuT78 cells stably expressing dCas9-VP64 were transduced with the lentiviral sgRNA expression vector “pCRISPRia-v2”, described in Horlbeck, et al, Elife 5, 2767 (2016). Cells were selected in media containing 1.5 μg/mL puromycin for days 2-5 post-infection to remove uninfected cells. After selection, cells were plated at 0.5x10^6 cells/mL in 1 mL of growth media per well in a 24-well plate. Cells were cultured for 48 hours and then harvested for RNA isolation.
Treatment protocols were specific to subsets of samples and are listed in the SAMPLES section.

value: GTGGGTGATTCTGTGGGCAG

value: Untreated


title: IL2RA CaRE4 sgRNA 1 rep2

value: TCTGAAGGAGGTATCTATTT


title: PBS rep2


title: IL2RA CaRE3 sgRNA 2 rep2

value: CCACGCTCCCAGAAAGCAAA


title: IL2RA CaRE3 sgRNA 2 rep1


title: IL2RA CaRE4 sgRNA 1 rep1


title: IL2RA CaRE3 sgRNA 1 rep1

value: ACTGTGAGCGTCCTCAGTGC


title: IL2RA TSS sgRNA 2 rep2

value: ATAAGCTGAGTCCTCCCCTC


title: IL2RA TSS sgRNA 1 rep2

value: TTATGGGCGTAGCTGAAGAA


title: IL2RA TSS sgRNA 1 rep1


title: IL2RA TSS sgRNA 2 rep1


title: Non-targeting sgRNA 2 rep1

value: GGCTCGGTCCCGCGTCGTCG


title: Non-targeting sgRNA 2 rep2


title: Fe-deficient 2


title: DegradomeSeq_w1118

city: Aarhus

first: Peter

last: Andersen

state: Denmark

address: Universitetsbyen 83

department: MBG

country: Denmark

institute: Aarhus University

description:
Degradome-seq (specific to 5'-monophosphorylated RNAs) from w1118 (wildtype) ovaries
DegradomeSeq_w1118

data_processing:
Library strategy: Degradome-Seq
basecalled with the instruments RTA (Run time analysis version 1.12.48.0): SCS (PN:RTA DS:Controlling software on instrument VN:1.13.48.0)
for small RNA sequencing fastq files were adapter 'AGATCGGAAGAGCACACGTCT' clipped using cutadapt, version 1.0, default settings and the first and last 4 bases were trimmed (both linkers contain 4 random nucleotides at their respective termini)
reads were mapped with bowtie v0.12.7 or STAR (RNAseq data only) to D. mel. genome (dm6, r6.10)
BigWig files were generated with Homertools v4.6
Genome_build: Drosophila melanogaster dm6, release 6.10
Supplementary_files_format_and_content: The processed data files are normalized bigwig tracks with uniquely mapping reads only

organism: Drosophila melanogaster

source_name: Adult ovaries

growth_protocol: Flies were kept at strandard conditions at 25º C.

extract_protocol:
Total RNA extraction with TRIzol followed by rRNA depletion using RiboZero
Protocol modified from PMID: 23260138
Ovaries were hand-dissected into ice-cold PBS

tag: genotype

tag: developmental stage

value: Adult

value: ovary


title: CapSeq_w1118

description:
Capseq (specific to 5' capped RNAs) from w1118 (wildtype) ovaries
CapSeq_w1118

data_processing:
Library strategy: Cap-Seq
basecalled with the instruments RTA (Run time analysis version 1.12.48.0): SCS (PN:RTA DS:Controlling software on instrument VN:1.13.48.0)
for small RNA sequencing fastq files were adapter 'AGATCGGAAGAGCACACGTCT' clipped using cutadapt, version 1.0, default settings and the first and last 4 bases were trimmed (both linkers contain 4 random nucleotides at their respective termini)
reads were mapped with bowtie v0.12.7 or STAR (RNAseq data only) to D. mel. genome (dm6, r6.10)
BigWig files were generated with Homertools v4.6
Genome_build: Drosophila melanogaster dm6, release 6.10
Supplementary_files_format_and_content: The processed data files are normalized bigwig tracks with uniquely mapping reads only

extract_protocol:
Total RNA extraction with TRIzol
Protocol modified from PMID: 23260138
Ovaries were hand-dissected into ice-cold PBS


title: Fe-sufficient 1

value: Sprague-Dawley

value: hippocampus


title: ChIPseq_input_moonshiner_KO

description:
Input sample of RNA polymerase II ChIPseq from moonshiner knock-out ovaries
ChIPseq_input_moonshiner_KO

data_processing:
basecalled with the instruments RTA (Run time analysis version 1.12.48.0): SCS (PN:RTA DS:Controlling software on instrument VN:1.13.48.0)
for small RNA sequencing fastq files were adapter 'AGATCGGAAGAGCACACGTCT' clipped using cutadapt, version 1.0, default settings and the first and last 4 bases were trimmed (both linkers contain 4 random nucleotides at their respective termini)
reads were mapped with bowtie v0.12.7 or STAR (RNAseq data only) to D. mel. genome (dm6, r6.10)
BigWig files were generated with Homertools v4.6
Genome_build: Drosophila melanogaster dm6, release 6.10
Supplementary_files_format_and_content: The processed data files are normalized bigwig tracks with uniquely mapping reads only

extract_protocol:
Lysis and sonication of formaldehyde-crosslinked ovaries
NEBNext® Ultra™ DNA Library Prep Kit for Illumina®
Ovaries were hand-dissected into ice-cold PBS

tag: chip antibody


title: ChIPseq_input_rhino_KO

description:
Input sample of RNA polymerase II ChIPseq from rhino knock-out ovaries
ChIPseq_input_rhino_KO


title: ChIPseq_input_w1118_1

description:
Input sample of RNA polymerase II ChIPseq from w1118 (wildtype) ovaries
ChIPseq_input_w1118_1


title: ChIPseq_antiRNAPol2_moonshiner_KO

description:
RNA polymerase II ChIPseq from moonshiner knock-out ovaries
ChIPseq_antiRNAPol2_moonshiner_KO

value: anti-RNA Pol II (8WG16, Abcam, ab817)


title: ChIPseq_antiRNAPol2_rhino_KO

description:
RNA polymerase II ChIPseq from rhino knock-out ovaries
ChIPseq_antiRNAPol2_rhino_KO


title: ChIPseq_antiRNAPol2_w1118_1

description:
RNA polymerase II ChIPseq from w1118 (wildtype) ovaries
ChIPseq_antiRNAPol2_w1118_1


title: IL2RA CaRE4 sgRNA 2 rep2


title: ChIPseq_antiRhino_Pld_promoter_deletion

description:
Rhino ChIPseq from Pld promoter deletion ovaries
ChIPseq_antiRhino_Pld_promoter_deletion

value: w ; Pld{delta}prom[m1-1] ;

value: anti-Rhino rabbit polyclonal antibody (PMID:24906153)


title: ChIPseq_antiRhino_Rhino_moonshiner_KO

description:
Rhino ChIPseq from moonshiner knock-out ovaries
ChIPseq_antiRhino_Rhino_moonshiner_KO


title: IL2RA CaRE3 sgRNA 1 rep2


title: ChIPseq_antiRhino_Rhino_rhino_KO

description:
Rhino ChIPseq from rhino knock-out ovaries
ChIPseq_antiRhino_Rhino_rhino_KO


title: ChIPseq_antiRhino_w1118_1

description:
Rhino ChIPseq from w1118 (wildtype) ovaries
ChIPseq_antiRhino_w1118_1


title: RNAseq_TRF2_KD

description:
RNAseq (RiboZero-treated) from TRF2 germline knock-down ovaries
RNAseq_TRF2_KD

extract_protocol:
Total RNA extraction with TRIzol followed by rRNA depletion using RiboZero
NEBNext® Ultra™ Directional RNA Library Prep Kit for Illumina®
Ovaries were hand-dissected into ice-cold PBS

value: w, MTD/+; MTD/+; MTD/P{TRiP.GL01076}[attP2]/;


title: RNAseq_TFIIA-S_KD

description:
RNAseq (RiboZero-treated) from TFIIA-S germline knock-down ovaries
RNAseq_TFIIA-S_KD

value: w, MTD/+; MTD/+; MTD/pW22>TFIIA-S_sh2[attP2]/;


title: RNAseq_moonshiner_KD

description:
RNAseq (RiboZero-treated) from moonshiner germline knock-down ovaries
RNAseq_moonshiner_KD


title: smallRNAseq_moonshiner_KO_bypass

description:
small RNA sequencing from moonshiner knock-out ovaries expressing a GFP-TRF2 transgene and a Deadlock-antiGFP-nanobody transgene
small RNAs
smallRNAseq_moonshiner_KO_bypass

extract_protocol:
Total RNA extraction with TRIzol
small RNA sequencing as described in PMID: 21890921
Ovaries were hand-dissected into ice-cold PBS

value: w, moon_FS2{delta}28; pNASA>LAP-TRF2s[attP40]/+; pNASA>Deadlock-vhhGFP[attP2]/+


title: RNAseq_control_KD

description:
RNAseq (RiboZero-treated) from control germline knock-down (w sh) ovaries
RNAseq_control_KD

value: w, MTD/+; MTD/+; MTD/pW20>w_sh[attP2]/;


