title: 2_CERV_BBB029_J7: Brain Endothetlail Cells (BECs)_Cerebellum _P0D7 _rep1

city: Chilly-Mazarin

first: Dominique

last: Lesuisse

address: 1, avenue Pierre Brossolette

department: Rare and neurologic diseases therapeutic area

country: France

institute: Sanofi

data_processing:
ArrayStudio (QIAGEN Inc.)
Filtering step to remove reads corresponding to rRNAs as well as reads having low quality score or shorter than 25 nt
Reads were  mapped to the macaque genome (Cyno.WashU2013) using OSA4 (Omicsoft Sequence Aligner, version 4) and quantified using Ensembl.R94 model of transcriptome.
Gene expression was measured calculating Fragments per Kilobase, per Million Mapped reads (FPKM) and raw counts.
Genome_build: Cyno.WashU 2013 genome based on the contigs assembled from a WGS project submitted by Washington University (WashU) in 2013
Supplementary_files_format_and_content: Matrix table with raw gene counts for every gene and every sample
Supplementary_files_format_and_content: Matrix table with FPKM values for every gene and every sample

organism: Macaca fascicularis

extract_protocol:
Cell pellets  were lysed using QIAzol Lysis Reagent (Qiagen, # 79306).
Libraries were prepared  using the NEBNext Ultra II Directional RNA Library Prep Kit for Illumina (New England Biolabs, #E 7765S) with the NEBNext rRNA Depletion Kit (New England Biolabs, #E6310L)
Paired-end sequencing (75 cycles x2) on the NovaSeq 6000 instrument (Illumina) using the NovaSeq 6000 SP Reagent Kit (300 cycles; # 20027465).

treatment_protocol:
P0D0: Brain microvessel isolation obtained through enzymatic digestion and gradient centrifugation yields a vascular fraction (P0D0) that is highly enriched in endothelial cells
P0D7: P0D0 cells were cultivated in endothelial cell medium (EBM-2 supplemented with Kit EGM-2 MV SingleQuots, Lonza) for 7 days (P0D7)
P1D7: Following 7 days of expansion, BECs from cortex, were further singularized and re-plated de novo for further 7-day cell expansion (P1D7)

tag: brain structure

value: Cerebellum

tag: fraction


title: 18_Striat_BBB036_J7: Brain Endothetlail Cells (BECs)_Striatum _P0D7 _rep7

status: Public on Jul 23 2020

type: SRA

library_source: transcriptomic

molecule: total RNA

value: Striatum


title: HCT116_EpCAMlow_1

city: Rotterdam

first: Mathijs

last: Verhagen

address: Doctor Molewaterplein 40

country: Netherlands

institute: Erasmus Medical Center

data_processing:
IIllumina paired-end reads of 76 bases were trimmed by removing the TrueSeq adapter sequences using Trimmomatic (v.0.33)
Subsequently, the reads were mapped in a two-pass procedure to the human reference genome build hg38 with the RNA-seq aligner STAR (v2.4.2a) and the Homo sapiens GENCODE v23 annotation.
Raw counts were summed with the summarize overlaps function with union mode from the Bioconductor Genomic Alignments package (v1.14.0).
Supplementary_files_format_and_content: tab-delimited text files include Ensemble Gene Id  and read count

molecule: polyA RNA

organism: Homo sapiens

source_name: colon cancer cell line

growth_protocol: Cell populations were sorted from established colon cancer cell lines grown in standard culture medium. Sorted cells were put in culture for 4 days before RNA was extracted.

extract_protocol:
RNA was harvested using Trizol reagent.
RNA-Seq was performed at the Erasmus MC Center for Biomics (Rotterdam, The Netherlands) according to manufacturer’s instructions (Illumina) and using the TruSeq RNA sample prep kit v2. Briefly, polyA containing mRNA molecules were purified using poly-T oligo attached magnetic beads. Following purification, the mRNA was fragmented into ~200 bp fragments using divalent cations under elevated temperature. The cleaved RNA fragments were copied into first strand cDNA using reverse transcriptase and random primers. This was followed by second strand synthesis using DNA polymerase I and RNaseH treatment. These cDNA fragments were end repaired, a single A base was added and Illumina adapters were ligated. The products were purified and size selected on gel and enriched by PCR. The PCR products were purified by Qiaquick PCR purification and used for cluster generation according to the Illumina cluster generation protocols (www.illumina.com).

treatment_protocol: no treatment was applied

tag: tissue

value: colon


title: H3K27ac_ARPE-19-1

city: Stanford

first: Fabian

last: Grubert

address: 300 Pasteur Drive

country: USA

institute: Stanford University

library_source: genomic

data_processing:
Illumina TruSeq adaptor sequences were trimmed using cutadapt in paired-end mode (non-default parameters: -q 15 -O 4 -m 20)
Reads were aligned to hg19 using bowtie (version 0.12.8; non-default parameters: -m 1 --fr --chunkmbs 500 -n 2 -l 50 --mapq 40 –best) after which duplicate reads were removed using Picard MarkDuplicates
peaks were called using MACS28 (non-default parameters: -q 0.01). Peaks across all samples were combined and overlapping peaks were merged to form a single interval spanning all overlapping peaks. Peaks seen in <2 samples and peaks overlapping ENCODE blacklisted regions [https://sites.google.com/site/anshulkundaje/projects/blacklists]  were removed from further consideration
Genome_build: hg19

molecule: genomic DNA

source_name: retinal pigmented epithelium

growth_protocol: All cell lines were cultured following the manufacturer’s recommendations

extract_protocol:
Nuclear lysates were sonicated using a Branson 250 Sonifier (power setting 2, 100% duty cycle for 7 x 30-s intervals). Clarified lysates corresponding to 20 million cells were treated with 1-5ug of antibody against H3K27ac [Abcam #4729]) coupled to Protein G Dynabeads (Life Technologies #10003D, New York). The protein-DNA complexes were washed with RIPA buffer and eluted in 1% SDS TE at 65 °C
ChIP DNA sequencing libraries were generated according to Illumina DNA  TruSeq DNA Sample Preparation Kit Instructions (Illumina Part # FC-121-2001, San Diego,  CA).

treatment_protocol: Cells were crosslinked with formaldehyde at a final concentration of 1% for 10 minutes at room temperature. The reaction was quenched with Glycine at a final concentration of 125mM

tag: disease

value: normal

tag: germ layer

value: Ectoderm


title: 4_CERV_BBB035_J7: Brain Endothetlail Cells (BECs)_Cerebellum _P0D7 _rep2


title: Stout39_12mo_S20

city: Oklahoma City

first: VICTOR

middle: AFRIYIE

last: ANSERE`

state: Oklahoma

address: Genes and Human Disease Research Program MS57, Oklahoma Medical Research Foundation 825 NE 13th Street

department: PHYSIOLOGY

institute: UNIVERSITY OF OKLAHOMA HEALTH SCIENCES CENTER

data_processing:
Sequenced reads were trimmed, aligned to the Mm10 mouse genome build, differential expression statistics and correlation analyses were performed using Strand NGS software package (Agilent).
Adapter trimming, fixed 5bp trim from 5’ and 2bp from 3’ ends and a maximum number of one novel splice was allowed per read. Read 1 and Read 2 were aligned directionally with Read 1 in the reverse and Read 2 in the forward orientation. Transcripts considered for quantitation and analysis had an average read count value >20 in at least 100% of the samples in at least one time-point.
A one-way ANOVA across the time points with a Benjamini-Hochberg Multiple Testing Correction was used to analyse differential expression across the timepoints. For anscripts that met this statistical criterion, we employed a fold change >|1.25| pairwise cutoff 6M, 9M, or 12M vs 3M) to  eliminate genes which were statistically significant but unlikely to be biologically significant.
Genome_build: mm10
Supplementary_files_format_and_content: matrix table of raw gene counts of all genes for each sample
Supplementary_files_format_and_content: matrix table of normalized gene counts for each sample
Supplementary_files_format_and_content: matrix table with all differentiallly expressed genes across the timepoints for each sample

organism: Mus musculus

source_name: Ovary

extract_protocol:
Ovaries were isolated and frozen with dry ice. RNA were isolated using the AllPrep DNA/RNA mini kit (Qiagen, Germantown, MD). 25ug of RNA with integrity >7 were subsequently used for the library construction.
Libraries were constructed using the NEBNext Ultra II Directional Library Prep Kit for Illumina (New England Biolabs Inc., Ipswich, MA) and standard Illumina sequencing protocols.

tag: strain

tag: age

tag: Sex

value: female


title: 16_Striat_BBB032_J7: Brain Endothetlail Cells (BECs)_Striatum _P0D7 _rep5


title: H3K27ac_GM12878-1

source_name: B cell; lymphoblastoid

value: Immortalized with EBV

value: Mesoderm


title: H3K27ac_DU145-2

source_name: prostate; metastatic to brain

value: carcinoma

value: Endoderm


title: Stout36_12mo_S17


title: H3K27ac_DU145-1


title: H3K27ac_ARPE-19-2


title: 4200-2%glu_3

city: Dallas

first: xueliang

last: lyu

state: Texas

address: 6001 Forest Park Road

department: Physiology

institute: UT Southwestern Medical Center

data_processing:
Sequencing reads were mapped to reference sequences using tophat2 with parameters --min-intron-length 4 --max-intron-length 5000 --library-type fr-firststrand --max-multihits 20 --read-mismatches 3 --read-gap-length 3 --read-edit-dist 3 --mate-inner-dist 0 --mate-std-dev 50 --no-coverage-search.
Reads Per Kilobase Million (RPKM) were calculated using cufflinks with parameters --min-intron-length 4 --max-intron-length 5000 --library-type fr-firststrand --multi-read-correct --upper-quartile-norm --frag-bias-correct
Genome_build: The reference sequence is whole genome sequence (supercontigs) with gff annotation file from NCBI database (assembly NC12).
Supplementary_files_format_and_content: Tab-delimited text files include RPKM values for each sample.

organism: Neurospora crassa

source_name: mycelia

growth_protocol: Fresh conidia (one week post inoculation on slants) of the WT strain and the cpc-1 deletion mutant were cultured in 2% glucose medium (1 × Vogel’s, 2% glucose) with or without 3-AT, or 0.1% glucose medium (1 × Vogel’s, 0.1% glucose and 0.17% arginine) at room temperate under constant light.

extract_protocol:
Total RNA was extracted using Trizol reagents (Invitrogen) and treated with DNase (Turbo DNase, Ambion) accoding to standard RNA extraction protocal.
Libraries were constucted using Illumina TruSeq reagents. All RNA libraries were prepared for sequencing using standard Illumina protocols.

treatment_protocol: For the samples cultured in 2% glucose medium or 0.1% glucose medium, no special treatment was performed. For the samples treated with 3-AT, the cultures were treated with 5 mM 3-AT for 8 h before sample collection.

value: wild-type (WT) strain FGSC4200

value: mycelia

tag: culture conditions


title: intervertebral disc of wild type SD rats

city: shanghai

first: george

last: yue

address: Room 4B, the 3rd Building, No. 401, Caobao Road, Xuhui District, Shanghai, China, 200233

country: China

institute: Genergy Bio-technology (Shanghai) Co., Ltd

data_processing:
CellRanger was used to base-calling, alignment, filtering and generation of normalized abundance measurements.
Read 1: contains the 16bp 10x barcode and the 10bp UMI (bases 17-26).Index contains the sample barcode. Read 2 contains the cDNA information
Genome_build: GRCh38
Supplementary_files_format_and_content: tab-delimited text files include normalized median scaling values for each cell

organism: Rattus norvegicus

source_name: purified wild type rat intervertebral disc cells

extract_protocol: For scRNA-seq, the IVD cells were dissociated into single cells suspensions using Ficoll-Paque buffer (Sigma), and single cell GEMs were generated by 10X genomics Chromium Controller. cDNA library was constructed following the manufacture’s protocol.

value: SD

value: 8 weeks

tag: genotype/variation

value: wild type

value: pooled male and female

tag: cell type

value: intervertebral disc (IVD) cells


title: Stout29_9mo_S15


title: 12_Striat_BBB027_J7: Brain Endothetlail Cells (BECs)_Striatum _P0D7 _rep1


title: Stout28_9mo_S14


title: Mouse-Cortex_HyPBase_scRNA

city: St. Louis

first: Robi

middle: D

last: Mitra

state: Missouri

address: 4515 McKinley Ave

department: Genetics

institute: Washington University in St. Louis

description:
Single cell RNA-seq library
SRRs represent biological replicates.