title: RNAseq_moonshiner_KO

description:
RNAseq (RiboZero-treated) from moonshiner knock-out ovaries
sequenced twice for increased coverage (L1: lane1, L2: lane2)
RNAseq_moonshiner_KO


title: RNAseq_rhino_KO

description:
RNAseq (RiboZero-treated) from rhino knock-out ovaries
sequenced twice for increased coverage (L1: lane1, L2: lane2)
RNAseq_rhino_KO


title: RNAseq_w1118

description:
RNAseq (RiboZero-treated) from w1118 (wildtype) ovaries
sequenced twice for increased coverage (L1: lane1, L2: lane2)
RNAseq_w1118


description:
small RNA sequencing from moonshiner knock-out ovaries expressing a Deadlock-antiGFP-nanobody transgene
small RNAs
smallRNAseq_moonshiner_KO_Deadlock-antiGFPnanobody

extract_protocol:
Total RNA extraction with TRIzol
small RNA sequencing as described in PMID: 21890920
Ovaries were hand-dissected into ice-cold PBS

value: w, moon_FS2{delta}28; CyO/+; pNASA>Deadlock-vhhGFP[attP2]/+


title: smallRNAseq_moonshiner_KO_2

description:
small RNA sequencing from moonshiner knock-out ovaries_2
small RNAs
smallRNAseq_moonshiner_KO_2

extract_protocol:
Total RNA extraction with TRIzol
small RNA sequencing as described in PMID: 21890918
Ovaries were hand-dissected into ice-cold PBS

value: w, moon_FS2{delta}28; ;


title: smallRNAseq_moonshiner_KO_GFP-TRF2

description:
small RNA sequencing from moonshiner knock-out ovaries expressing a GFP-TRF2 transgene
small RNAs
smallRNAseq_moonshiner_KO_GFP-TRF2

extract_protocol:
Total RNA extraction with TRIzol
small RNA sequencing as described in PMID: 21890919
Ovaries were hand-dissected into ice-cold PBS

value: w, moon_FS2{delta}28; pNASA>LAP-TRF2s[attP40]/+; Sb/+


description:
small RNA sequencing from ovaries of flies with deletion of the right promoter of the cluster42AB piRNA source locus
small RNAs
smallRNAseq_cluster42AB_rightpromoter_deletion

extract_protocol:
Total RNA extraction with TRIzol
small RNA sequencing as described in PMID: 21890917
Ovaries were hand-dissected into ice-cold PBS

value: w; cluster42AB{delta}RIGHTprom[3];


description:
small RNA sequencing from ovaries of flies with deletion of both promoters of the cluster38C1 piRNA source locus - combined with moonshiner knock-out
small RNAs
smallRNAseq_cluster38C1_bothpromoter_deletion_moon_KO

extract_protocol:
Total RNA extraction with TRIzol
small RNA sequencing as described in PMID: 21890916
Ovaries were hand-dissected into ice-cold PBS


description:
small RNA sequencing from ovaries of flies with deletion of both promoters of the cluster38C1 piRNA source locus
small RNAs
smallRNAseq_cluster38C1_bothpromoter_deletion

extract_protocol:
Total RNA extraction with TRIzol
small RNA sequencing as described in PMID: 21890915
Ovaries were hand-dissected into ice-cold PBS


description:
small RNA sequencing from ovaries of flies with deletion of the right promoter of the cluster38C1 piRNA source locus - combined with moonshiner knock-out
small RNAs
smallRNAseq_cluster38C1_rightpromoter_deletion_moon_KO

extract_protocol:
Total RNA extraction with TRIzol
small RNA sequencing as described in PMID: 21890914
Ovaries were hand-dissected into ice-cold PBS


description:
small RNA sequencing from ovaries of flies with deletion of the right promoter of the cluster38C1 piRNA source locus
small RNAs
smallRNAseq_cluster38C1_rightpromoter_deletion

extract_protocol:
Total RNA extraction with TRIzol
small RNA sequencing as described in PMID: 21890913
Ovaries were hand-dissected into ice-cold PBS

value: w; cluster38C1{delta}RIGHTprom[1-1];


description:
small RNA sequencing from ovaries of flies with deletion of the left promoter of the cluster38C1 piRNA source locus - combined with moonshiner knock-out
small RNAs
smallRNAseq_cluster38C1_leftpromoter_deletion_moon_KO

extract_protocol:
Total RNA extraction with TRIzol
small RNA sequencing as described in PMID: 21890912
Ovaries were hand-dissected into ice-cold PBS


description:
small RNA sequencing from ovaries of flies with deletion of the left promoter of the cluster38C1 piRNA source locus
small RNAs
smallRNAseq_cluster38C1_leftpromoter_deletion

extract_protocol:
Total RNA extraction with TRIzol
small RNA sequencing as described in PMID: 21890911
Ovaries were hand-dissected into ice-cold PBS

value: w; cluster38C1{delta}LEFTprom[1-6];


title: smallRNAseq_w1118_2

description:
small RNA sequencing from w1118 (wildtype) ovaries_2
small RNAs
smallRNAseq_w1118_2

extract_protocol:
Total RNA extraction with TRIzol
small RNA sequencing as described in PMID: 21890907
Ovaries were hand-dissected into ice-cold PBS

value: Adult


title: smallRNAseq_TFIIA-S_KD

description:
small RNA sequencing from TFIIA-S germline knock-down ovaries
small RNAs
smallRNAseq_TFIIA-S_KD

extract_protocol:
Total RNA extraction with TRIzol
small RNA sequencing as described in PMID: 21890905
Ovaries were hand-dissected into ice-cold PBS

value: ovary


title: smallRNAseq_TRF2_KD

description:
small RNA sequencing from TRF2 germline knock-down ovaries
small RNAs
smallRNAseq_TRF2_KD

extract_protocol:
Total RNA extraction with TRIzol
small RNA sequencing as described in PMID: 21890906
Ovaries were hand-dissected into ice-cold PBS


title: smallRNAseq_Pld_promoter_deletion

description:
small RNA sequencing from Pld promoter deletion ovaries
small RNAs
smallRNAseq_Pld_promoter_deletion

extract_protocol:
Total RNA extraction with TRIzol
small RNA sequencing as described in PMID: 21890908
Ovaries were hand-dissected into ice-cold PBS


title: smallRNAseq_moonshiner_KD

description:
small RNA sequencing from moonshiner germline knock-down ovaries
small RNAs
smallRNAseq_moonshiner_KD

extract_protocol:
Total RNA extraction with TRIzol
small RNA sequencing as described in PMID: 21890904
Ovaries were hand-dissected into ice-cold PBS


title: PCHiC_D2_Exp2

city: Odense M

first: Susanne

last: Mandrup

address: Campusvej 55

department: Biochemistry and Molecular Biology

institute: University of Southern Denmark

description: Genomic DNA

data_processing:
mRNA-seq reads were aligned to mm9 by STAR (Dobin et al., 2013) and differentially expressed genes were identified by DESeq2 (Love et al., 2014)
ChIP-seq reads were aligned to mm9 by STAR, peaks were identified by HOMER, and significantly changing binding sites were identified by DESeq2.
Hi-C and PCHi-C raw sequencing reads were aligned and filtered by HiCUP (Wingett et al., 2015). Hi-C data was further processed by HOMER (Heinz et al., 2010). PCHi-C data was analysed by CHiCAGO (Cairns et al., 2016) to identify chromatin interactions.
Genome_build: mm9
Supplementary_files_format_and_content: txt, bedGraph

organism: Mus musculus

source_name: 3T3-L1 preadipocytes

growth_protocol: Mouse 3T3-L1 cells (obtained from ATCC) were grown in Dulbecco’s Modified Eagle’s Medium (DMEM, #52-100-021, Gibco) supplemented with 10% calf serum (#B15-005, PAA) and 1% pen/strep (#DE 17-602E, Lonza).