data_processing:
scRNA-seq libraries were sequenced on either Illumina HiSeq 2500 or NovaSeq S1 machines. Reads were analyzed using 10x Genomics’ Cell Ranger with the following settings: --expect-cells=6000 --chemistry=SC3Pv2 --localcores=16 --localmem=30. The digital gene expression matrices from 10x were then further processed with scanpy (Wolf et al., 2018) for identification of highly variable genes, batch correction, dimensionality reduction, and Louvain clustering. Processed scRNA-seq datasets were stored as .loom files (http://loompy.org). We cross-referenced gene expression data with published datasets (Rosenberg et al., 2018; Rouillard et al., 2016; Saunders et al., 2018; Tasic et al., 2018; Zeisel et al., 2018) to assign cell types.
Genome_build: mm10
Supplementary_files_format_and_content: Loom files contain processed scRNA-seq data

source_name: Mouse cortex

growth_protocol: All mouse experiments were done following procedures described in (Cammack et al., 2019). In brief, we cloned the PB-SRT-tdTomato and HyPBase constructs into AAV vectors. The Hope Center Viral Vectors Core at Washington University in St. Louis packaged each construct in AAV9 capsids. Titers for each virus ranged between 1.1e13 and 2.2e13 viral genomes/ml. All animal practices and procedures were approved by the Washington University in St. Louis Institutional Animal Care and Use Committee (IACUC) in accordance with National Institutes of Health (NIH) guidelines.

extract_protocol:
Mouse cortical tissues were dissociated to single suspensions following a modification of previously published methods (Avey et al., 2018; Saxena et al., 2012). We incubated samples in a papain solution containing Hibernate-A with 5% v/v trehalose, 1x B-27 Supplement, 0.7 mM EDTA, 70 µM 2-mercaptoethanol, and 2.8 mg/ml papain. After incubation at 37ºC, cells were treated with DNaseI, triturated through increasingly narrow fire-polished pipettes, and passed through a 40-micron filter prewetted with resuspension solution: Hibernate-A containing 5% v/v trehalose, 0.5% Ovomucoid Trypsin Inhibitor, 0.5% Bovine Serum Albumin (BSA), 33 µg/ml DNaseI (Worthington), and 1x B-27 Supplement. The filter was washed with 6 ml of resuspension solution. The resulting suspension was centrifuged for 4 minutes at 250 g. The supernatant was discarded. The pellet was then resuspended in 2 ml of resuspension solution and resuspended by gentle pipetting. We eliminated subcellular debris using gradient centrifugation. We first prepared a working solution of 30% w/v OptiPrep Density Gradient Medium mixed with an equal volume of 1x Hank’s Balanced Salt Solution (HBSS) with 0.5% BSA. We then prepared solutions of densities 1.057, 1.043, 1.036, and 1.029 g/ml using by combining the working solution with resuspension solution at ratios of 0.33:0.67, 0.23:0.77, 0.18:0.82, and 0.13:0.87, respectively. We layered 1 ml aliquots of each solution in a 15 ml conical tube beginning with the densest solution on the bottom. The cell suspension was added last to the tube and centrifuged for 20 minutes at 800g at 12ºC. The top layer was then aspirated and purified cells were isolated from the remaining layers. These cells were then resuspended in FACS buffer: 1x HBSS, 2 mM MgCl2, 2 mM MgSO4, 1.25 mM CaCl2, 1 mM D-glucose, 0.02% BSA, and 5% v/v trehalose. Cells were centrifuged for 4 minutes at 250 g, the supernatant was discarded, and the pellet was resuspended in FACS buffer by gentle pipetting. Cells were then sorted based on fluorescence activity. As a gating control, we analyzed cells from cortices injected with AAV9-PB-SRT-tdTomato only. We then collected cells from brains transfected with AAV9-PB-SRT-tdTomato and AAV9-HyPBase whose fluorescence values exceeded the gate. After sorting, cells were centrifuged for 3 minutes at 250 g. The supernatant was discarded and cells were resuspended in FACS buffer at a concentration appropriate for 10x Chromium 3’ scRNA-seq library preparation. Single cell RNA-seq libraries were prepared using 10x Genomics’ Chromium Single Cell 3’ Library and Gel Bead Kit. Each replicate was targeted for recovery of 6,000 cells. Library preparation followed a modified version of the manufacturer’s protocol. We prepared the Single Cell Master Mix without RT Primer, replacing it with an equivalent volume of Low TE Buffer. Gel-in-emulsion (GEM) generation and GEM-RT incubation proceeded as instructed. At the end of Post GEM-RT cleanup, we added 36.5 µl Elution Solution I and transferred 36 µl of the eluted sample to a new tube (instead of 35.5 µl and 35 µl, respectively). The eluate was split into two 18 µl aliquots and kept at –20ºC until ready for further processing. One fraction was kept for single cell calling cards library preparation, while the other half was further processed into a single cell RNA-seq library.
We added the RT Primer sequence to the products of the scRNA-seq aliquot. We created an RT master mix by adding 20 µl of Maxima 5X RT Buffer, 20 µl of 20% w/v Ficoll PM-400, 10 µl of 10 mM dNTPs, 2.5 µl RNase Inhibitor and 2.5 µl of 100 µM 10x_TSO. To this solution we added 18 µl of the first RT product and 22 µl of ddH2O. Finally, we added 5 µl Maxima H Minus Reverse Transcriptase, mixed by flicking, and centrifuged briefly. This reaction was incubated at 25ºC for 30 minutes followed by 50ºC for 90 minutes and heat inactivated at 85ºC for 5 minutes. The solution was purified using DynaBeads MyOne Silane following 10x Genomics’ instructions, beginning at “Post GEM-RT Cleanup – Silane DynaBeads” step D. The remainder of the single cell RNA-seq protocol, including purification, amplification, fragmentation, and final library amplification, followed manufacturer’s instructions.

treatment_protocol: We mixed equal volumes of each virus and performed intracranial cortical injections of the mixture into newborn wild-type C57BL/6J pups (P0-2). As a gating control, we injected one litter-matched animal with AAV9-PB-SRT-tdTomato only. After 2 to 4 weeks, we sacrificed mice and dissected the cortex (8 libraries) or hippocampus (1 library).

value: cerebral cortex


title: 10_Hippo_BBB032_J7: Brain Endothetlail Cells (BECs)_Hippocampus_P0D7 _rep6

value: Hippocampus


title: 17_Striat_BBB035_J7: Brain Endothetlail Cells (BECs)_Striatum _P0D7 _rep6

value: Striatum


title: goat_low_spleen3

city: Chengdu

first: Siyuan

last: Feng

state: Sichuan

address: Huiming Road #211, Wenjiang District

department: College of Animal Science and Technology

institute: Sichuan Agriculcural University

data_processing:
Incipient bioinformatics analysis (base calling) was performed with CASAVA 1.8 (Illumina) to generate raw reads (in FASTQ form).
Raw reads was subjected to a series of stringent filters (such as removing low quality-reads, repeated sequences and adaptor sequences). Filtered sequences were then mapped to goat reference genome (assembly ARS1) with stringent criteria (0 mismatch in the whole length) using Bowtie. Next, mappable reads were submitted to miRDeep version 2.0.0.7 to detect miRNAs with default parameters, while mature miRNA sequences of goat and all other annotated mammalian species in miRbase release 21 were selected as reference. miRNAs with readcount no less than 3 in at least one sample were remained for further analysis.
Genome_build: goat representative genome (ARS1)
Supplementary_files_format_and_content: expression of identified miRNAs in each library

organism: Capra hircus

source_name: spleen tissue from low-altitude goat

extract_protocol:
Three adult females (~3 years old) from each of the low-altitude goat and Tibetan goat populations residing at distinct altitudes in southwest China (600 m and 3000 m) were used in this study. Animals were fed with free access to food and water, and killed humanely to ameliorate suffering. All of the animals procedures complied the guidelines for the care and use of experimental animals established by the Ministry of Agriculture of China. Six typical hypoxia-sensitive tissues (heart, kidney, liver, lung, longissimus muscle and spleen) were rapidly excised from the carcass, immediately frozen in liquid nitrogen, and then stored at −80 °C until RNA isolation.
Small RNA libraries were constructed using the Illumina TruSeq Small RNA Sample Prep kit. Libraries were assessed using the Agilent 2200 TapeStation and sequenced on the Illumina HiSeq 2500 platforms.

tag: population

value: Chuandong White Goat

value: spleen

value: > 3 year


title: goat_low_muscle2

source_name: muscle tissue from low-altitude goat

value: muscle


title: goat_low_lung3

source_name: lung tissue from low-altitude goat

value: lung


title: Stout37_12mo_S18

value: female


title: goat_low_heart2

source_name: heart tissue from low-altitude goat

value: heart


title: goat_low_kidney2

source_name: kidney tissue from low-altitude goat

value: kidney


title: 15_Striat_BBB031_J7: Brain Endothetlail Cells (BECs)_Striatum _P0D7 _rep4


title: 14_Striat_BBB029_J7: Brain Endothetlail Cells (BECs)_Striatum _P0D7 _rep3


title: Stout16_6mo_S10


title: goat_low_lung2


title: Stout35_12mo_S16


title: goat_low_muscle1


title: goat_low_muscle3


title: Stout15_6mo_S9


title: goat_low_kidney3


title: goat_low_heart3


title: 13_Striat_BBB028_J7: Brain Endothetlail Cells (BECs)_Striatum _P0D7 _rep2


title: goat_low_liver3

source_name: liver tissue from low-altitude goat

value: liver


title: 11_Hippo_BBB035_J7: Brain Endothetlail Cells (BECs)_Hippocampus_P0D7 _rep7


title: Stout14_6mo_S8


title: Stout26_9mo_S13


title: 9_Hippo_BBB031_J7: Brain Endothetlail Cells (BECs)_Hippocampus_P0D7 _rep5


title: goat_low_spleen2


title: Stout25_9mo_S12


title: 5_Hippo_BBB027_J7: Brain Endothetlail Cells (BECs)_Hippocampus_P0D7 _rep1


title: goat_low_lung1


title: Stout24_9mo_S11


title: 8_Hippo_BBB030_J7: Brain Endothetlail Cells (BECs)_Hippocampus_P0D7 _rep4


title: 7_Hippo_BBB029_J7: Brain Endothetlail Cells (BECs)_Hippocampus_P0D7 _rep3


title: goat_low_kidney1


title: 6_Hippo_BBB028_J7: Brain Endothetlail Cells (BECs)_Hippocampus_P0D7 _rep2


title: goat_low_spleen1


title: goat_low_heart1


title: Stout13_6mo_S7


title: goat_low_liver1


title: goat_low_liver2


title: 22_TronC_BBB030_J7: Brain Endothetlail Cells (BECs)_Brainstem_P0D7 _rep4

value: Brainstem


title: Stout11_6mo_S6


title: 24_TronC_BBB032_J7: Brain Endothetlail Cells (BECs)_Brainstem_P0D7 _rep6


title: Stout9_3mo_S5


title: goat_high_spleen3

source_name: spleen tissue from high-altitude goat

value: Tibetan Goat


title: 23_TronC_BBB031_J7: Brain Endothetlail Cells (BECs)_Brainstem_P0D7 _rep5


title: goat_high_kidney3

source_name: kidney tissue from high-altitude goat


title: Stout7_3mo_S4


title: 21_TronC_BBB029_J7: Brain Endothetlail Cells (BECs)_Brainstem_P0D7 _rep3


title: goat_high_liver3

source_name: liver tissue from high-altitude goat


title: goat_high_heart3

source_name: heart tissue from high-altitude goat


title: goat_high_lung3

source_name: lung tissue from high-altitude goat


title: goat_high_kidney2


title: goat_high_muscle2

source_name: muscle tissue from high-altitude goat


title: goat_high_muscle3


title: Stout6_3mo_S3

source_name: Ovary


title: goat_high_liver2


title: goat_high_lung2


title: 20_TronC_BBB028_J7: Brain Endothetlail Cells (BECs)_Brainstem_P0D7 _rep2


title: 19_TronC_BBB027_J7: Brain Endothetlail Cells (BECs)_Brainstem_P0D7 _rep1


title: Stout4_3mo_S1


title: goat_high_muscle1


title: 22_BBB036_P1J7: Brain Endothetlail Cells (BECs)_Cortex-P1D7_rep6

value: Cortex


title: goat_high_heart1


title: goat_high_kidney1


title: 21_BBB035_P1J7: Brain Endothetlail Cells (BECs)_Cortex-P1D7_rep5


title: endothelial_lineage_positive_cells

city: Charlottesville

first: Gary

middle: K

last: Owens

state: VA

address: 415 Lane Road MR5 Building Room 1322

institute: University of Virginia

data_processing:
Basecalls performed using CASAVA
Reads were aligned against mouse genome mm10 using the CellRanger software (10x Genomics).
Gene-barcode matrices were analyzed in R using Seurat v342. Cells were filtered for 200-5000 reads per UMI, 10% or less mitochondrial and less than 5% hemoglobin gene content.
Significant principal components of variation (PCs) were calculated using JackStraw test with 10000 repetitions, and clusters were calculated with 19 PCs
Differential expression analysis was done using MAST
Genome_build: mm10
Supplementary_files_format_and_content: Cell-gene-barcode matrices