extract_protocol:
RNA: Cells were harvested in Isol-RNA lysis reagent (5 PRIME) and purified according to the manufacturer's instructions. ChIP-seq: Crosslinking: For MED1/TRAP220, SMC1, P300, HDAC2, HDAC3 and NCoR ChIP-seq: Cells were crosslinked by 2 mM disuccinimidyl glutarate (DSG) for 20-30 min at RT followed by 1% formaldehyde for 10 min at RT. Crosslinking was quenched by addition of 0.125M glycine for 10 min. For CTCF and histone mark ChIP-seq: Cells were crosslinked in formaldehyde only as described above. Sonication: For MED1/TRAP220, SMC1, CTCF and histone marks: Crosslinked cells were scraped in ChIP lysis buffer (0.1% SDS, 1% Triton X-100, 0.15M NaCl, 1mM EDTA, 20mM Tris, pH=8) and sonicated for 40 cycles (approximately 10-12 million crosslinked cells in 1.5 ml lysis buffer was sonicated in 15 ml tubes, 30 sec on, 30 sec off, maximum intensity) in a Bioruptor (Diagenode). For P300, HDAC2, HDAC3 and NCoR: Crosslinked cells were scraped in high SDS lysis buffer (1% SDS,  20mM EDTA, 50mM Tris, pH=8) and rotated 2h at 4C to release nuclei. A buffer change was done on pelleted nuclei to the ChIP lysis buffer described above before sonication for 7 cycles (nuclei from approximately 10-12 million crosslinked cells in 0.3 ml lysis buffer was sonicated in 1.5 ml tubes, 30 sec on, 30 sec off, maximum intensity) in a Bioruptor (Diagenode). Immunoprecipitation was performed overnight using specific antibodies. After washing, IP’ed chromatin was decrosslinked overnight at 65 °C and DNA was purified by phenol-chloroform extraction. Crosslinking of cells for Hi-C and PCHi-C: Approximately 50-80 millions of 3T3-L1 cells were crosslinked for 10 min by 2% formaldehyde in PBS. Crosslinking was quenched by addition of 0.125M glycine. Cells were incubated at room temperature for 5 min and then on ice for 15 min. Crosslinked cells were washed once in cold PBS, and the cell pellets were flash frozen in liquid nitrogen and stored at -80°C.
mRNA-seq: Libraries were prepared from purified RNA (TruSeq Illumina mRNA prep kit) and sequenced according to the manufacturer’s instructions (HiSeq v4 Illumina SR sequencing kit). ChIP-seq: Libraries were prepared from the purified DNA using the NEB ChIPseq prep kit and samples were sequenced according to the manufacturer’s instructions (HiSeq v4 Illumina SR sequencing kit). Hi-C : Hi-C library generation was carried out with in-nucleus ligation as described previously (Nagano et al., 2015). PCHi-C: Capture Hi-C of promoters was carried out with SureSelect target enrichment (SureSelectXT Custom 3-5.9Mb library), using the custom-designed biotinylated RNA bait library and custom paired-end blockers according to the manufacturer’s instructions (Agilent Technologies). Between 500 ng to 1 µg of Hi-C library were captured. After library enrichment, a post-capture PCR amplification step was carried out using PE PCR 1.0 and PE PCR 2.0 primers with 4 PCR amplification cycles.
RNA-seq, ChIP-seq, Hi-C, PCHi-C

treatment_protocol: Cells were induced to differentiate two days post confluency by addition of differentiation media (DMEM supplemented with 10% fetal bovine serum (#S 01 15, Biochrom) and 1% pen/strep) supplemented with 1 μg/ml insulin (Sigma-Aldrich, I9278), 390 ng/ml dexamethasone (Sigma-Aldrich, D4902) and 1115 μg/ml 3-Isobutyl-1-methylxanthine (Sigma-Aldrich, I5879). Two days after induction of differentiation, fresh differentiation media supplemented with 1 μg/ml insulin were added. From day four and onwards, differentiated cells were maintained in differentiation media.

value: adipocyte


title: smallRNAseq_rhino_KD

description:
small RNA sequencing from rhino germline knock-down ovaries
small RNAs
smallRNAseq_rhino_KD

extract_protocol:
Total RNA extraction with TRIzol
small RNA sequencing as described in PMID: 21890903
Ovaries were hand-dissected into ice-cold PBS

value: w, MTD/+; MTD/+; MTD/pW20>rhi_sh[attP2]/;


title: RNAseq_rhino_KD

description:
RNAseq (RiboZero-treated) from rhino germline knock-down ovaries
RNAseq_rhino_KD

extract_protocol:
Total RNA extraction with TRIzol followed by rRNA depletion using RiboZero
NEBNext® Ultra™ Directional RNA Library Prep Kit for Illumina®
Ovaries were hand-dissected into ice-cold PBS


title: PCHiC_D2_Exp1

description: Genomic DNA


description:
small RNA sequencing from ovaries of flies with Pld promoter deletion and expression of Pld cDNA in trans
small RNAs
smallRNAseq_Pld_promoter_deletion_with_Pld_transgene

extract_protocol:
Total RNA extraction with TRIzol
small RNA sequencing as described in PMID: 21890909
Ovaries were hand-dissected into ice-cold PBS

value: w ; Pld{delta}prom[m1-1], tub40>Pld_cDNA[attP40] ;


title: smallRNAseq_control_KD

description:
small RNA sequencing from control germline knock-down (w sh) ovaries
small RNAs
smallRNAseq_control_KD

extract_protocol:
Total RNA extraction with TRIzol
small RNA sequencing as described in PMID: 21890902
Ovaries were hand-dissected into ice-cold PBS


title: smallRNAseq_rhino_KO

description:
small RNA sequencing from rhino knock-out ovaries
small RNAs
smallRNAseq_rhino_KO

extract_protocol:
Total RNA extraction with TRIzol
small RNA sequencing as described in PMID: 21890900
Ovaries were hand-dissected into ice-cold PBS


title: smallRNAseq_moonshiner_KO

description:
small RNA sequencing from moonshiner knock-out ovaries
small RNAs
smallRNAseq_moonshiner_KO

extract_protocol:
Total RNA extraction with TRIzol
small RNA sequencing as described in PMID: 21890901
Ovaries were hand-dissected into ice-cold PBS


title: smallRNAseq_w1118

description:
small RNA sequencing from w1118 (wildtype) ovaries
small RNAs
smallRNAseq_w1118

extract_protocol:
Total RNA extraction with TRIzol
small RNA sequencing as described in PMID: 21890899
Ovaries were hand-dissected into ice-cold PBS


title: PCHiC_4h_Exp2

description: Genomic DNA


title: PCHiC_4h_Exp1

description: Genomic DNA


title: DRSC_plate3_day3_H9_DRSC31627_FBgn0261283_SREBP

city: Bethesda

first: Brian

last: Oliver

state: MD

address: 50 South Drive

department: LBG

institute: NIDDK, NIH

description: 3day RNAi incubation

data_processing:
Base calling was done with Illumina CASAVA 1.8.2 (Illumina, San Diego, CA).
We aligned short reads from RNA-Seq to Drosophila reference genome release 6 from Flybase (http://flybase.org, McQuilton et al., 2012. PMID:22127867) with ERCC spike-in sequences (Jiang et al., 2011. PMID: 21816910).  We used TopHat 2.0.10 aligner in doing that (Trapnell et al., 2009. PMID: 19289445) with -g 1 and -G parameters.  We used Flybase gene model version 6.06 with only major chromosome arms.
We counted gene level mapped reads using HTSeq (Anders et al., 2014. PMID: 25260700) with strand-specific, otherwise default parameters.  We also measured transcript abundance in FPKMs (rapnell et al., 2009. PMID: 19289445) using Cufflinks 2.2.1 (Trapnell et al., 2010. PMID: 20436464).  We have used -G, -b, and -u parameters.
Genome_build: BDGP release 6 (from FlyBase (http://www.flybase.org)
Supplementary_files_format_and_content: We provide tab delimited text files that were generated from HTseq and Cufflink. HTseq results have information on read counts. Cufflinks results contain gene level FPKM values.

growth_protocol: We used protocols (Ramadan et al., 2007. PubMed ID: 17853882) from the Harvard Drosophila RNAi Screening Center (Boston, MA) to culture Drosophila S2R+ cells (http://www.flyrnai.org/DRSC-PRC.html ).

extract_protocol:
We used RNeasy 96 kits (Qiagen, Valencia, CA) according to the product manual (Protocol for Isolation of Total RNA from Animal Cells using spin technology, Cat#19504) to isolate total RNA from the RNAi-treated cells
Strand-specific RNA-Seq libraries were prepared as in our previous study (Lee et al., 2016, PMID:27599372) with the following exceptions. 1) We used 1ug of total RNA as an input, 2) we purified results from reverse transcription, second strand synthesis, end-repair, and A-tailing with 1.6 volume of MagNA beads (Rohland et al., 2012. PMID:22267522) instead of 1.9 volume, and 3) We ran 13 cycles of PCR amplification instead of 14 cycles. Briefly, we selected polyA+ RNA from total isolated RNA using Dynabeads Oligo(dT)25 (Life Technologies, Grand Island, NY).  Purified polyA+ RNA was fragmented with spike-in RNA from External RNA Control Consortium Standard Reference Material 2374 ( Zook et al., 2012. PMID:22859977).  We used pool 78A (Lee et al., 2016.  PMID:27512518).  We performed strand specific reverse transcription from the fragmented RNA by adding dUTP in the second strand synthesis.  We ligated 24 differently indexed adaptors from TruSeq v2 kit (Illumina, San Diego, CA) to the end-repaired, and 3’ adenylated DNA.  We removed dUTP-incorporated strands using DNA glycoxylase before PCR amplification of libraries.  Our libraries were sequenced on Illumina HiSeq 2500 (76 bp read-length, single-end. Illumina, San Diego, CA).

treatment_protocol: 2.7 ug of dsRNA in 27ul were pre-plated to 96-well plates before adding cells.  We removed serum-containing media from confluent S2R+ cells by centrifugation.  We re-suspended cells in serum-free media to have 100,000 cells per ml.  53 ul of cell suspension was transferred to each well of the dsRNA-containing plates.  We incubated cells for 30 minute, and added 160 ul serum-containing media (10% FBS).   We incubated cells in humidified chamber at 25C for 24, 48, and 72 hours.  After incubation, we removed media with 12-well vacuum wands, and lysed cells with 150 ul of RLT lysis buffer (Qiagen, Valencia, CA, Cat. No. 79216).  We stored lysed cells at -80C until RNA isolation.

value: S2R+ cells

tag: RNAi target gene id (flybase)

tag: RNAi target gene name

value: SREBP

tag: RNAi reagent from harvard drosophila rnai screening center

tag: time point

value: 3day RNAi incubation


title: PCHiC_D0_Exp2

description: Genomic DNA

tag: cell type


title: HiC_D2_Exp1

description: Genomic DNA


title: PCHiC_D0_Exp1

description: Genomic DNA


title: HiC_4h_Exp2

description: Genomic DNA


title: HiC_D2_Exp2

description: Genomic DNA


description:
small RNA sequencing from ovaries of flies with alpha-Catenin promoter deletion and expression of alpha-Catenin cDNA in trans
small RNAs
smallRNAseq_aCat_promoter_deletion_with_aCat_transgene

extract_protocol:
Total RNA extraction with TRIzol
small RNA sequencing as described in PMID: 21890910
Ovaries were hand-dissected into ice-cold PBS


title: DRSC_plate3_day3_H5_DRSC35747_FBgn0001078_ftz-f1

description: 3day RNAi incubation


title: DRSC_plate3_day3_H6_DRSC40439_FBgn0003870_ttk

description: 3day RNAi incubation

value: ttk


title: HiC_4h_Exp1

description: Genomic DNA


title: DRSC_plate3_day3_H7_DRSC37439_FBgn0259938_cwo

description: 3day RNAi incubation

value: cwo


title: HiC_D0_Exp2

description: Genomic DNA


title: DRSC_plate3_day3_H8_DRSC37675_FBgn0003866_tsh

description: 3day RNAi incubation

value: tsh


title: HiC_D0_Exp1

description: Genomic DNA


title: SMC1_D7_Exp2

description: ChIP-enriched DNA

value: SMC1 antibody, Cat#A300-055A; RRID:AB_2192467


title: SMC1_D7_Exp1

description: ChIP-enriched DNA


title: SMC1_D4_Exp2

description: ChIP-enriched DNA


title: SMC1_D4_Exp1

description: ChIP-enriched DNA


title: DRSC_plate3_day3_H3_DRSC31616_FBgn0001168_h

description: 3day RNAi incubation

value: h


title: DRSC_plate3_day3_H1_DRSC40479_FBgn0003345_sd

description: 3day RNAi incubation

value: sd


title: DRSC_plate3_day3_H2_DRSC37384_FBgn0267978_ap

description: 3day RNAi incubation

value: ap


title: DRSC_plate3_day3_H4_DRSC32506_FBgn0052296_Mrtf

description: 3day RNAi incubation

value: Mrtf


title: DRSC_plate3_day3_H12_DRSC40438_FBgn0004198_ct

description: 3day RNAi incubation

value: ct


title: SMC1_D2_Exp2

description: ChIP-enriched DNA


title: SMC1_D2_Exp1

description: ChIP-enriched DNA


title: SMC1_D0_Exp2

description: ChIP-enriched DNA


title: DRSC_plate3_day3_H10_DRSC32737_FBgn0001133_grau

description: 3day RNAi incubation

value: grau


title: SMC1_D0_Exp1

description: ChIP-enriched DNA


title: SMC1_4h_Exp2

description: ChIP-enriched DNA


title: DRSC_plate3_day3_G7_DRSC37681_FBgn0004050_z

description: 3day RNAi incubation

value: z


title: P300_D0_Exp2

description: ChIP-enriched DNA

value: adipocyte


title: DRSC_plate3_day3_G8_DRSC31645_FBgn0003118_pnt

description: 3day RNAi incubation

value: pnt


title: SMC1_4h_Exp1

description: ChIP-enriched DNA


title: DRSC_plate3_day3_G6_DRSC37529_FBgn0267821_da

description: 3day RNAi incubation

value: da


title: DRSC_plate3_day3_G5_DRSCNA_NA_NA

description: 3day RNAi incubation

value: S2R+ cells

value: DRSCNA


title: P300_D0_Exp1

description: ChIP-enriched DNA


title: P300_4h_Exp2

description: ChIP-enriched DNA


title: DRSC_plate3_day3_G4_DRSC30099_FBgn0000611_exd

description: 3day RNAi incubation

value: exd

value: 3day RNAi incubation


title: P300_4h_Exp1

description: ChIP-enriched DNA


title: NCoR_D0_Exp2

description: ChIP-enriched DNA

value: NCoR antibody, Cat#A301-145A; RRID:AB_873085


title: DRSC_plate3_day3_G3_DRSC26006_FBgn0261239_Hr39

description: 3day RNAi incubation


title: DRSC_plate3_day3_G2_DRSC30105_FBgn0052296_Mrtf

description: 3day RNAi incubation


title: DRSC_plate3_day3_G1_DRSC32517_FBgn0016694_Pdp1

description: 3day RNAi incubation


title: NCoR_D0_Exp1

description: ChIP-enriched DNA


title: NCoR_4h_Exp2

description: ChIP-enriched DNA


title: DRSC_plate3_day3_G12_DRSC34905_FBgn0013799_Deaf1

description: 3day RNAi incubation


title: DRSC_plate3_day3_G10_DRSC40910_FBgn0003117_pnr

description: 3day RNAi incubation

value: pnr


title: MED1_D7_Exp2

description: ChIP-enriched DNA

value: MED1/TRAP220 antibody (M-255) Cat#sc-8998, RRID:AB_2144021


title: NCoR_4h_Exp1

description: ChIP-enriched DNA


title: DRSC_plate3_day3_G11_DRSC29174_FBgn0000520_dwg

description: 3day RNAi incubation

value: dwg


title: DRSC_plate3_day3_F9_DRSCNA_NA_NA

description: 3day RNAi incubation


title: DRSC_plate3_day3_H11_DRSC35845_FBgn0011656_Mef2

description: 3day RNAi incubation


title: DRSC_plate3_day3_G9_DRSC32788_FBgn0000097_aop

description: 3day RNAi incubation

value: aop


title: MED1_D7_Exp1

description: ChIP-enriched DNA


title: DRSC_plate3_day3_F8_DRSC23558_FBgn0265784_CrebB

description: 3day RNAi incubation

value: CrebB


title: MED1_D4_Exp1

description: ChIP-enriched DNA


title: DRSC_plate3_day3_F7_DRSC20260_FBgn0002914_Myb

description: 3day RNAi incubation

value: Myb


title: DRSC_plate3_day3_F6_DRSC37640_FBgn0003345_sd

description: 3day RNAi incubation

value: sd


title: MED1_D2_Exp1

description: ChIP-enriched DNA


title: DRSC_plate3_day3_F5_DRSC30106_FBgn0001168_h

description: 3day RNAi incubation

value: h


title: MED1_D0_Exp1

description: ChIP-enriched DNA


title: MED1_D0_Exp2

description: ChIP-enriched DNA


title: DRSC_plate3_day3_F2_DRSC25100_FBgn0004198_ct

description: 3day RNAi incubation

value: ct


title: DRSC_plate3_day3_F3_DRSC25984_FBgn0013799_Deaf1

description: 3day RNAi incubation


title: MED1_4h_Exp2

description: ChIP-enriched DNA

value: adipocyte


title: DRSC_plate3_day3_F1_DRSC37618_FBgn0039044_p53

description: 3day RNAi incubation


title: DRSC_plate3_day3_F4_DRSC27700_FBgn0001222_Hsf

description: 3day RNAi incubation

value: Hsf


title: MED1_4h_Exp1

description: ChIP-enriched DNA


title: DRSC_plate3_day3_F12_DRSC31100_FBgn0002914_Myb

description: 3day RNAi incubation


title: HDAC3_D0_Exp1