source_name: Atherosclerosis plaques

extract_protocol:
RNA-seq: Individual atherosclerotic plaques from the brachiocephalic artery were removed from underlying media with forceps and deposited into FACS buffer (1% BSA in PBS) plus 1 ug/mL Actinomycin D,(Gibco, #11805017). Samples were then minced and digested with 1mL Liberase (Roche, #355374) plus 1ug/mL ActinomycinD for 1 hour at 37 degrees, then resuspended in PBS plus 0.04% UltraPure non-Acetylated BSA (ThermoFisher #AM2616) and filtered through 30um filters on ice. Samples were then either counted and submitted directly for scRNAseq or sorted by flow cytometry. Samples for flow were stained with Sytox Blue viability dye and sorted with a BD FACSAria Fusion cell sorter with gating on live, single cells that were tdTomato+, GFP+, or eYFP+. Cells were collected into LoBind tubes containing 0.04% non-acetylated BSA.
RNA-seq: 2,000 cells in each group were targeted in Chromium 10X genomics libraries, which after barcoding, were pooled and sequenced on the Illumina MiSeq, 150 cycle high-output.
ChIP-seq: Segments of the aorta from the arch to the aortic root and up to the carotid bifurcation isolated from 18 week western diet fed  SMC-Oct4+/+, and SMC-Oct4Δ/Δ (n = 15 each group) mice were snap-frozen in liquid nitrogen and then processed for ChIP assays using the  Oct4-specific antibody and sequenced using Illumina TruSeq Chip Library Kit and sequenced on the HiSeq2000. Sequencing reads were aligned to the mouse genome release M21 (GRCm38.p6) using the BOWTIE alignment tool39. These aligned reads were then processed involving removal of duplicate reads and format conversions using the SAMtools40 suite and converted into bam/bai format and loaded in the Integrative Genomics Viewer for visualization. The reads were also converted to BED format and peaks were identified using MACS241 callpeaks function against the mouse genome with a qvalue < 0.05.
ChIP-seq: Illumina TruSeq Chip Library Kit according to the manufacturer's protocol and sequenced on the HiSeq2000

treatment_protocol: Animals were maintened in a Western Diet for 18 weeks

value: Sorted eYFP+ endothelial cells

tag: diet

value: Western Diet


title: goat_high_spleen1


title: endothelial_lineage_unsorted_cells

value: Mixture of cell types (SMC, Epithelial, Macrophage, others)


title: goat_high_heart2


title: goat_high_liver1


title: goat_high_spleen2


title: TgBAC(dbx1b:GFP) positive cells 2

city: Boston

first: Tony

middle: Yu-Chen

last: Tsai

state: MA

address: 200 Longwood Avenue, WAB 536

department: Systems Biology

institute: Harvard Medical School

data_processing:
Sequenced reads were filtered by fastqc 0.11.3, then mapped to GRCz10 whole genome using bowtie 1.1.1 with parameters -q -n 2 -l 28 -e 1000 -I 0 -X 1000 -k 1 -m 200
Reads Per Kilobase of exon per Megabase of library size (RPKM) were calculated.
Genome_build: GRCz10
Supplementary_files_format_and_content: tab-delimited text files include gene names, gene length, mapped count, unmapped count
Supplementary_files_format_and_content: excel spreadsheet with gene names and transcript count in each sample

organism: Danio rerio

source_name: FACS sorted dbx1b (+) neural progenitors

extract_protocol:
14hpf zebrafish embryos were dissected to remove the brain and tail region. The trunk region is dissociated by mechanical pipetting in FACSMax solution. The dissociated cells are resuspended in PBS, FACS sorted for GFP positive cells. The sorted cells are spun down and the pellet was flash frozen in Trizol for RNA extraction.
RNA libraries were prepared for sequencing using standard Illumina protocols

tag: genotype

value: WT


title: 19_BBB031_P1J7: Brain Endothetlail Cells (BECs)_Cortex-P1D7_rep3


title: 20_BBB032_P1J7: Brain Endothetlail Cells (BECs)_Cortex-P1D7_rep4


title: 18_BBB030_P1J7: Brain Endothetlail Cells (BECs)_Cortex-P1D7_rep2


title: aorta

source_name: aorta


title: TgBAC(dbx1b:GFP) positive cells 1


title: bca_unsorted_03


title: goat_high_lung1


title: TgBAC(olig2:GFP) positive cells 1

source_name: FACS sorted olig2 (+) neural progenitors


title: bca_unsorted_01


title: Stout5_3mo_S2


title: TgBAC(olig2:GFP) positive cells 2


title: Aorta Adventitia Macrophage rep 3

first: Maxim

last: Artyomov

address: 425 S Euclid Ave

department: Pathology & Immunology

institute: WASHINGTON UNIVERSITY IN ST LOUIS

data_processing:
quality control: RSeQC version 2.3.
alignment: STAR version 2.0.4b.
quantification: Subread: featureCount version 1.4.5.
Genome_build: GRCm38 (Ensembl release 76)
Supplementary_files_format_and_content: Gene counts tables

source_name: Aorta Adventitia Macrophages

extract_protocol: The cDNA libraries were prepared from cell lysates in 10X Single-Cell Lysis Buffer (Takara-Clontech) containing 5% RNase inhibitor. ds-cDNA was prepared using the SMARTer Ultra Low RNA kit for Illumina Sequencing (Takara-Clontech) per the manufacturer’s protocol. cDNA was fragmented using a Covaris E220 sonicator with the following parameters: duty cycle, 10; intensity, 5; cycles/burst, 200; and time, 180 seconds. The cDNAs were blunt ended, supplemented with an A base at the 3’ end, and then ligated with Illumina sequencing adapters. The ligated fragments were amplified for 12 cycles using primers that incorporated unique index tags. The amplified fragments were sequenced on an Illumina HiSeq-3000 using single reads extending 50 bases.

treatment_protocol: For intima macrophages isolation (C57BL/6) thoracic aorta (including the aortic sinus, arch, descending thoracic aorta) was carefully isolated, and the perivascular fat was removed. To remove the adventitia, the whole aorta was incubated with collagenase II (400U/ml)  with hyaluronidase (90 U/ml) at 37°C for 8 min, and then the adventitia was physically removed. The aortic tissue was cut into 2–5 mm pieces and incubated at 37°C for 70 min with gentle shaking in enzyme mixture containing DNase I (90 U/ml), collagenase I (675 U/ml), collagenase XI (187.5 U/ml), hyaluronidase (90 U/ml). Live aorta intima resident macrophages (PI-CD64+ CD11c+ MHC II+) were sorted out using flow cytometry (BD FACSAria III). For adventitia macrophages (ApoE-/-) whole  aorta (including the aortic sinus, arch, descending thoracic and abdominal aorta) was carefully isolated, and the perivascular fat was removed. To remove the adventitia, the whole aorta was incubated with collagenase II (400U/ml)  with hyaluronidase (90 U/ml) at 37°C for 8 min, and then the adventitia was physically removed and further digested in the same exzyme mixture macrophages (PI- CD64+) were sorted out using flow cytometry (BD FACSAria III).The sorted cells were quickly added to single cell lysis buffer.

value: ApoE-/-

value: Aorta Adventitia Macrophages


title: Aorta Intima Resident Macrophage rep 1

source_name: Aorta Intima Resident Macrophages

value: Aorta Intima Resident Macrophages


title: Aorta Intima Resident Macrophage rep 2


title: TgBAC(nkx2.2a:memGFP) positive cells 1

source_name: FACS sorted nkx2.2a (+) neural progenitors


title: 16_BBB036_J7: Brain Endothetlail Cells (BECs)_Cortex_P0D7_rep8


title: TgBAC(nkx2.2a:memGFP) positive cells 2


title: tdTomato_SMC_sorted_cells01

value: Sorted tdTomato+ SMC cells


title: 15_BBB035_J7: Brain Endothetlail Cells (BECs)_Cortex_P0D7_rep7


title: Aorta Adventitia Macrophage rep 2


title: gfp_SMC_sorted_cells03

value: Sorted eYFP+ SMC cells


title: Aorta Adventitia Macrophage rep 1


title: gfp_SMC_sorted_cells02


title: 14_BBB032_J7: Brain Endothetlail Cells (BECs)_Cortex_P0D7_rep6


title: 13_BBB031_J7: Brain Endothetlail Cells (BECs)_Cortex_P0D7_rep5


title: tdTomato_SMC_sorted_cells02


title: 12_BBB030_J7: Brain Endothetlail Cells (BECs)_Cortex_P0D7_rep4


title: 17_BBB029_P1J7: Brain Endothetlail Cells (BECs)_Cortex-P1D7_rep1


title: gfp_SMC_sorted_cells01


title: SMC_WT_eYFP_sorted


title: 11_BBB029_J7: Brain Endothetlail Cells (BECs)_Cortex_P0D7_rep3


title: SMC_WT_eYFP_unsorted


title: SKIN-HV-011

type: RNA

city: Bethesda

first: Adriana

middle: R

last: Marques

state: MD

institute: NIAID/NIH

hyb_protocol: Hybridization to the GeneChip Human Genome U133A 2.0 or U133Plus 2.0 Array (Affymetrix, St. Clara, CA) performed according to the manufacturer’s protocol.

scan_protocol: Microarrays were scanned at 570 nm using Affymetrix Genechip Scanner 3000 7G.

data_processing: CEL files were analyzed using Affymetrix Expression Console and the MAS5 algorithm. Resulting Signal Intensity values and Present-Absent calls for each of the probe- sets were processed in JMP statistical package (SAS, Cary, NC) using the Mathematical and Statistical Computing Laboratory (MSCL) Analyst’s Toolbox, written by and freely available (http://abs.cit.nih.gov/MSCLtoolbox/) from one of the authors (P.J.M.). The 22 277 probe sets in common between HG-U133A 2.0 and HG-U133 plus 2.0 chips were used for further analysis. The quantile-normalizing, variance-stabilizing “S10” transform was applied to the Signal Intensity data

label: biotin

source_name: skin- control

label_protocol: Complementary RNA (cRNA) synthesis and labeling performed according to the manufacturer’s protocol.

growth_protocol: not applicable

extract_protocol: Total RNA was extracted from skin biopsies using a modified RNeasy Fibrous Tissue Mini kit (QIAGEN, Germantown, MD).

treatment_protocol: All skin specimens were stored at −80°C in RNAlater solution (Ambion) before RNA isolation.

value: skin

tag: type

value: skin- control

tag: batch


title: Klf4_KO_unsorted02


title: 9_BBB027_J7: Brain Endothetlail Cells (BECs)_Cortex_P0D7_rep1


title: 7_BBB037_J0: Isolated brain microvessels_Cortex_P0D0_rep7

source_name: Isolated brain microvessels from Cortex


title: 8_BBB038_J0: Isolated brain microvessels_Cortex_P0D0_rep8


title: HCT-116_HyPBase

description:
Bulk DNA calling cards library
SRRs represent biological replicates.