description: ChIP-enriched DNA

value: HDAC3 antibody (H-99), Cat#sc-11417; RRID:AB_2118706


title: HDAC3_4h_Exp1

description: ChIP-enriched DNA


title: DRSC_plate3_day3_F11_DRSC28794_FBgn0266411_sima

description: 3day RNAi incubation

value: S2R+ cells

value: sima


title: DRSC_plate3_day3_E9_DRSC29930_FBgn0264075_tgo

description: 3day RNAi incubation

value: tgo


title: HDAC2_D0_Exp2

description: ChIP-enriched DNA

value: HDAC2 antibody, Cat#A300-705A; RRID:AB_533399


title: HDAC2_D0_Exp1

description: ChIP-enriched DNA


title: DRSC_plate3_day3_E7_DRSC24447_FBgn0003964_usp

description: 3day RNAi incubation

value: usp

value: 3day RNAi incubation


title: MED1_D4_Exp2

description: ChIP-enriched DNA

value: MED1/TRAP220 antibody (M-255) Cat#sc-8998, RRID:AB_2144021


title: HDAC2_4h_Exp2

description: ChIP-enriched DNA


title: DRSC_plate3_day3_E6_DRSC23134_FBgn0004050_z

description: 3day RNAi incubation

value: z


title: DRSC_plate3_day3_F10_DRSC26054_FBgn0016076_vri

description: 3day RNAi incubation

value: vri


title: DRSC_plate3_day3_E5_DRSC16977_FBgn0001297_kay

description: 3day RNAi incubation

value: kay


title: MED1_D2_Exp2

description: ChIP-enriched DNA


title: DRSC_plate3_day3_E4_DRSC37563_FBgn0261283_SREBP

description: 3day RNAi incubation

value: SREBP


title: H3K4me2_D0_Exp1

description: ChIP-enriched DNA

value: H3K4me2 antibody, Cat#9726; RRID:AB_915839


title: DRSC_plate3_day3_E2_DRSCNA_NA_NA

description: 3day RNAi incubation

value: DRSCNA


title: H3K4me2_D0_Exp2

description: ChIP-enriched DNA


title: HDAC2_4h_Exp1

description: ChIP-enriched DNA


title: DRSC_plate3_day3_E1_DRSC34559_FBgn0001222_Hsf

description: 3day RNAi incubation


title: HDAC3_D0_Exp2

description: ChIP-enriched DNA


title: H3K4me1_D0_Exp2

description: ChIP-enriched DNA

value: H3K4me1 antibody, Cat#ab8895; RRID:AB_306847


title: H3K4me1_D0_Exp1

description: ChIP-enriched DNA

value: adipocyte


title: DRSC_plate3_day3_E11_DRSC18766_FBgn0000520_dwg

description: 3day RNAi incubation

value: dwg


title: DRSC_plate3_day3_E12_DRSC37262_FBgn0264075_tgo

description: 3day RNAi incubation


title: HDAC3_4h_Exp2

description: ChIP-enriched DNA

value: HDAC3 antibody (H-99), Cat#sc-11417; RRID:AB_2118706


title: H3K27ac_D0_Exp2

description: ChIP-enriched DNA

value: H3K27ac antibody, Cat#ab4729; RRID:AB_2118291


title: DRSC_plate3_day3_E8_DRSC27606_FBgn0001291_Jra

description: 3day RNAi incubation

value: S2R+ cells

value: Jra


title: DRSC_plate3_day3_D9_DRSC25045_FBgn0013263_Trl

description: 3day RNAi incubation

value: Trl


title: CTCF_D7_Exp1

description: ChIP-enriched DNA

value: CTCF antibody, Cat#07-729; RRID:AB_441965


title: DRSC_plate3_day3_D6_DRSC29336_FBgn0003507_srp

description: 3day RNAi incubation

value: srp

value: 3day RNAi incubation


title: DRSC_plate3_day3_D7_DRSC11122_FBgn0004396_CrebA

description: 3day RNAi incubation

value: CrebA


title: CTCF_D4_Exp1

description: ChIP-enriched DNA


title: CTCF_D4_Exp2

description: ChIP-enriched DNA


title: CTCF_D2_Exp1

description: ChIP-enriched DNA


title: CTCF_D2_Exp2

description: ChIP-enriched DNA


title: DRSC_plate3_day3_D5_DRSC27078_FBgn0003870_ttk

description: 3day RNAi incubation

value: ttk


title: DRSC_plate3_day3_D4_DRSC25219_FBgn0259938_cwo

description: 3day RNAi incubation

value: cwo


title: CTCF_D0_Exp2

description: ChIP-enriched DNA


title: CTCF_D0_Exp1

description: ChIP-enriched DNA


title: DRSC_plate3_day3_E3_DRSC40784_FBgn0267821_da

description: 3day RNAi incubation

value: da


title: DRSC_plate3_day3_D3_DRSC21341_FBgn0040305_MTF-1

description: 3day RNAi incubation


title: CTCF_4h_Exp2

description: ChIP-enriched DNA


title: DRSC_plate3_day3_D2_DRSC08706_FBgn0025725_alphaCOP

description: 3day RNAi incubation

value: alphaCOP


title: CTCF_4h_Exp1

description: ChIP-enriched DNA


title: DRSC_plate3_day3_D1_DRSC30999_FBgn0011766_E2f1

description: 3day RNAi incubation


title: CTCF_D7_Exp2

description: ChIP-enriched DNA


title: H3K27ac_D0_Exp1

description: ChIP-enriched DNA


title: RNA_D7_Exp2

description: RNA


title: DRSC_plate3_day3_E10_DRSC39162_FBgn0000097_aop

description: 3day RNAi incubation

value: aop


title: RNA_D2_Exp2

description: RNA


title: DRSC_plate3_day3_D8_DRSC22973_FBgn0016694_Pdp1

description: 3day RNAi incubation


title: DRSC_plate3_day3_D12_DRSC32152_FBgn0033846_mip120

description: 3day RNAi incubation


title: RNA_D1_Exp2

description: RNA


title: DRSC_plate3_day3_D11_DRSC29572_FBgn0011766_E2f1

description: 3day RNAi incubation


title: RNA_D0_Exp2

description: RNA


title: RNA_D0_Exp1

description: RNA


title: RNA_4h_Exp2

description: RNA


title: RNA_4h_Exp1

description: RNA


title: DRSC_plate3_day3_C8_DRSC29182_FBgn0033846_mip120

description: 3day RNAi incubation


title: K562-invFAF2_clone1-VP-RNF44

city: Montpellier

first: David

last: Martin

address: 1919 route de Mende

country: France

institute: IGMM/CNRS

data_processing:
Library strategy: 4C-seq
Reads were aligned to the hg19 genome assembly using Bowtie2 version 2.1.0 with default parameters.
Aligned reads have been processed with the bioconductor package Basic4CSeq.
Genome_build: hg19 (GRCh37)
Supplementary_files_format_and_content: bedGraph files

growth_protocol: K562 (CCL-243) was grown in RPMI (Gibco) supplemented with 10% FBS (Gold, PAA) at 37 °C, 5% CO2.

extract_protocol:
Chromatin preparation, genome digestion and ligation procedures were performed as described in Stadhouders R. et al., Nat Protoc. 2013 Mar;8(3):509-24. doi: 10.1038/nprot.2013.018.
Library preparation was performed as described in Stadhouders R. et al., Nat Protoc. 2013 Mar;8(3):509-24. doi:10.1038/nprot.2013.018.
The purified DNA was captured on an Illumina flow cell for cluster generation. Libraries were run on Illumina NextSeq500.

treatment_protocol: Cells were not treated.