data_processing:
Reads were demultiplexed by the N7 index sequences added during the final PCR. Read 1 began with the 3 base-pair barcode followed by the end of the transposon terminal repeat, culminating with the insertion site motif (TTAA in the case of piggyBac; TA in the case of Sleeping Beauty) before entering the genome. piggyBac reads were checked for exact matches to the barcode, transposon sequence, and insertion site at the beginning of reads before being hard trimmed using cutadapt (Martin, 2011) with the following settings: -g "^NNNTTTACGCAGACTATCTTTCTAGGGTTAA" --minimum-length 1 --discard-untrimmed -e 0 --no-indels, where NNN is replaced with the primer barcode. Sleeping Beauty libraries were trimmed with the following settings: -g "^NNNTAAGTGTATGTAAACTTCCGACTTCAACTGTA" --minimum-length 1 --discard-untrimmed -e 0 --no-indels. Reads passing this filter were then trimmed of any trailing Nextera adapter sequence, again using cutadapt and the following settings: -a "CTGTCTCTTATACACATCTCCGAGCCCACGAGACTNNNNNNNNNNTCTCGTATGCCGTCTTCTGCTTG" --minimum-length 1. The remaining reads were aligned to the human genome (build hg38) with NovoAlign and the following settings: -n 40 -o SAM -o SoftClip. Aligned reads were validated by confirming that they mapped adjacent to the insertion site motif. Successful reads were then converted to calling card format (.ccf.txt; see http://wiki.wubrowse.org/Calling_card) using custom programs (available at https://github.com/arnavm/calling_cards) and visualized on the WashU Epigenome Browser v46 (Zhou et al., 2011) (http://epigenomegateway.wustl.edu/legacy/).
Genome_build: hg38
Supplementary_files_format_and_content: BAM files contain aligned and annotated calling card reads. ccf.txt files contain processed calling card data. Peaks file provides details on regions significantly enriched for calling card insertions.  describes calling card-specific BAM tags.

growth_protocol: Cells were cultured in Dulbecco’s Modified Eagle Medium (DMEM) supplemented with 10% fetal bovine serum (FBS) and 1% of either penicillin-streptomycin or antibiotic-antimycotic. Cells were grown at 37ºC with 5% carbon dioxide (CO2). Media was replenished every 2 days.

extract_protocol:
Total RNA was isolated from each replicate using the RNEasy Plus Mini Kit following manufacturer’s instructions. Briefly, cell pellets were resuspended in 600 µl of Buffer RLT Plus with 1% 2-mercaptoethanol. Cells were homogenized by vortexing. DNA was removed by running lysate through gDNA Eliminator spin columns, while RNA was bound by passing the flow-through over RNEasy spin columns. An on-column treatment with DNaseI was also performed. After washing, RNA was eluted in 40 µl RNase-free H2O. RNA was quantitated using the Qubit RNA HS Assay Kit. We performed first strand synthesis on each replicate with Maxima H Minus Reverse Transcriptase. We mixed 2 µg of total RNA with 1 µl 10 mM dNTPs and 1 µl of 50 µM SMART_dT18VN primer (for a complete list of oligonucleotides, see Table S5), brought the total volume up to 14 µl, and incubated it at 65ºC for 5 minutes. After transferring to ice and letting rest for 1 minute, we added 4 µl 5X Maxima RT Buffer, 1 µl RNaseOUT, and 1 µl of 1:1 Maxima H Minus Reverse Transcriptase diluted in 1x RT Buffer (100 U). The solution was mixed by pipetting and incubated at 50ºC for 1 hour followed by heat inactivation at 85ºC for 10 minutes. Finally, we digested with 1 µl RNaseH at 37ºC for 30 minutes. cDNA was stored at -20ºC.
The PCR conditions for amplifying self-reporting transcripts (i.e. transcripts derived from self-reporting transposons) involved mixing 1 µl cDNA template with 12.5 µl Kapa HiFi HotStart ReadyMix, 0.5 µl 25 µM SMART primer, and either 1 µl of 25 µM SRT_PAC_F1 primer (in the case of puromycin selection) or 0.5 µl of 25 µM SRT_tdTomato_F1 primer (in the case of tdTomato screening). The mixture was brought up to 25 µl with ddH2O. Thermocycling parameters were as follows: 95ºC for 3 minutes; 20 cycles of: 98ºC for 20 seconds–65ºC for 30 seconds–72ºC for 5 minutes; 72ºC for 10 minutes; hold at 4ºC forever. As a control, cDNA quality can be assessed with exon-spanning primers for β-actin (see Table S5 for examples of human primers (Raff et al., 1997)) under the same thermocycling settings. PCR products were purified using AMPure XP beads. 12 µl of resuspended beads were added to the 25 µl PCR product and mixed homogenously by pipetting. After a 5-minute incubation at room temperature, the solution was placed on a magnetic rack for 2 minutes. The supernatant was aspirated and discarded. The pellet was washed twice with 200 µl of 70% ethanol (incubated for 30 seconds each time), discarding the supernatant each time. The pellet was left to dry at room temperature for 2 minutes. To elute, we added 20 µl ddH2O to the pellet, resuspended by pipetting, incubated at room temperature for 2 minutes, and placed on a magnetic rack for one minute. Once clear, the solution was transferred to a clean 1.5 ml tube. DNA concentration was measured on the Qubit 3.0 Fluorometer using the dsDNA High Sensitivity Assay Kit. Calling card libraries from bulk RNA were generated using the Nextera XT DNA Library Preparation Kit. One nanogram of PCR product was resuspended in 5 µl ddH2O. To this mixture we added 10 µl Tagment DNA (TD) Buffer and 5 µl Amplicon Tagment Mix (ATM). After pipetting to mix, we incubated the solution in a thermocycler preheated to 55ºC. The tagmentation reaction was halted by adding 5 µl Neutralization Tagment (NT) Buffer and was kept at room temperature for 5 minutes. The final PCR was set up by adding 15 µl Nextera PCR Mix (NPM), 8 µl ddH2O, 1 µl of 10 µM transposon primer (e.g. OM-PB-NNN) and 1 µl Nextera N7 indexed primer. The transposon primer anneals to the end of the transposon terminal repeat–piggyBac, in the case of OM-PB primers, or Sleeping Beauty, in the case of OM-SB primers–and contains a 3 base pair barcode sequence. Every N7 primer contains a unique index sequence that is demultiplexed by the sequencer. Each replicate was assigned a unique combination of barcoded transposon primer and indexed N7 primer, enabling precise identification of each library’s sequencing reads. The final PCR was run under the following conditions: 95ºC for 30 seconds; 13 cycles of: 95ºC for 10 seconds–50ºC for 30 seconds–72ºC for 30 seconds; 72ºC for 5 minutes; hold at 4ºC forever. After PCR, the final library was purified using 30 µl (0.6x) AMPure XP beads, as described above. The library was eluted in 11 µl ddH2O and quantitated on an Agilent TapeStation 4200 System using the High Sensitivity D1000 ScreenTape.

treatment_protocol: We cotransfected 10-12 replicates of HCT-116 cells with 5 µg of PB-SRT-Puro plasmid and 5 µg PBase plasmid via Neon electroporation Each replicate contained 2e6 cells. As a negative control, we transfected one replicate of HCT-116 cells with 5 µg PB-SRT-Puro plasmid only. We used the following settings–pulse voltage: 1,530 V; pulse width: 20 ms; pulse number: 1. We used the same experimental setup for experiments with PB-SRT-Puro and each of SP1-PBase, HyPBase, and SP1-HyPBase plasmids, as well as with SB-SRT-Puro and SB100X plasmids. After transfection, each replicate was plated into a 10 cm dish. Puromycin was added after 24 hours to a final concentration of 2 µg/ml. Cells were grown under selection for one week, by which time almost all negative control transfectants were dead. After 7 days, we dissociated each replicate with trypsin-EDTA and created single cell suspensions in phosphate-buffered saline (PBS) Aliquots of each replicate were cryopreserved in cell culture media supplemented with 5% DMSO. The remaining cells were pelleted by centrifugation at 300g for 5 minutes. Cell pellets were either processed immediately or kept at -80ºC in RNAProtect Cell Reagent.

tag: cell line

value: colorectal cancer


title: HCT-116_SP1-PBase


title: 10_BBB028_J7: Brain Endothetlail Cells (BECs)_Cortex_P0D7_rep2

tag: brain structure


title: SKIN-HV-010


title: SKIN-HV-009


title: 6_BBB036_J0: Isolated brain microvessels_Cortex_P0D0_rep6

tag: fraction


title: Klf4_KO_sorted_SMCs01


title: KYSE-150-KD, biological rep3

city: Linhai

first: suna

last: zhou

state: zhejiang

address: No. 150 Ximen Street

institute: The Affiliated Taizhou Hospital, Wenzhou Medical University

description: Gene expression data from KYSE-150 KD cells

hyb_protocol: Following fragmentation, 10ug of cRNA were hybridized for 16 hr at 45C on PrimeView Human Gene Expression Array. GeneChips were washed and stained in the GeneChip Fluidics Station 450.

scan_protocol: GeneChips were scanned using the Hewlett-Packard GeneArray Scanner 3000 .

data_processing: The data were analyzed with RMA using Affymetrix default analysis settings and global scaling as normalization method.

source_name: Human esophageal carcinoma cells

label_protocol: Biotinylated cRNA were prepared according to the standard Affymetrix protocol from 12ug total RNA (Expression Analysis Technical Manual, 2001, Affymetrix).

growth_protocol: KYSE-150 NC and KD cell lines were cultured in 1640 supplemented with 10% fetal bovine serum.

extract_protocol: Total RNA were extracted using Trizol. The RNA quantity and quality were measured by NanoDrop 2000.

treatment_protocol: KYSE-150 Cells were transfected with CDKL3-shRNA and NC lentivirus.

tag: transfection


title: SKIN-HV-008

tag: tissue


title: KYSE-150-KD, biological rep1


title: KYSE-150-NC, biological rep3

description: Gene expression data from KYSE-150 NC cells


title: SKIN-HV-004


title: KYSE-150-NC, biological rep1


title: Klf4_WT_sorted_SMCs01


title: KYSE-150-NC, biological rep2


title: SKIN-HV-007


title: SKIN-HV-006


title: SKIN-HV-003


title: KYSE-150-KD, biological rep2


title: Klf4_KO_unsorted01


title: HCT-116_PBase


title: Klf4_WT_sorted_SMCs02


title: Klf4_KO_sorted_SMCs02


title: 5_BBB035_J0: Isolated brain microvessels_Cortex_P0D0_rep5


title: Mjan_NaCNBH3

city: Tel Aviv

first: Aldema

last: Sas-Chen

state: Israel

address: Tel Aviv University

department: Shmunis School of Biomedicine and Cancer Research

country: Israel

institute: Tel Aviv University

data_processing:
Library strategy: ac4C-seq
alignment using STAR (V. 2.5.3a)
base-calling using JACUSA v2.0.0 under default parameters
calculation of misincorporation rates per base
Genome_build: ASM996v1 for Thermococcus kodakarensis, ASM27560v1 for Pyrococcus furiosus, ASM15120v2 was used for Thermococcus sp. AM4, ASM700v1 for Sulfolobus solfataricus and ASM9166v1 for Methanocaldococcus jannaschii. For human GRCh37/hg19 with UCSC Genes annotations. For Saccharomyces cerevisiae samples the sacCer3 assembly.
Supplementary_files_format_and_content: SasChen_TableS2.xlsx an excel file detailing ac4C sites identified in each organism including the level of modification of each site.

organism: Methanocaldococcus jannaschii

source_name: archaeal cells

growth_protocol:
T. kodakarensis and T. sp AM4 strains – TS559 can derivatives thereof – were grown as previously described (Santangelo et al. 2007; Hileman and Santangelo 2012; Gehring et al. 2017) in artificial sea water (ASW) medium supplemented with vitamins and trace minerals. Pyrococcus furiosus strain COM1 was cultured at 75-95˚C in an artificial sea water based medium supplemented with cellobiose, maltose, yeast extract, S˚, trace minerals, cysteine and sodium tungstate as previously described (Lipscomb et al, APPLIED AND ENVIRONMENTAL MICROBIOLOGY, Apr. 2011, p. 2232–2238).
HEK-293T and HeLa (WT and Nat10 depleted) cells were grown in DMEM media.
Saccharomyces cerevisiae strains were grown at 30°C in standard YEP medium (1% yeast extract, 2% Bacto Peptone) supplemented with 2% galactose (YPG), or 2% dextrose (YPD).

extract_protocol:
Total RNA was isolated from T. kodakarensis using TRIZOL according to manufacturer’s protocol. For the indicated samples of T. kodakarensis ,rRNAs were depleted according to  Morlan et al.(Morlan et al. 2012) using reagents provided in the NEBNext® rRNA Depletion Kit (NEB #E6310). For indicated samples purifications of T. kodakarensis ribosomes of the wild-type and TkNat10 knockout were conducted similar to previously documented procedures (Matzov et al. 2017).
Total RNA from Human Cells was extracted using TRIZOL according to manufacturer's protocol. For the indicated samples Poly(A) RNA from yeast and human total RNA was isolated by two rounds of purification using the  GenElute mRNA miniprep kit (Sigma) according to manufacturer's protocol.  500ug total RNA was used per purification column.  Typical yield after two rounds of isolation was 1.2%.
Total RNA was isolated from Yeast using hot acidic phenol.  Briefly, frozen yeast (S. cerevisiae) pellet suspended in 1.0 mL AES buffer (50mM NaOAc, 10mM EDTA ph 8.0, 1% SDS) per 0.5mL pellet volume.  To suspended pellet, 1.0 mL Acid buffered Phenol per mL of AES buffer used was added. Sample was mixed by vortexing and incubated in a 65˚C water bath for 30 min, vortexing every 2 minutes to mix.  Samples were put on ice for 10 minutes and 1.0 mL Chloroform:isoamyl alcohol (24:1) was added for each 1.0 mL phenol used. Sample was vortexed to mix and centrifuged 5000 rcf for 15 minutes. Aqueous layer (top) was transferred to a clean tube and extracted 3X with an equal volume of acid buffered phenol:chlorofom:isoamyl alcohol (24:23:1).  After each extraction sample was centrifuged at 5000 rcf for 10 minutes and aqueous layer transferred to a new tube. A final extraction with chloroform:isoamyl alcohol was carried out to remove residual phenol. Aqueous layer was transferred to a clean tube and RNA was precipitated by the addition of an equal volume of 100% isopropanol and 1/9th volume of 3M Sodium Acetate.  Samples were incubated -20˚C for 30 minutes and centrifuged 12000 rcf at 4˚C for 15 minutes. Supernatant was decanted and the pellet washed with 4 mL ice fold 70% ethanol. RNA pellet was resuspended by briefly heating at 50 ˚C in 1.0 mL 1X TE buffer pH 8.0. Samples were quantified by UV absorbance and stored at -80˚C. Typical extractions were carried out with 1.0 mL volume cell pellets and yielded 20mg of total RNA
Fragmentation, 3' adapter ligation, cDNA synthesis, second adapter ligation and enrichment

treatment_protocol: HEK-293T cells transfected with 3xFLAG-tagged NAT10 and myc-tagged THUMPD1 using FuGENE® 6 transfection reagent (Promega, #E2691) 24 hours prior to harvesting.

tag: genetic background

tag: treatment

tag: growth condition

value: 85 degrees


title: Mjan_mock

value: mock


title: Tkod_ribosomes_NaCNBH3

description: purified ribosomes

organism: Thermococcus kodakarensis


title: SKIN-HV-005


title: Klf4_WT_unsorted02

tag: strain


title: Libhp4

city: San Francisco

first: Jim

last: Wells

state: CA

department: Pharmaceutical Chemistry

institute: ucsf

data_processing:
Library strategy: SPD-NGS
Raw NGS data from Lib 10AA, “*.fastq.gz” sequencing files were converted into a table with DNA sequences, amino acid sequences, counts/frequency, RPM as four columns, which were saved as .csv files for further analysis.
For Lib hP, reads were aligned to a reference dataset using bowtie2 to generate Sam files. Sam files were then converted into Bam files using Samtools, which were then parsed using a suite of in-house analysis tools (R) to make a table with gene, ref.name, individual phage counts/frequency, peptide sequence, etc. as columns. These tables are saved as .csv files for further analysis.
Supplementary_files_format_and_content: Lib10AA data was processed directly by R script. Lib hP data was first aligned to reference dataset using bowtie2 to generate sam file. The sam files were then converted to bam files using Samtools. Bam files  can be processed using R script.

organism: synthetic construct

source_name: phage

growth_protocol: phage was amplified using XL1-Blue culture

extract_protocol:
We used phage amplification overnight cultures for NGS sample preparation after boiling the phage to extract ssDNA as templates for PCR reactions.
For all samples from both Lib 10AA and Lib hP, a barcoding PCR process was performed using primers listed in Supplemental Figure 21.The thermal PCR profile for Lib 10AA was: 98 °C (20 s), 63 °C (15 s), 72 °C (30 s), 15 cycle. The thermal profile for Lib hP was: 98 °C (30 s), 72 °C (30 s), 15 cycle to prevent heterodimerization due to the 24-residue overlap. The number of cycles was determined empirically to prevent product laddering, assessed by agarose gel electrophoresis. PCR products were gel purified on 1.2 % agarose. Libraries were sequenced on a NextSeq or Highseq4000(Illumina) using single-read 50 or 150 base pair reads. A custom sequencing primer was used (order as shown): ATCGGATCCGCCGCTCTGAAAGTACAGATTCTC (reverse, Tm =67 ◦C, GC% = 52) or CTGCAGTATGTGGCATGAATCTGGTGGTGGT (reverse, Tm =67 ◦C, GC% = 52) for Lib 10AA and GTATGACGTCGCCATCCGCAGTTCGAGAAA (forward, Tm =67 ◦C, GC% = 52) for Lib hp.

treatment_protocol: To perform a screening, 1 mL of biotinylated substrate phage library was incubated with 100 µL magnetic StreptAvidin (SA) beads for 30 min and after that, the SA beads were then stringently washed with PBS buffer supplemented with 0.05% Tween20 and 0.2% BSA to get rid of any free phage. After the SA beads were treated with protease of interest, released phage in supernatant were collected and propagated as protease-sensitive pool, and those bound on the beads can also be propagated as protease-resistant pool after released by non-specific proteases like trypsin. After three rounds of selection, phage polymerase chain reaction (PCR) was conducted, followed by NGS to identify sensitive and resistant substrate sequences in each pool.

tag: fragment size

tag: sample prep

value: phage_PCR and gel extraction


title: Tkod_mRNA_95deg_mock

description: rRNA depleted RNA


title: 3_BBB031_J0: Isolated brain microvessels_Cortex_P0D0_rep3


title: SKIN-HV-001


title: Libhp3


title: Tkod_mRNA_85deg_NaCNBH3

description: rRNA depleted RNA


title: Tkod_mRNA_95deg_NaCNBH3

description: rRNA depleted RNA


title: Klf4_WT_unsorted01

tag: cell type


title: 2_BBB030_J0: Isolated brain microvessels_Cortex_P0D0_rep2


title: Tkod_95deg_mock_rep1

data_processing:
Library strategy: ac4C-seq
alignment using STAR (V. 2.5.3a)
base-calling using JACUSA v2.0.0 under default parameters
calculation of misincorporation rates per base
Genome_build: ASM996v1 for Thermococcus kodakarensis, ASM27560v1 for Pyrococcus furiosus, ASM15120v2 was used for Thermococcus sp. AM4, ASM700v1 for Sulfolobus solfataricus and ASM9166v1 for Methanocaldococcus jannaschii. For human GRCh37/hg19 with UCSC Genes annotations. For Saccharomyces cerevisiae samples the sacCer3 assembly.
Supplementary_files_format_and_content: TableS2_SasChen.xlsx an excel file detailing ac4C sites identified in each organism including the level of modification of each site.

growth_protocol: T. kodakarensis and T. sp AM4 strains – TS559 can derivatives thereof – were grown as previously described (Santangelo et al. 2007; Hileman and Santangelo 2012; Gehring et al. 2017) in artificial sea water (ASW) medium supplemented with vitamins and trace minerals. Pyrococcus furiosus strain COM1 was cultured at 75-95˚C in an artificial sea water based medium supplemented with cellobiose, maltose, yeast extract, S˚, trace minerals, cysteine and sodium tungstate as previously described (Lipscomb et al, APPLIED AND ENVIRONMENTAL MICROBIOLOGY, Apr. 2011, p. 2232–2238).

extract_protocol:
Total RNA was isolated from T. kodakarensis using TRIZOL according to manufacturer’s protocol. For the indicated samples of T. kodakarensis ,rRNAs were depleted according to  Morlan et al.(Morlan et al. 2012) using reagents provided in the NEBNext® rRNA Depletion Kit (NEB #E6310). For indicated samples purifications of T. kodakarensis ribosomes of the wild-type and TkNat10 knockout were conducted similar to previously documented procedures (Matzov et al. 2017).
Fragmentation, 3' adapter ligation, cDNA synthesis, second adapter ligation and enrichment

treatment_protocol: growth at various temperatures

value: 95 degrees


title: Libhp2


title: Tkod_95deg_NaCNBH3_rep1


title: SKIN-HV-002


title: Tkod_TK0754del_NaCNBH3_rep2

value: TK0754 deletion


title: 1_BBB029_J0: Isolated brain microvessels_Cortex_P0D0_rep1

data_processing:
ArrayStudio (QIAGEN Inc.)
Filtering step to remove reads corresponding to rRNAs as well as reads having low quality score or shorter than 25 nt
Reads were  mapped to the macaque genome (Cyno.WashU2013) using OSA4 (Omicsoft Sequence Aligner, version 4) and quantified using Ensembl.R94 model of transcriptome.
Gene expression was measured calculating Fragments per Kilobase, per Million Mapped reads (FPKM) and raw counts.
Genome_build: Cyno.WashU 2013 genome based on the contigs assembled from a WGS project submitted by Washington University (WashU) in 2013
Supplementary_files_format_and_content: Matrix table with raw gene counts for every gene and every sample
Supplementary_files_format_and_content: Matrix table with FPKM values for every gene and every sample

extract_protocol:
Cell pellets  were lysed using QIAzol Lysis Reagent (Qiagen, # 79306).
Libraries were prepared  using the NEBNext Ultra II Directional RNA Library Prep Kit for Illumina (New England Biolabs, #E 7765S) with the NEBNext rRNA Depletion Kit (New England Biolabs, #E6310L)
Paired-end sequencing (75 cycles x2) on the NovaSeq 6000 instrument (Illumina) using the NovaSeq 6000 SP Reagent Kit (300 cycles; # 20027465).

value: Cortex


title: SKIN-EM-016

source_name: skin- erythema migrans

value: erythema migrans


title: Libhp1


title: 4_BBB032_J0: Isolated brain microvessels_Cortex_P0D0_rep4


title: Lib_hP_ADAM17_R3


title: Tkod_TK0754del_NaCNBH3_rep1


title: P1_uHC_GFP_rep1_RNA

city: Los Angeles

first: Neil

last: Segil

address: 1425 san pablo st, BCC 503K

department: BCC

institute: University of Southern California

data_processing:
Sequenced reads were mapped to Gencode Mm10v11 genome using STAR (v2.5.4b).
Transcript quantification was performed with RSEM.
Genome_build: mm10
Supplementary_files_format_and_content: TSV files of transcript quantification was generated with RSEM

source_name: Utricle

growth_protocol: see samples column

extract_protocol:
Cells were FACS purified and collected in RNA lysis buffer. Total RNA was extracted with Quick-RNA Microprep kit (Zymo Research).
Libraries were prepared using QIAseq FX Single Cell RNA Library Kit (Qiagen)

tag: developmental stage

value: post natal day 1

value: Utricle

tag: growth protocol

value: Utricles were dissected from P1 mice and enzymatically dissociated with 0.25% trypsin. Cells were pelleted and resuspended in DMEM/F12 and then FACS purified to collect the Atoh1-GFP positive cells.