tag: cell line

value: chronic myelogenous leukemia cell line

tag: enzyme

value: NlaIII/DpnII

tag: viewpoint

value: RNF44 promoter

tag: genotype/variation

value: Inverted FAF2 promoter


title: DRSC_plate3_day3_C7_DRSCNA_FBgn0260635_NA

description: 3day RNAi incubation

value: DRSCNA


title: K562-deltaFAF2_clone1-VP-RNF44

value: FAF2 delta e-promoter


title: K562-deltaFAF2_clone2-VP-RNF44


title: DRSC_plate3_day3_C4_DRSC41279_FBgn0000611_exd

description: 3day RNAi incubation

value: S2R+ cells

value: exd


title: DRSC_plate3_day3_C5_DRSC25347_FBgn0003117_pnr

description: 3day RNAi incubation

value: pnr


title: DRSC_plate3_day3_C6_DRSC26931_FBgn0004606_zfh1

description: 3day RNAi incubation


title: RNA_D7_Exp1

description: RNA

value: adipocyte


title: RNA_D1_Exp1

description: RNA


title: DRSC_plate3_day3_C9_DRSC40777_FBgn0039044_p53

description: 3day RNAi incubation

value: 3day RNAi incubation


title: K562-WT-VP-RNF44

value: Wild type


title: DRSC_plate3_day3_C2_DRSC40819_FBgn0011763_Dp

description: 3day RNAi incubation

value: Dp


title: K562-deltaFAF2_clone2-rep2-VP-FAF2

value: FAF2 promoter


title: DRSC_plate3_day3_C1_DRSC34422_FBgn0004396_CrebA

description: 3day RNAi incubation

value: CrebA


title: K562-WT-rep2-VP-FAF2


title: DRSC_plate3_day3_C11_DRSCNA_NA_NA

description: 3day RNAi incubation


title: K562-deltaFAF2_clone1-rep2-VP-FAF2


title: DRSC_plate3_day3_C12_DRSC37320_FBgn0265784_CrebB

description: 3day RNAi incubation

value: CrebB


title: RNA_D2_Exp1

description: RNA


title: DRSC_plate3_day3_C10_DRSC37552_FBgn0001078_ftz-f1

description: 3day RNAi incubation


title: K562-WT-rep1-VP-FAF2


title: DRSC_plate3_day3_D10_DRSC27330_FBgn0001133_grau

description: 3day RNAi incubation

value: grau


title: K562-deltaFAF2_clone2-rep1-VP-FAF2

value: chronic myelogenous leukemia cell line


title: DRSC_plate3_day3_B8_DRSC03346_FBgn0261239_Hr39

description: 3day RNAi incubation


title: DRSC_plate3_day3_B9_DRSC03740_FBgn0003866_tsh

description: 3day RNAi incubation

value: tsh


title: DRSC_plate3_day3_C3_DRSC04917_FBgn0267978_ap

description: 3day RNAi incubation

value: ap


title: hSS

city: Menlo Park

first: Christina

last: Fan

state: CALIFORNIA

address: 4040, Campbell Ave

institute: BD Biosciences

description: mRNA

data_processing:
BD Resolve pipeline was used to identify the molecular index sequences (position 49-56) and align reads to predfined cell label sequences (position 1-8,22-30,43-51) on Read1, and to align reads (Read2) to the human transcriptome (hg19, gencode annotation) using STAR with default parameters. A unique cell label is a combination of the cell label sequences in the 3 indicated positions on Read1.
cell label sequence for position 1-8:TTAACGCG,TGTGCTGA,TCGAGTTG,TTGGCGAT,GCATCCAA,TATACGCG,GAGTTCTG,ATGCTTGG,TCCTCAAC,CCTAATCC,CGAACGTT,TTGCCGTT,AGCTCAAC,ATCCGGTA,AAGCAAGC,CACTCAGA,CAAGGTTC,AGGATGGT,TAGGATGG,ACACACAC,CACATCTC,TCCACTTG,TCCATGCT,GCAAGGAT,GGATATGG,TTGCATCC,CAGACACT,GGATAACG,CTAGTGGT,TGTGACTG,TATAGCCG,TCTGGTCT,ATGGATGG,GCAAGGTA,TGGATGCT,CAACTCGA,GCATGCAT,TCCTTGGT,AAGCAACG,AACGCGAA,TCTCTGTG,AATACGCC,AACGATGG,TCGACAAC,TAGCCGAT,TGCACTTC,GATCCATG,TGGTTGGT,CAACGTAC,ATTAGCCG,TCCAACCT,CTTGGATG,ACGAACCT,CCATTAGG,GCAAGCTT,ACAGCACT,AGTCTCTC,AGTGAGTG,TTCCATGC,CCATAACG,TGGACTTG,TTAAGCCG,TCGACTTC,GCATGGTT,CTTCCAAC,CAACCATG,TCCTAGCT,GTGACACT,GCAATAGG,TGACACAC,TTGCCGAA,TTCCAAGG,AACCTAGC,AAGCCGTT,AACGCCTA,TTAACGGC,ATCCTTGC,GGTACGAA,AACGGCTT,CACTGAGT,GGTTCCTT,CTTCTCCT,CTAGGTTG,AAGGATGC,TCAGACTG,TGTGGACT,TTCCTTGG,TAGGTTCG,AGCTTGGT,ATTACGCG,AACCATGC,TGCAACGT,AGGTGAAC,TGTCTGTC,GATCTGGT,CTGACTGA,
cell label sequence for position 22-30:TGGTCTTC,CAACAGCT,GGTACGTT,TGGAGTTC,TTGCAAGC,TTATGGCG,CAACACGT,CCAACCAA,TCTCTGAC,CTTGTGCT,AGCTTCCT,AGTCCTCT,GAACCATC,CTTGACGA,CATCCTTG,CCTATTGC,TGGTTCCT,TGCATGGT,CCTAGCTT,GCATAAGG,CACATGAC,CCAATTCC,AAGCCGAA,ACTCAGAC,GCAATTCG,GATCTCCT,GACTCTGA,AGCAACGA,ACTGACTG,GCCGTTAA,CAACCAAC,AAGCTTGC,CAAGTGCT,GATGACCT,ACCTGATG,GTCTGTGA,CAACCTTC,AGGTACGA,ATCCAAGC,AGCTCATG,GTTCTGCT,TGCTTGCT,ACTGCTCT,ATGCCGTA,CCATATCC,AGTGGTCT,TGAGTCTG,CGATATGC,CGATTAGC,GATGACGA,TTCGATGG,GTTGCTTC,CTGAACTG,TAGCCGTA,ACCACAAC,GAACGATG,TGCTGTTC,AATTGCCG,TGTGCTCT,GGATGGAT,GATGGATG,GATGGTTC,ACGATGCT,CAAGGATG,GCAACGTT,TGCTCTTG,ATTACGGC,TGCTGATG,CTCAACTC,TGCACATG,CTTGGTTC,ACTGGACT,CTCTTCTC,TGGTTCGA,GTTCCATC,GGTTGCAT,TGGTACGT,GCGGTATT,GTTGTCCT,TTGCTTCG,ACACGACT,CATGGTAC,TCCTCATG,GGATGCAA,TTGCATGG,GATCCAAC,GATCAGCT,TGACCTGA,TGCATCCT,CGAATAGG,ATCCATCC,CTTCCTTC,AGGTCTTG,TGACTGAC,GGTTCGAT,GTTGCATG,
cell label sequence for position 43-51:CTGACTCT,GACTGACT,TAGCTTGG,CATGGTTG,CCATAAGC,TTCGAACG,CTCTCTGA,ACACTGTG,ATATGCCG,AACGGCAA,GCGGTTAT,ACTGTCTC,TGTCCACT,GAACTGCT,TCAGACAC,ACACACTG,ATAACGCC,GGTATTGC,CCAATACG,TCTCTCTC,TCTGACTC,AAGGTACG,GATGTGCT,GCCGTATA,TCTGTCTG,GTCTTCTG,TCGACATG,TCCTTCCT,CGATAAGG,TTGCAACG,GGTTCCAA,TGAGGAGT,GCATCCTT,CATCTGCT,CACACTCT,GATGAGGT,CGAACCAT,ACTGACAC,ACTCCACT,GCCGTAAT,TGAGGTCT,GTCTTGTC,AAGCATCC,CTGTTCTG,TCGAGTAC,CCTAATGG,CATGTCCT,GATGCTTG,GTGAGTCT,GCCGATAT,TCAGCTCT,GACTCTCT,CGAACGAA,TGCAAGCT,CAGAACTC,TCTGAGTG,ATGCATGC,AGTGGAGT,TGACCTCT,AGGAGTAC,TAGCTTCC,CTAGCAAC,GGTATTCG,CCTTCGAT,CTTCAGCT,TTCCGGTT,TCCAAGGT,GACATCTG,GGTTAAGG,GGATGGTA,TCCTGTTG,ACCTGAAC,ACTGAGTC,GTCTAGTG,ATGCGGTT,AACCAAGG,AGGACATG,AAGGTTGG,GCATTACG,TTGGTTGG,CGAATTGC,ACGACATC,CCAATAGC,ATGGTTGC,CTAGTCCT,TGACTCTC,GCAACCAT,AGGAGTTG,TCTGGTGA,AGCTGTTG,CTTGCTTG,CTCAAGTG,ATGCAAGG,CTAGGTAC,CCATGGTA,TATTGGCG,
Reads with the identical cell label, molecular index, and gene identity, are collapsed and counted as 1 molecule
Counts of molecule per gene per cell label are organized in a cell by gene matrix format, which is the final processed data file
Cell labels with proportion of mitochondrial gene >25% are filtered out
Psuedogenes and mitochondrial genes from each cell are removed for clustering and visualization
Genome_build: hg19
Supplementary_files_format_and_content: Data tables containing number of molecules per gene per cell

source_name: hiPSC-derived 3D culture (subpallium)

growth_protocol: Differentiation of human iPSC into hCS and hSS

extract_protocol:
Dissociated single cells from each of hCS and hSS were loaded onto the BD Resolve single cell gene expression analysis system
mRNA from individual cells were captured by barcoded oligo probes on a BD Resolve bead. Capture probes contain cell labels (total of 24 bases) and molecule index (8 randomers) such that when cDNAs were extended by reverse transcriptase, each cDNA molecule was tagged with a cell specific sequence and a molecular index sequence. cDNA were then universally amplified following by adaptor ligation. 3' end of transcripts were enriched using random priming and sequenced on Illumina NextSeq.

treatment_protocol: Patterning for pallium and subpallium; single cell trituration (no reporter used)


title: hCS

description: mRNA

source_name: hiPSC-derived 3D culture (pallium)


title: DRSC_plate3_day3_B5_DRSC23794_FBgn0000567_Eip74EF

description: 3day RNAi incubation


title: YapTaz_r1

city: Manchester

first: Zoi

last: Diamantopoulou

address: Wilmslow road, Withington

department: Cell Signalling

country: United Kingdom

institute: CRUK MI

data_processing:
Basecall files generated from NextSeq sequencing run were converted to FASTQ format with Illumina’s bcl2fastq v2.17.1.14. Lane-wise alignment was performed by Tophat2 (version 2.0.12) to human reference genome GRCh37 (Ensembl v75) with the added parameter "--library-type fr-firststrand --no-novel-juncs". Resulting lane-wise BAM files was filtered for uniquely mapped reads with the samtools command "samtools view -b -q 50". The filtered BAM from the same sequence library was merged into one BAM file used for downstream analysis. Read counts from BAM files were extracted under the R environment (R v3.1.0) with the package Rsubread version 1.13.13. Only genes with at least 1 count per million in at least three samples were retained for differential expression analysis. Count table was loaded to edgeR v3.8.5 and differentially expressed genes between groups were identified by exact test function.
Genome_build: GRCh37

growth_protocol: SW620 cells were cultured in DMEM-AQ media supplemented with 10% FBS.

extract_protocol:
Total RNA was collected using the Rneasy Mini Kit (Qiagen) according to the manufacture's instructions.
Indexed PolyA libraries were prepared using  200ng of Total RNA and 14 cycles of amplification in the Agilent Sure Select Strand Specific RNA Library Prep Kit for Illumina Sequencing  (Agilent  Cat No: G9691A). Libraries were quantified by qPCR using a Kapa Library Quantification Kit for Illumina sequencing platforms  (Kapa Biosystems Inc. Cat No: KK4835). Paired-end  75bp sequencing was carried out by clustering  2pM of the equimolar pooled libraries on a NextSeq 500 sequencer  (Illumina inc.)

treatment_protocol: SW620 cells were transfected with NT, TIAM1 or YAP/TAZ siRNAs using Lipofectamine RNAiMAX according to the manufacture's instructions.

tag: source tissue

value: colon

value: YAP/TAZ KD


title: DRSC_plate3_day3_B6_DRSC29116_FBgn0003118_pnt

description: 3day RNAi incubation

value: pnt


title: DRSC_plate3_day3_B4_DRSCNA_NA_NA

description: 3day RNAi incubation

value: S2R+ cells

value: DRSCNA


title: DRSC_plate3_day3_B3_DRSC03462_FBgn0010287_Trf

description: 3day RNAi incubation

value: Trf


title: DRSC_plate3_day3_B2_DRSC03320_FBgn0024371_E2f2

description: 3day RNAi incubation


title: DRSC_plate3_day3_B1_DRSC31026_FBgn0004606_zfh1

description: 3day RNAi incubation


title: Tiam1_r3

value: TIAM1 KD


title: Tiam1_r1


title: Tiam1_r2


title: NT_r3

value: WT


title: DRSC_plate3_day3_B11_DRSC26287_FBgn0032940_Mondo

description: 3day RNAi incubation

value: Mondo

value: 3day RNAi incubation


title: NT_r2


title: K562-deltaFAF2_clone1-rep1-VP-FAF2

value: NlaIII/DpnII

value: FAF2 promoter

value: FAF2 delta e-promoter


title: SW480_WT_TAZ_Sigma

data_processing:
Basecall files generated from NextSeq sequencing run were converted to FASTQ format with Illumina’s bcl2fast v2.17.1.14. Lane-wise alignment was performed by bowtie2 v2.2.1 to o human reference genome GRCh37 (Ensembl v75) with default parameters. Generated SAM files from bowtie2 alignment were converted to BAM files by samtools v0.1.19. Parameters for samtools SAM to BAM conversion: -bS -q 10 -F 260. Resulting lane-wise BAM files from the same sequence library was merged into one BAM file used for downstream analysis. Duplicated reads were removed with picard tools version 1.129. Peaks were called using the program macs2 (version 2.1.0.20150420) with the settings "-p 0.001".
Genome_build: GRCh37
Supplementary_files_format_and_content: BigWig files were generated with the homer program. The function "makeUCSCfile" was used with the following paramerters "-o auto -noadj -bigWig style chipseq -fsize 1e20"

source_name: SW480_wild type

growth_protocol: SW480 cells were cultured in DMEM-AQ media supplemented with 10% FBS.

extract_protocol:
Micrococcal nuclease treated chromatin was pulled down with TAZ antibody.
The MicroPlex Library Preparation Kit v2 (Diagenode) was used to prepare indexed libraries from 1ng of ChIP DNA. The resulting libraries were then size-selected (selecting a 200-800bp range) by adding 0.55X volume of AMPure XP beads (Beckman Coulter). To the supernatant, 0.3X the original sample volume of AMPure XP beads were added. The supernatant was discarded and the beads washed with 70% ethanol before drying and eluting the size-selected library. Library quality was checked using the Agilent Bioanalyzer. Libraries were quantified by qPCR using the KAPA Library Quantification Kit for Illumina (Kapa Biosystems Inc.). 1.7 pM pooled libraries were loaded onto the NextSeq 500 and 2x75bp sequencing was carried out using a NextSeq 500/550 High Output v2 kit (Illumina Inc.).

value: TAZ antibody


title: NT_r1


title: DRSC_plate3_day3_B10_DRSC25554_FBgn0011656_Mef2

description: 3day RNAi incubation


title: DRSC_plate3_day3_B12_DRSC31967_FBgn0040305_MTF-1

description: 3day RNAi incubation


title: SW480_WT_Input


title: DRSC_plate3_day3_A9_DRSC34303_FBgn0003507_srp

description: 3day RNAi incubation

value: srp


title: SW480_TIAM1 KO_TAZ_Sigma

source_name: SW480_TIAM1 knockout

value: TIAM1 knockout


title: DRSC_plate3_day3_A7_DRSC31022_FBgn0001297_kay

description: 3day RNAi incubation

value: kay


title: DRSC_plate3_day3_A8_DRSC31684_FBgn0013263_Trl

description: 3day RNAi incubation

value: Trl


title: DRSC_plate3_day3_B7_DRSC35831_FBgn0034534_maf-S

description: 3day RNAi incubation

value: maf-S


title: SW480_TIAM1 KO_Input

value: colon


title: YapTaz_r2

value: YAP/TAZ KD


title: DRSC_plate3_day3_A5_DRSC35830_FBgn0034534_maf-S

description: 3day RNAi incubation

value: S2R+ cells


title: DRSC_plate3_day3_A6_DRSC41069_FBgn0032940_Mondo

description: 3day RNAi incubation


title: DRSC_plate3_day3_A2_DRSC30795_FBgn0001291_Jra

description: 3day RNAi incubation

value: Jra


title: DRSC_plate3_day3_A1_DRSC37534_FBgn0011763_Dp

description: 3day RNAi incubation

value: Dp


title: YapTaz_r3


title: U266_H3K27AC

city: Boston

first: Nikhil

middle: C

last: Munshi

state: MA

address: 450 Brookline Avenue

department: Hematologic Oncology

institute: Dana-Farber Cancer Institute

description: ChIP-Seq for histone modification H3K27AC in U266 cells

data_processing:
Sequenced reads were aligned to HG19 referenece genome using bowtie2 with default parameters with the exception of '-k 1'
Peaks were called using MACS1.4.2 with  p-val =1e-9 and background datasets as described in characteristic: input
Genome_build: hg19
Supplementary_files_format_and_content: Processed ChIP-Seq data files are in wiggle format

source_name: Multiple Myeloma

growth_protocol: All MM cell lines were cultured in RPMI–1640 media containing 10% fetal bovine serum (FBS, GIBCO, Life technologies, Carlsbad, CA, United States), 2 mmol/L L-glutamine, 100 U/mL penicillin, and 100 mg/mL streptomycin (GIBCO, Life technologies, Carlsbad, CA, United States).

extract_protocol:
Lysates were clarified from sonicated nuclei and chromatin-DNA complexes were isolated with antibody.
Libraries were prepared using the Rubicon Thruplex FD library preparation kit (cat#: 400427) per manufacturers instructions.

tag: input

tag: treatment

tag: barcode

value: CGATGT

value: anti H3K27AC

tag: antibody maker

value: Abcam

tag: antibody catalog number


title: U266_WCE_2

description: ChIP-Seq whole cell extract (WCE) input control in U266 cells

value: ATCACG

tag: antibody


title: U266_E2F1

description: ChIP-Seq for transcription factor E2F1 in U266 cells

value: anti E2F1

value: Cell Signaling

value: Cell Signaling #3742


title: U266_DP1

description: ChIP-Seq for transcription factor DP1 in U266 cells

value: anti DP1

value: Santa Cruz


title: MM1S_E2F1

description: ChIP-Seq for transcription factor E2F1 in MM1.S cells


title: MM1S_WCE_MUNSHI

description: ChIP-Seq whole cell extract (WCE) input control in MM1.S cells


title: MM1S_DP1

description: ChIP-Seq for transcription factor DP1 in MM1.S cells


title: DRSC_plate3_day3_A11_DRSC40711_FBgn0000567_Eip74EF

description: 3day RNAi incubation

value: 3day RNAi incubation


title: DRSC_plate2_day2_H9_DRSC31627_FBgn0261283_SREBP

description: 2day RNAi incubation

value: SREBP

value: 2day RNAi incubation


title: DRSC_plate2_day2_H8_DRSC37675_FBgn0003866_tsh

description: 2day RNAi incubation

value: tsh


title: Sample 22_ YJ_lil-1_9xMyc_1LIL_r7 MethylC-seq

city: La Jolla

first: Julie

middle: Ann

last: Law

address: 10010 North Torrey Pines Rd

department: PLANT MOLECULAR AND CELLULAR BIOLOGY LABORATORY

institute: The Salk Institute for Biological Studies

data_processing:
Illumina Casava software used for basecalling and demultiplexing.
Read Filtering and Mapping: Illumina sequence reads were filtered to remove duplicate reads either with prinseq [71], using the remove exact duplicates option (-derep 1), or the BSseeker2[72] filterReads.py script, using default conditions.  The reads were then mapped to the YJ or LUCH transgene sequences (LUC_transgene6.fa or LUC_transgene7.fa) using bsmap-2.74 [73] with the default parameters, allowing 2 mismatches (-v 2) and allowing uniquely mapping reads and multi-mapping reads with the maximum number of equal best hits set to 2 (-w 2). In addition, the reads were also mapped to the tair10 genome using the BSseeker2 bs_seeker2-align.py script, using the bowtie1 aligner (version bowtie-1.0.0)[74] and allowing for 2 mismatches (-m 2).
Methylation levels: The percent methylation level at each cytosine was calculated using the BSseeker2 bs_seeker2-call_methylation.py script requiring a minimum coverage of 4 reads (-r 4) for mapping to the Tair10 genome or the bsmap methratio.py script, reporting loci with zero methylation ratios (-z) for the YJ and LUCH transgenes. In house scripts were used to convert the resulting .CGmap or .txt files, respectively, into wiggle files containing the percent methylation levels of cytosines covered by at least 4 reads in the CG, CHG, and CHH contexts individually.
DMR calling: DMRs were identified using parameters outlined in Stroud et al. [54] using in house scripts. Briefly, the genome was split into 100bp, non-overlapping bins and the methylation level across each bin in the CG, CHG, or CHH context was calculated independently using the percent methylation values in the wiggle files. The methylation values in each bin were then compared between two samples to call DMRs with the following requirements: (1) To account for 100bp regions of the genome with low numbers of cytosines and for regions that display lower than average coverage, only bins with at least 4 cytosines in the given context that are covered by at least 4 reads in both samples being compared were included in the DMR analysis.  (2) Only bins with an absolute change in methylation of 0.4, 0.2, and 0.1 for the CG, CHG, and CHH contexts, respectively, and with an adjusted p-value of ≤0.01 were identified as DMRs.
Genome_build: TAIR10; LUC_Transgene6.fa; LUC_Transgene7.fa
Supplementary_files_format_and_content: wiggle (.wig) file, header with Chromosome, and two columns: position, % methylation
Supplementary_files_format_and_content: bed file, which has 3 columns:Chr#, start position, end position

organism: Arabidopsis thaliana

source_name: YJ_lil-1_9xMyc_1LIL_r7 MethylC-seq

growth_protocol: Arabidopsis seeds were surface-sterilized and then planted on either half-strength Murashige and Skoog (MS) media supplemented with 8% agar and 1% sucrose or Linsmaier and Skoog (LS) media (Caissen, LSP03) supplemented with 8% agar. Seed were then stratified at 4°C for three days. Plants were grown in a growth incubator at 23°C under either continuous light or short day conditions (8 hrs light, 16 hrs dark). 10-day-old seedlings were used for all of the experiments.