title: Tkod_85deg_mock_rep4

value: WT


title: P1_uHC_GFP_rep2_RNA

tag: genotype


title: Tkod_85deg_mock_rep2


title: Oct4-KO/KO

source_name: BCA/Arch/region

value: Mixture of cell types (SMC, Epithelial, Macrophage, others)

tag: isolation

value: Segments of the aorta from the arch to the aortic root and up to the carotid bifurcation isolated from 18 week western diet fed  SMC-Oct4+/+, and SMC-Oct4Δ/Δ (n = 15 each group) mice were snap-frozen in liquid nitrogen and then processed for ChIP assays using the  Oct4-specific antibody and sequenced using Illumina TruSeq Chip Library Kit. Sequencing reads were aligned to the mouse genome release M21 (GRCm38.p6) using the BOWTIE alignment tool39. These aligned reads were then processed involving removal of duplicate reads and format conversions using the SAMtools40 suite and converted into bam/bai format and loaded in the Integrative Genomics Viewer for visualization. The reads were also converted to BED format and peaks were identified using MACS241 callpeaks function against the mouse genome with a qvalue < 0.05.

tag: diet

value: Western Diet


title: Lib_hP_ADAM17_R2


title: SKIN-EM-018

value: skin

tag: type

tag: batch


title: Tkod_85deg_NaCNBH3_rep4


title: SKIN-EM-017


title: Tkod_85deg_NaCNBH3_rep3


title: Tkod_75deg_NaCNBH3_rep2

value: 75 degrees


title: Tkod_85deg_NaCNBH3_rep2


title: Lib_hP_ADAM10_R2


title: Mouse whole brain 3

city: La Jolla

first: Benjamin

last: Gastfriend

address: 9500 Gilman Dr

institute: University of California, San Diego

data_processing:
Basecalling was performed by the Illumina HiSeq 2000
FASTQ files were aligned to the mouse (mm10) or human (hg38) genomes using STAR (v 2.6.0b-1) implemented in Galaxy on the public server at usegalaxy.org
Gene-level transcript abundances were calculated using featureCounts (v 1.6.3) implemented in Galaxy on the public server at usegalaxy.org
Normalized transcript abundances (FPKM, TPM) were calculated from raw counts and featureLengths file (output from featureCounts) using Excel
Genome_build: mm10, hg38
Supplementary_files_format_and_content: Raw counts, FPKM values, and TPM values are provided in an Excel file (xlsx)

source_name: Brain

extract_protocol:
Tissue was lysed with buffer RLT (Qiagen) supplemented with 1% beta-mercaptoethanol. The RNeasy Micro Kit (Qiagen) was used to extract total RNA per manufacturer instructions. DNase I treatment was used to remove gDNA.
Total RNA was subjected to rRNA-depletion and libaries prepared using the NuGEN Ovation Single Cell RNA-Seq High Volume Beta Kit according to manufacturer instructions.

value: whole brain


title: Lib_hP_ADAM10_R1


title: Mouse whole brain 1


title: Tkod_75deg_mock_rep1


title: Ssol_mock

organism: Saccharolobus solfataricus

value: 85 degrees


title: Tkod_75deg_NaCNBH3_rep1


title: Tkod_65deg_mock_rep1

value: mock

value: 65 degrees


title: Mouse brain microvessels 3

value: brain


title: Tkod_65deg_NaCNBH3_rep2


title: Tkod_75deg_mock_rep2


title: Ssol_NaCNBH3


title: Lib10AA_for_casps

value: phage_PCR and gel extraction


title: Mouse brain microvessels 2


title: Mouse brain microvessels 1


title: Tkod_65deg_NaCNBH3_rep1


title: Lib_hP_ADAM10_R3


title: Lib10AA_ADAM17_R3


title: Pfur_95deg_mock

organism: Pyrococcus furiosus

value: 95 degrees


title: Tkod_85deg_mock_rep3


title: Pu_1-KO-LPS-4hr-2

city: Ansan

first: Young Gyu

last: Chai

state: Gyeonggi-do

address: 55 Hanyangdaehak-ro

department: Molecualr & Life Science

country: South Korea

institute: Hanyang Univ.

data_processing:
Illumina CASAVA v1.8.2 software was used for basecalling.
To avoid low-quality data, we clipped and trimmed the reads using Trimmomatic (v0.33)
For the analysis of differentially expressed genes, the data of quality-checked reads for each condition were processed with the STAR (version 2.5.1) software based on reference genome sequence (Homo sapiens UCSC hg19) and the differential gene expressed values of each sample were calculated by the DESeq2 Bioconductor package.
The number of reads mapped to gene ID features were counted using STAR.
Genome_build: Homo sapiens UCSC hg19
Supplementary_files_format_and_content: tab-delimited text files including normalized read counts for each sample

source_name: Pu.1 gene has been knocking-out using CRISPR-Cas9 treated with LPS (10ng/ml) for 2 hrs

growth_protocol: The murine microglial cell line BV-2 was cultured in high glucose Dulbecco’s modified Eagle’s medium supplemented with 10% fetal bovine serum and 1% penicillin-streptomycin (Life Technologies, California, USA) in a humidified CO2 incubator.

extract_protocol:
Total RNA was extracted using RNAiso Plus (Takara, Shiga, Japan) and QIAGEN RNeasy® Mini kit (QIAGEN, Hilden, Germany). hMSCs or LPS-treated hMSCs were completely lysed with RNAiso Plus, then 200 μl of chloroform was added, and the tubes inverted for 5 min. The mixture was centrifuged at 12,000 x g for 15 min at 4°C, and the upper phase was placed into a new tube and 600 μl of 70% ethanol was added. The mixture was applied to an RNeasy mini column. The column was washed with wash buffer. To elute, RNase-free water (30 μl) was added directly onto the RNase mini column, followed by centrifugation at 12,000 x g for 3 min at 4°C. For ribosomal RNA (rRNA) depletion in total RNA, the RiboMinus Eukaryote kit (Life Technologies, Carlsbad, CA) was employed according to the manufacturer’s instructions. RNA libraries were created using the NEBNext® Ultra™ directional RNA library preparation kit for Illumina® (New England Biolabs, Ipswich, MA). rRNA-depleted total RNA was fragmented into small pieces using divalent cations at elevated temperatures. The first-strand cDNA was synthesized using reverse transcriptase and random primers, and then second-strand cDNA synthesis was performed using DNA polymerase I and RNase H. The cDNA fragments were processed using an end-repair reaction after the addition of a single ‘A’ base, followed by adapter ligation. These products were purified and amplified through PCR to generate the final cDNA library.
RNA libraries were prepared for sequencing using standard Illumina protocols

treatment_protocol: Cells were treated with different concentrations of LPS (Sigma-Aldrich, St. Louis, USA) for different durations.

tag: cell line

value: BV2 cells

value: microglia

value: Pu.1 KO

value: LPS-treated


title: Tkod_55deg_mock_rep1

value: WT

value: 55 degrees


title: Mouse whole brain 2


title: Lib10AA_ADAM17_R2


title: Pu_1-KO-LPS-4hr-1


title: Pfur_95deg_NaCNBH3


title: Tkod_85deg_mock_rep1


title: Oct4-WT/WT

value: Mixture of cell types (SMC, Epithelial, Macrophage, others)

value: Segments of the aorta from the arch to the aortic root and up to the carotid bifurcation isolated from 18 week western diet fed  SMC-Oct4+/+, and SMC-Oct4Δ/Δ (n = 15 each group) mice were snap-frozen in liquid nitrogen and then processed for ChIP assays using the  Oct4-specific antibody and sequenced using Illumina TruSeq Chip Library Kit. Sequencing reads were aligned to the mouse genome release M21 (GRCm38.p6) using the BOWTIE alignment tool39. These aligned reads were then processed involving removal of duplicate reads and format conversions using the SAMtools40 suite and converted into bam/bai format and loaded in the Integrative Genomics Viewer for visualization. The reads were also converted to BED format and peaks were identified using MACS241 callpeaks function against the mouse genome with a qvalue < 0.05.


title: Lib10AA_ADAM17_R1


title: Lib10AA_for_ADAMs


title: Pfur_85deg_mock


title: Tkod_85deg_NaCNBH3_rep1


title: BV2-Pu1-K-2

source_name: Pu.1 gene has been knocking-out using CRISPR-Cas9

value: not treated


title: BV2-Pu1-K-1


title: Lib10AA_ADAM10_R3


title: Pfur_85deg_NaCNBH3


title: Lib10AA_ADAM10_R2


title: Tkod_85deg_deacetylation_rep1

value: deacetylation


title: Pfur_75deg_mock

value: 75 degrees


title: Ssol_deacetylation

value: 85 degrees


title: HepG2_FOXA2-HyPBase_scRNA

data_processing:
scRNA-seq libraries were sequenced on either Illumina HiSeq 2500 or NovaSeq S1 machines. Reads were analyzed using 10x Genomics’ Cell Ranger with the following settings: --expect-cells=6000 --chemistry=SC3Pv2 --localcores=16 --localmem=30. The digital gene expression matrices from 10x were then further processed with scanpy (Wolf et al., 2018) for identification of highly variable genes, batch correction, dimensionality reduction, and Louvain clustering. Processed scRNA-seq datasets were stored as .loom files (http://loompy.org). We cross-referenced gene expression data with published datasets (Rosenberg et al., 2018; Rouillard et al., 2016; Saunders et al., 2018; Tasic et al., 2018; Zeisel et al., 2018) to assign cell types.
Genome_build: hg38
Supplementary_files_format_and_content: Loom files contain processed scRNA-seq data

extract_protocol:
Single cell RNA-seq libraries were prepared using 10x Genomics’ Chromium Single Cell 3’ Library and Gel Bead Kit. Each replicate was targeted for recovery of 6,000 cells. Library preparation followed a modified version of the manufacturer’s protocol. We prepared the Single Cell Master Mix without RT Primer, replacing it with an equivalent volume of Low TE Buffer. Gel-in-emulsion (GEM) generation and GEM-RT incubation proceeded as instructed. At the end of Post GEM-RT cleanup, we added 36.5 µl Elution Solution I and transferred 36 µl of the eluted sample to a new tube (instead of 35.5 µl and 35 µl, respectively). The eluate was split into two 18 µl aliquots and kept at –20ºC until ready for further processing. One fraction was kept for single cell calling cards library preparation, while the other half was further processed into a single cell RNA-seq library.
We added the RT Primer sequence to the products of the scRNA-seq aliquot. We created an RT master mix by adding 20 µl of Maxima 5X RT Buffer, 20 µl of 20% w/v Ficoll PM-400, 10 µl of 10 mM dNTPs, 2.5 µl RNase Inhibitor and 2.5 µl of 100 µM 10x_TSO. To this solution we added 18 µl of the first RT product and 22 µl of ddH2O. Finally, we added 5 µl Maxima H Minus Reverse Transcriptase, mixed by flicking, and centrifuged briefly. This reaction was incubated at 25ºC for 30 minutes followed by 50ºC for 90 minutes and heat inactivated at 85ºC for 5 minutes. The solution was purified using DynaBeads MyOne Silane following 10x Genomics’ instructions, beginning at “Post GEM-RT Cleanup – Silane DynaBeads” step D. The remainder of the single cell RNA-seq protocol, including purification, amplification, fragmentation, and final library amplification, followed manufacturer’s instructions.

treatment_protocol: HepG2 cells were electroporated with the following settings–pulse voltage: 1,200 V; pulse width: 50 ms; pulse number: 1.  Each replicate for electroporation was comprised of 2e6 cells. For single cell calling cards analysis of FOXA2 binding in HepG2 cells, we transfected six replicates each with 5 µg PB-SRT-Puro; three of these replicates were co-transfected with 5 µg HyPBase, while the other three were co-transfected with 5 µg FOXA2-HyPBase. All cells were allowed to recover for 24 hours before undergoing puromycin selection. A negative control replicate, transfected only with PB-SRT-Puro, was treated identically in parallel. Replicates were harvested once the negative control cells had died.

value: hepatocellular carcinoma


title: Pfur_75deg_NaCNBH3


title: Tkod_TK2097del_NaCNBH3_rep2

value: TK2097 deletion


title: Tkod_TK2097del_NaCNBH3_rep1


title: HepG2_HyPBase_scRNA


title: Tkod_55deg_NaCNBH3_rep1


title: shADORA1#3

city: Shanghai

first: Youqiong

last: Ye

address: 280 Chongqing south road

institute: Shanghai Jiao Tong University School of Medicine

data_processing:
All reads were aligned to the human reference genome (hg19) using hisat2 version 2.0.5 with the default setting.
Stringtie v1.3.3 was used to calculate the read count for each gene
Genome_build: hg19
Supplementary_files_format_and_content: Excel file included read count for each gene

growth_protocol: We collected  samples from Sk-mel-28 melanoma cell lines with stable expression of ADORA1 knockdown and scrambled-shRNA control cells

extract_protocol:
Total RNA was extracted used the TRizol reagent.
RNA libraries were prepared for sequencing using standard Illumina protocols

treatment_protocol: The shRNA sequence with the best knockout efficiency were selected. The tumor cells were stably transfected and screened by purinomycin, and then were divided into three parts when the appropriate number of cells were grown. The supernatant medium was removed and total RNA was extracted by TRizol reagent.