extract_protocol:
For the MethylC-seq libraries corresponding to the “_r2, _r5, and_B” series (Table S4), genomic DNA was extracted using the DNeasy Plant Mini  Kit (Qiagen, Cat# 69104). One microgram of genomic DNA was sonicated into  fragments 150 to 300 bp in length using a Diagenode Bioruptor, followed by purification with the PureLink PCR Purification Kit (Invitrogen, Cat#  K3100-01). For all the remaining MethylC-seq libraries (Table S4), genomic  DNA was extracted from 10-day-old seedlings using the DNeasy Plant Mini Kit  (Qiagen, Cat# 69104). 2 ug of genomic DNA was sonicated into 200 bp fragments using a Covaris S2 Sonicator, followed by purification with Sera-Mag Magnetic SpeedBeads (ThermoScientific, Cat# 65152105050250)."
For the MethylC-seq libraries corresponding to the '_r2, _r5, and_B' series  (Table S4), the sonicated and purified genomic DNA fragments were repaired using the End-It DNA End-Repair Kit (Epicentre, Cat# ER0720), and the DNA fragments were purified using the Agencourt AMPure XP-PCR Purification system (Beckman Coulter, Cat# A63880). The purified DNAs were adenylated at the 3? end using the polymerase activity of Klenow Fragment (3?"5? exo-) (New England Biolabs, Cat# M0212), followed by purification using the Agencourt AMPure XP-PCR Purification system. The methylated adapters in the TruSeq DNA Sample Preparation Kit (Illumina, Cat# FC-121-2001) were ligated to the DNA fragments using T4 DNA Ligase (New England Biolabs, Cat# M0202). After purification using AMPure XP beads, less than 400 ng DNA was subjected to bisulfite conversion using the MethylCode Bisulfite Conversion Kit (Invitrogen, Cat# MECOV-50). PfuTurbo Cx Hotstart DNA polymerase (Agilent, Cat# 600414) and the following PCR conditions were used for amplification: 95°C for 2 minutes; 9 cycles of 98°C for 15 seconds, 60°C for 30 seconds and 72°C for 4 minutes; and 72°C for 10 minutes. The resulting libraries were quantified, pooled and sequenced using HiSeq 2000 with the 101-cycle single-end sequencing mode (Illumina). For all the remaining MethylC-seq libraries (Table S4), the sonicated and purified genomic DNA fragments were repaired using the End-It DNA End-It DNA End-Repair Kit (Epicentre, Cat# ER81050), and the DNA fragments were purified using Sera-Mag Magnetic SpeedBeads. The purified DNAs were adenylated at the 3? end using the polymerase activity of Klenow Fragment (3?"5? exo-) (New England Biolabs, Cat# M0212), followed by purification using Sera-Mag Magnetic SpeedBeads. The methylated adapters in the NEXTflex Bisulfite-Seq Barcodes Kit (BIOO Scientific, Cat# 511911) were ligated to the DNA fragments using T4 DNA Ligase (New England Biolabs, Cat# M0202). After purification using Sera-Mag Magnetic SpeedBeads, the DNA was subjected to bisulfite conversion using the MethylCode Bisulfite Conversion Kit (Invitrogen, Cat# MECOV-50). KAPA HiFi HotStart Uracil+ ReadyMix PCR Kit (KAPA Biosystems, Cat# KK2801) and the following PCR conditions were used for amplification: 95°C for 2 minutes; 98°C for 30 seconds; 4 cycles of 98°C for 15 seconds, 60°C for 30 seconds and 72°C for 4 minutes; and 72°C for 10 minutes. After purification using Sera-Mag Magnetic SpeedBeads, the libraries were quantified, pooled and sequenced at single-end 50 base-pair using the Illumina HiSeq 2500 v4 at the Salk NGS Core. Raw sequencing data was de-multiplexed and converted into FASTQ files using CASAVA (v1.8.2)

tag: ecotype

value: Columbia

tag: tissue

value: ten-day-old seedlings

value: YJ_rdr6-11_lil-1 + 9xMyc-LIL


title: DRSC_plate2_day2_H6_DRSC40439_FBgn0003870_ttk

description: 2day RNAi incubation

value: S2R+ cells

tag: RNAi target gene id (flybase)

tag: RNAi target gene name

value: ttk


title: DRSC_plate2_day2_H5_DRSC35747_FBgn0001078_ftz-f1

description: 2day RNAi incubation

tag: RNAi reagent from harvard drosophila rnai screening center

tag: time point


title: Sample 23_ YJ_lil-1_9xMyc_2LIL_r7 MethylC-seq

source_name: YJ_lil-1_9xMyc_2LIL_r7 MethylC-seq


title: DRSC_plate2_day2_H7_DRSC37439_FBgn0259938_cwo

description: 2day RNAi incubation

tag: cell type

value: cwo


title: Sample 20_ YJ_r7 MethylC-seq

source_name: YJ_r7 MethylC-seq


title: DRSC_plate3_day3_A4_DRSC33100_FBgn0266411_sima

description: 3day RNAi incubation

value: sima


title: DRSC_plate2_day2_H4_DRSC32506_FBgn0052296_Mrtf

description: 2day RNAi incubation

value: Mrtf


title: Sample 21_ YJ_lil-1_r7 MethylC-seq

source_name: YJ_lil-1_r7 MethylC-seq


title: Sample 19_Ler_YJ_mbd7-5 + MBD73xHA_r6 MethylC-seq

source_name: Ler_YJ_mbd7-5 + MBD73xHA_r6 MethylC-seq

value: Landsberg erecta

value: YJ_rdr6-11_mbd7-5 + MBD7-3xHA


title: Sample 18_Ler_YJ_mbd7-5_r6 MethylC-seq

source_name: Ler_YJ_mbd7-5_r6 MethylC-seq


title: DRSC_plate3_day3_A3_DRSC31698_FBgn0016076_vri

description: 3day RNAi incubation

value: vri


title: DRSC_plate3_day3_A12_DRSC37538_FBgn0024371_E2f2

description: 3day RNAi incubation


title: DRSC_plate2_day2_H3_DRSC31616_FBgn0001168_h

description: 2day RNAi incubation

value: h


title: Sample 17_Ler_YJ_r6 MethylC-seq

source_name: Ler_YJ_r6 MethylC-seq


title: DRSC_plate3_day3_A10_DRSC36079_FBgn0003964_usp

description: 3day RNAi incubation

value: usp


title: DRSC_plate2_day2_H1_DRSC40479_FBgn0003345_sd

description: 2day RNAi incubation

value: sd


title: Sample 14_rdd_r5 MethylC-seq

source_name: rdd_r5 MethylC-seq

value: rdd


title: Sample 15_Ler_r6 MethylC-seq

source_name: Ler_r6 MethylC-seq

value: wild type (Landsberg erecta)


title: Sample 16_Ler_mbd7-5_r6 MethylC-seq

source_name: Ler_mbd7-5_r6 MethylC-seq


title: Sample 13_Col_r5 MethylC-seq

source_name: Col_r5 MethylC-seq

value: wild type (Columbia)


title: DRSC_plate2_day2_H2_DRSC37384_FBgn0267978_ap

description: 2day RNAi incubation

value: ap

value: 2day RNAi incubation


title: Sample 12_Col_r1_r2 MethylC-seq

source_name: Col_r1_r2 MethylC-seq

value: Columbia

value: ten-day-old seedlings


title: DRSC_plate2_day2_H12_DRSC40438_FBgn0004198_ct

description: 2day RNAi incubation

value: S2R+ cells

value: ct


title: DRSC_plate2_day2_G9_DRSC32788_FBgn0000097_aop

description: 2day RNAi incubation

value: aop


title: DRSC_plate2_day2_G8_DRSC31645_FBgn0003118_pnt

description: 2day RNAi incubation

value: pnt


title: DRSC_plate2_day2_H10_DRSC32737_FBgn0001133_grau

description: 2day RNAi incubation

value: grau


title: Sample 9_YJ_r5 MethylC-seq

source_name: YJ_r5 MethylC-seq


title: Sample 10_YJ_lil-1_r5 MethylC-seq

source_name: YJ_lil-1_r5 MethylC-seq


title: DRSC_plate2_day2_G7_DRSC37681_FBgn0004050_z

description: 2day RNAi incubation

value: z


title: Sample 5_LUCH_r2 MethylC-seq

source_name: LUCH_r2 MethylC-seq