value: malignant melanoma cell line

tag: tissue source

value: skin

tag: genotype/variation


title: OCM1A_BAP1-HyPBase_scRNA

treatment_protocol: For single cell calling cards analysis of BAP1 binding in OCM-1A cells, we lipofected (as described above) six replicates each with 1.25 µg PB-SRT-Puro; three of these replicates were co-transfected with 1.25 µg HyPBase, while the other three were co-transfected with 1.25 µg BAP1-HyPBase. We used the TransIT-LT1 transfection reagent following manufacturer’s protocol for 6-well plates. A negative control replicate, transfected only with PB-SRT-Puro, was treated identically in parallel. Replicates were harvested once the negative control cells had died.

tag: tissue

value: uveal melanoma


title: shADORA1#2


title: OCM1A_HyPBase_scRNA


title: shADORA1#1


title: SK1_trisomic_6h

city: New York

first: Hajime

last: Murakami

state: NY

address: 1275 york ave

department: Molecular Biology

institute: Memorial Sloan Kettering Cancer Center

description: Spo11-associated DNA oligonucleotides

data_processing:
Base calls perfomed using HiSeq Control Software v.2.0.10.0 for samples sequenced on Illumina HiSeq 2500
Adapters (AGATCGGAAGAGCACACGTCTGAACTCCAGTCAC) were clipped from reads using fastx_clipper, part of the FASTX Toolkit 0.0.13.2, with parameters -l 15 -n -v -Q33 -i
First five bases (diffusion primer added in library prep) in each sequence were trimmed and trimmed sequence was recorded in the FASTQ ID tag for downstream tracking
Clipped reads were mapped using SHRiMP 2.1.1b with parameters -N 15 -U -g -1000 -q -1000 -m 10 -i -20 -h 100 -r 50% -n 1 -s 1111111111,11110111101111,1111011100100001111,1111000011001101111 -o 1001 -Q -E --sam-unaligned --strata
Sequences clipped from a read during mapping should be polyC on the 3' end or polyG on the 5' end. If clipped bases were not polyC or polyG the alignment score was corrected by deducting the mismatch score for each non-C,G base in the clipped sequence.
Maps of 5' ends of sequence reads were compiled and analyzed in R (RStudio version 0.98.1091, R version 3.0.1)
Genome_build: For wild-derived S. cerevisiae strains YPS128 and UWOPS03-461.4, reads were mapped to the SGD assembly released in June 2008, called "sacCer2" by UCSC: http://genome.ucsc.edu/cgi-bin/hgGateway?hgsid=340067645; for S. paradoxus YPS138, reads were mapped to the strain's genome assembly from SGRP (Saccharomyces Genome Resequencing Project, Wellcome Trust Sanger Institute); for S. mikatae IFO1815 and S. kudriavzevii ZP591, reads were mapped to the strains' genome assemblies from Scannell et al. 2011 G3 1: 11-25, available on http://www.saccharomycessensustricto.org.
Supplementary_files_format_and_content: tab delimited text files include Spo11-oligo hitcount at each nucleotide position in the genome

organism: Saccharomyces cerevisiae SK1

source_name: Saccharomyces cerevisiae

growth_protocol: For samples 1-8: Cells were grown in two stages of SPS (0.5% yeast extract, 2% peptone, 2% glucose) pre-growth culturing beofre transfeering into sporulation medium. For samples 9-10:  Cells were grown in YPA (1% yeast extract, 2% Bacto Peptone, 1% potassium acetate) for 13.5–14 hr at 30ºC, harvested, resuspended in 2% potassium acetate, and sporulated at 30ºC.

extract_protocol:
Based on Pan et al. 2011 Cell 144:719-731. Whole cell extracts were prepared by grinding frozen cells in a bead mill and resuspending powder in 10%TCA. Pellets were then extracted with SDS and soluble protein was subjected to affinity purification of Spo11-Flag fusion with Protein-G Dynabeads (for samples 1-8) or PrA fusion with IgG Sepharose Fast Flow beads (for samples 9-11). Protein was digested with proteinase K and Spo11-associated oligonucleotides were recovered by ethanol precipitation.
Based on Pan et al. 2011 Cell 144:719-731. Oligonucleotides were tailed with ribo-G using rGTP and terminal transferase, then ligated to duplex DNA adaptors with a dC overhang using T4 RNA ligase 2. Complementary strands were synthesized with Klenow, then purified by denaturing polyacrylamide electrophoresis. Purified complementary strands were subjected to 3' tailing with ribo-G using rGTP and terminal transferase, then another duplex adaptor was ligated using T4 RNA ligase 2. Ligated material was amplified with 16 cycles of PCR to add Illumina HiSeq adaptors. For sample 5, a spike-in oligonucleotides were added during library preparation to increase DNA amount, sequece is: TGGATCCGTGGAATAATGCACAATT

tag: epitope tag

value: epitope tag:Protein A

tag: meiotic time-point

value: meiotic time-point:6 hr in meiosis

value: meiotic Saccharomyces cerevisiae culture


title: Control#3

value: scrambled-shRNA transfected


title: Control#2


title: SK1_trisomic_4h

description: Spo11-associated DNA oligonucleotides

tag: strain

value: meiotic time-point:4 hr in meiosis


title: OCM1A_BAP1-HyPBase

data_processing:
Reads were demultiplexed by the N7 index sequences added during the final PCR. Read 1 began with the 3 base-pair barcode followed by the end of the transposon terminal repeat, culminating with the insertion site motif (TTAA in the case of piggyBac; TA in the case of Sleeping Beauty) before entering the genome. piggyBac reads were checked for exact matches to the barcode, transposon sequence, and insertion site at the beginning of reads before being hard trimmed using cutadapt (Martin, 2011) with the following settings: -g "^NNNTTTACGCAGACTATCTTTCTAGGGTTAA" --minimum-length 1 --discard-untrimmed -e 0 --no-indels, where NNN is replaced with the primer barcode. Sleeping Beauty libraries were trimmed with the following settings: -g "^NNNTAAGTGTATGTAAACTTCCGACTTCAACTGTA" --minimum-length 1 --discard-untrimmed -e 0 --no-indels. Reads passing this filter were then trimmed of any trailing Nextera adapter sequence, again using cutadapt and the following settings: -a "CTGTCTCTTATACACATCTCCGAGCCCACGAGACTNNNNNNNNNNTCTCGTATGCCGTCTTCTGCTTG" --minimum-length 1. The remaining reads were aligned to the human genome (build hg38) with NovoAlign and the following settings: -n 40 -o SAM -o SoftClip. Aligned reads were validated by confirming that they mapped adjacent to the insertion site motif. Successful reads were then converted to calling card format (.ccf.txt; see http://wiki.wubrowse.org/Calling_card) using custom programs (available at https://github.com/arnavm/calling_cards) and visualized on the WashU Epigenome Browser v46 (Zhou et al., 2011) (http://epigenomegateway.wustl.edu/legacy/).
Genome_build: hg38
Supplementary_files_format_and_content: BAM files contain aligned and annotated calling card reads. ccf.txt files contain processed calling card data.  describes calling card-specific BAM tags.

growth_protocol: Cells were cultured in RPMI 1640 Medium supplemented with 10% fetal bovine serum (FBS) and 1% of either penicillin-streptomycin or antibiotic-antimycotic. Cells were grown at 37ºC with 5% carbon dioxide (CO2). Media was replenished every 2 days.

treatment_protocol: We transfected 10 replicates of OCM-1A cells 1.25 µg of PB-SRT-Puro and 1.25 µg of either HyPBase or BAP1-HyPBase (gifts from Dr. Michael Onken) using the TransIT-LT1 transfection reagent following manufacturer’s protocol for 6-well plates. Puromycin was added after 24 hours to a final concentration of 2 µg/ml. Cells were grown under selection for one week, by which time almost all negative control transfectants were dead. After 7 days, we dissociated each replicate with trypsin-EDTA and created single cell suspensions in phosphate-buffered saline (PBS) Aliquots of each replicate were cryopreserved in cell culture media (see above) supplemented with 5% DMSO. The remaining cells were pelleted by centrifugation at 300g for 5 minutes. Cell pellets were either processed immediately or kept at -80ºC in RNAProtect Cell Reagent.


title: K562_SP1-HyPBase_scRNA

treatment_protocol: K562 cells were transfected using Neon electroporation with the aforementioned settings. Each replicate for electroporation was comprised of 2e6 cells. For single cell calling cards analysis of SP1 binding in K562 cells, we transfected four replicates each with 5 µg PB-SRT-Puro and 5 µg SP1-HyPBase. These libraries were not mixed. We used the demultiplexed data from the cell line mixing experiment with HyPBase as controls. Cells were cultured independently and mixed immediately prior to generating single cell emulsions. All cells were allowed to recover for 24 hours before undergoing puromycin selection. A negative control replicate, transfected only with PB-SRT-Puro, was treated identically in parallel. Replicates were harvested once the negative control cells had died.

value: leukemia


title: SK1_trisomic_5h

description: Spo11-associated DNA oligonucleotides

value: meiotic time-point:5 hr in meiosis


title: SK1_homeologous_5h

description: Spo11-associated DNA oligonucleotides

source_name: Saccharomyces cerevisiae plus Saccharomyces pastorianus chromsome V

value: epitope tag:Spo11-Flag


title: 12_BAFA_674

city: Paris

first: Franck

last: Letourneur

address: 22 rue mechain

institute: INSERM

scan_protocol: After overnight hybridization, chips are washed on the fluidic station FS450 following specific protocols (Affymetrix) and scanned using the GCS3000 7G

data_processing: The scanned images are then analyzed with Expression Console software (Affymetrix) to obtain raw data (cel files) and metrics for Quality Controls. The observations of some of these metrics and the study of the distribution of raw data show no outlier experiment. RMA normalization is performed using R with Version 21 of Entrezgene CDF brain array for ClariomD Mouse array.

label: Biotin

source_name: Healthy liver tissue from WT mouse

label_protocol: 80 ng of total RNA is reverse transcribed following the Ovation FFPE WTA System (Tecan). Briefly, the resulting double strand cDNA is used for amplification based on SPIA technology. After purification according to Nugen protocol, 5.5 ug of Sens Target DNA are fragmented and biotin labelled using Encore Biotin Module kit (Nugen). After control of fragmentation using Bioanalyzer 2100, cDNA is then hybridized to GeneChip® MTA1.0 ST (Affymetrix) at 45°C for 17 hours

extract_protocol: After deparaffinization, FFPE tissues were lysed for 24h in tissue lysis buffer  supplemented with proteinase K at 60C and total RNA were extracted with trizol reagent.

treatment_protocol: Liver tissues were fixed overnight in 4% formalin buffer and embedded in paraffin

value: liver

tag: tissue region

value: healthy liver

value: wildtype


title: Lib10AA_ADAM10_R1

tag: fragment size

tag: sample prep

value: phage_PCR and gel extraction


title: 11_BAFA_573

hyb_protocol: After control of fragmentation using Bioanalyzer 2100, cDNA is then hybridized to GeneChip® MTA1.0 ST (Affymetrix) at 45°C for 17 hours


title: Control#1

tag: cell type


title: 10_BAFA_670


title: SK1_homeologous_4h

description: Spo11-associated DNA oligonucleotides


title: HCT-116_N2a_HyPBase_scRNA

description: Single cell RNA-seq library

data_processing:
scRNA-seq libraries were sequenced on either Illumina HiSeq 2500 or NovaSeq S1 machines. Reads were analyzed using 10x Genomics’ Cell Ranger with the following settings: --expect-cells=6000 --chemistry=SC3Pv2 --localcores=16 --localmem=30. The digital gene expression matrices from 10x were then further processed with scanpy (Wolf et al., 2018) for identification of highly variable genes, batch correction, dimensionality reduction, and Louvain clustering. Processed scRNA-seq datasets were stored as .loom files (http://loompy.org). We cross-referenced gene expression data with published datasets (Rosenberg et al., 2018; Rouillard et al., 2016; Saunders et al., 2018; Tasic et al., 2018; Zeisel et al., 2018) to assign cell types. The species mixing analysis was performed using Drop-seq_tools (Macosko et al., 2015)
Genome_build: hg38 & mm10
Supplementary_files_format_and_content: Loom files contain processed scRNA-seq data

organism:
Homo sapiens
Mus musculus

treatment_protocol: HCT-116 cells were transfected using Neon electroporation with the aforementioned settings. N2a cells were electroporated with the following settings–pulse voltage: 1,050 V; pulse width: 30 ms; pulse number: 2. Each replicate for electroporation was comprised of 2e6 cells. For the species mixing experiment, we transfected one replicate each of HCT-116 and N2a cells with 5 µg PB-SRT-Puro and 5 µg HyPBase. All cells were allowed to recover for 24 hours before undergoing puromycin selection. A negative control replicate, transfected only with PB-SRT-Puro, was treated identically in parallel. Replicates were harvested once the negative control cells had died.

value: mixed species


title: OCM1A_HyPBase


title: Tam4_85deg_mock

organism: Thermococcus sp. AM4

tag: genetic background

value: WT

tag: treatment

value: mock

tag: growth condition

value: 85 degrees


title: Treatment_Liver 2

city: 无锡

first: 李

middle: 鸣

last: 霄

address: 滨湖区雪浪街道南泉薛家里69号无锡渔业学院

institute: 中国水产科学研究院淡水渔业研究中心

data_processing:
Illumina Casava1.7 software used for basecalling.
Sequenced reads were trimmed for adaptor sequence, and masked for low-complexity or low-quality sequence, then mapped to mm8 whole genome using bowtie v0.12.2 with parameters -q -p 4 -e 100 -y -a -m 10 --best --strata
Reads Per Kilobase of exon per Megabase of library size (FPKM) were calculated using a protocol from Chepelev et al., Nucleic Acids Research, 2009. In short, exons from all isoforms of a gene were merged to create one meta-transcript. The number of reads falling in the exons of this meta-transcript were counted and normalized by the size of the meta-transcript and by the size of the library.
Data filtering steps.Prior to assembly, the low quality reads(1,reads containing sequencing adaptors; 2,reads containing sequencing primer;3, nucleotide with q quality score lower than 20 )were removed .software CutAdapter,parameters -o 5 -p 100
Read alignment software,software HISAT ,version 2.0 ,parameters -l fr-firststrand -mi 20 -mx 500000 -p 2 -b dta -q phred33-quals -x 9
perform expression level for mRNAs by calculating FPKM.stimate the expression levels of all transcripts.software StringTie ,version 1.3.0,parameters -b dta -q phred33-quals.
Genome_build: ASM372403v1
Supplementary_files_format_and_content: tab-delimited text files include FPKM values for each Sample

organism: Tachysurus fulvidraco x Tachysurus vachelli

source_name: Liver

extract_protocol:
Total RNA was extracted using Trizol reagent (Invitrogen, CA, USA) following the manufacturer's procedure. The total RNA quantity and purity were analysis of Bioanalyzer 2100 and RNA 6000 Nano LabChip Kit (Agilent, CA, USA) with RIN number >7.0. Approximately 10 ug of total RNA representing a specific adipose type was subjected to isolate Poly (A) mRNA with poly-T oligoattached magnetic beads (Invitrogen). Following purification, the poly(A)- or poly(A)+ RNA fractions is fragmented into small pieces using divalent cations under elevated temperature. Then the cleaved RNA fragments were reverse-transcribed to create the final cDNA library in accordance with the protocol for the mRNA-Seq sample preparation kit (Illumina, San Diego, USA), the average insert size for the paired-end libraries was 300 bp (±50 bp). And then we performed the paired-end sequencing on an Illumina Novaseq™ 6000 at the (lc-bio, China) following the vendor's recommended protocol.
RNA libraries were prepared for sequencing using standard Illumina protocols

value: Liver

tag: age

value: Feeding for 8 weeks

value: 4g/kg Acanthopanax  senticosus supplementation in the diet


title: Lib_hP_ADAM17_R1


title: Treatment_Liver 3

address: 滨湖区雪浪街道南泉薛家里69号无锡渔业学院

institute: 中国水产科学研究院淡水渔业研究中心


title: SK1_zip3Δ homeologous_2

description: Spo11-associated DNA oligonucleotides

value: meiotic Saccharomyces cerevisiae culture


title: 09_BAFA_669

tag: genotype


title: SK1_ecm11Δ_2

description: Spo11-associated DNA oligonucleotides


title: 08_BAFA_562_KO

source_name: RBC region from  [Apc-Arid1a]ko-focal liver mouse

value: RBC region


title: 06_BAFA_263_KO


title: 07_BAFA_562_WT

source_name: Adjacent region from  [Apc-Arid1a]ko-focal liver mouse

value: Adjacent region


title: SK1_zip3Δ homeologous_1

description: Spo11-associated DNA oligonucleotides


title: Control_Liver 3

address: 滨湖区雪浪街道南泉薛家里69号无锡渔业学院

institute: 中国水产科学研究院淡水渔业研究中心

value: 0g/kg Acanthopanax  senticosus supplementation in the diet


title: Tam4_75deg_mock

value: 75 degrees


title: 05_BAFA_263_WT


title: Tam4_75deg_NaCNBH3


title: Control_Liver 2

address: 滨湖区雪浪街道南泉薛家里69号无锡渔业学院

institute: 中国水产科学研究院淡水渔业研究中心


title: SK1_zip3Δ_2

description: Spo11-associated DNA oligonucleotides

value: epitope tag:Spo11-Flag

value: meiotic time-point:5 hr in meiosis


title: SK1_zip3Δ_1

description: Spo11-associated DNA oligonucleotides


title: 03_BAFA_239_WT

value: liver


title: Tam4_85deg_NaCNBH3


title: Control_Liver 1

address: 滨湖区雪浪街道南泉薛家里69号无锡渔业学院

institute: 中国水产科学研究院淡水渔业研究中心


title: Tam4_65deg_mock

value: 65 degrees


title: 02_BAFA_114GR_KO


title: myrAKT#4

status: Public on Jul 22 2020

city: Hangzhou

first: Ting

last: Tao

state: Zhejiang

address: 3333 Binsheng Road

institute: Children's Hospital, Zhejiang University School of Medicine

data_processing:
After acquiring the raw fastq format paired-end RNA-seq reads, we inspected the sequencing quality using fastqc v0.11.5, and removed standard Illumina adapters from the 3’ end using Trim Galore v0.4.4.
The processed reads were then aligned to the GRCz10 version of the zebrafish reference genome using Tophat v2.1.1. The GTF format UCSC gene models, obtained from the iGenomes website, was provided as an annotation reference (-G option of Tophat).
After the alignment, the BAM format files were sorted by reads name using Samtools v1.5, and the reads mapped to each gene were counted using HTSeq-count v0.6.1.
edgeR library was applied to normalize read counts across samples and identify genes that are differentially expressed between conditions in R.
Genome_build: GRCz10
Supplementary_files_format_and_content: counts

source_name: Zebrafish ganglioneuroma tumor cells

growth_protocol: EGFP+ neuroblastoma cells or mCherry+ ganglioneuroma cells were harvested from tumor-bearing MYCN or myrAkt transgenic zebrafish. Tumors were mechanically homogenized and EGFP+ or mCherry+ cells were sorted directly into Trizol-LS

extract_protocol:
EGFP+ or mCherry+ tumor cells were sorted, then total RNA was extracted with Trizol-LS.
Total RNA was depleted of ribosomal RNA with the SMARTer Universal Low Input RNA Kit (Clontech). Enriched mRNA was then applied to library preparation according to manufacturer’s protocol (NEBNext Ultra). After repeated quality control for average DNA input size of 300 bp, the samples were sequenced on aIllumina NextSeq 500 with 2X75bp paired-end reads.

value: myrAKT transgenic fish

value: ganglioneuroma cells


title: 04_BAFA_239_KO


title: Tam4_65deg_NaCNBH3

value: WT


title: myrAKT#1


title: 01_BAFA_114S_WT


title: Treatment_Liver 1

address: 滨湖区雪浪街道南泉薛家里69号无锡渔业学院

institute: 中国水产科学研究院淡水渔业研究中心


title: MYCN#3

source_name: Zebrafish neuroblastoma tumor cells

value: MYCN transgenic fish

value: neuroblastoma cells


title: MYCN#2


title: myrAKT#3


title: IL-17A-/- Knockout CD4+CD62L+ T cells replicate 3

first: Matthew

middle: J

last: Brooks

address: 6 Center Dr Bldg 6, Rm 303

department: NEI

institute: NIH

description:
KO.3
replicate 3

data_processing:
Reads passing Illumina chastity filtering were aligned to reference assembly GRCm38 with transcript annotation Ensembl v73. Transcript quantitation was accomplished by streaming read alignment from Bowtie2 v2.1.0 (Langmead and Salzberg, 2012) to eXpress v1.3.1 (Roberts and Pachter, 2013; Roberts et al., 2013). Bowtie2 alignment settings were used as previously described (Roberts et al., 2013) and eXpress setting –r-stranded was used for directional transcript analysis. Bias corrected effective counts were used for downstream analysis.
Genome_build: GRCm38 assembly, Ensembl v73 annotation
Supplementary_files_format_and_content: tab-delimited text files of bias corrected count values.

source_name: CD4+CD62L+ T cells

extract_protocol:
Splenocytes were isolated, ACK lysed, cell population was selected using CD4+CD62L+ T Cell Isolation Kit II (Miltenyi Biotech, Cambridge, MA)
RNA was isolated using the RNeasy Mini Kit (Qiagen) and processed as per manufacturer’s instructions.
Stranded libraries were prepared using a modified Illumina TruSeq mRNA protocol (Brooks, et al. Meth Mol Biol 2012)

treatment_protocol: CD4+CD62L+ cells were in-vitro cultured for three days under Th17 polarization condition. Culture medium was supplemented with 10 ng/ml IL-6, 10 ng/ml IL-23, 1 ng/ml TGF-β, 10 μg/ml anti-IFN-γ and 10 μg/ml anti-IL-4 antibodies

value: CD4+CD62L+ T cells

tag: background strain

value: IL-17A-/- Knockout

value: Th17 polarized


title: IL-17A-/- Knockout CD4+CD62L+ T cells replicate 1

description:
KO.1
replicate 1


title: Treg-psoriasis-3

city: Xi'an

first: luting

last: Yang

state: Shaanxi

address: No.620, South Chang’an Road, Xi’an, 710062, China

institute: Shaanxi normal university

description: circRNA expression

hyb_protocol: 1 μg of each labeled cRNA was fragmented by adding 5 μl 10 × Blocking Agent and 1 μl of 25 × Fragmentation Buffer, then heated the mixture at 60 °C for 30 min, finally 25 μl 2 × Hybridization buffer was added to dilute the labeled cRNA. 50 μl of hybridization solution was dispensed into the gasket slide and assembled to the circRNA expression microarray slide. The slides were incubated for 17 hours at 65°C in an Agilent Hybridization Oven. The hybridized arrays were washed, fixed and scanned using the Agilent Scanner G2505C.

scan_protocol: the arrays were scanned by the Agilent Scanner G2505C

data_processing:
data processing were performed using the R software package. After quantile normalization of the raw data, low intensity filtering was performed, and the circRNAs that at least 3 out of 6 samples have flags in “P” or “M” (“All Targets Value”) were retained for further analyses.
circRNAs having fold changes ³1.5 and p-values £ 0.05 are selected as the significantly differentially expressed.

source_name: Peripheral blood, Treg,psoriatic patients

label_protocol: the enriched circular RNAs were amplified and transcribed into fluorescent cRNA utilizing a random priming method (Arraystar Super RNA Labeling Kit; Arraystar). The labeled cRNAs were purified by RNeasy Mini Kit (Qiagen).

extract_protocol: Total RNA was extracted from Tregs using TRIzol reagent (Invitrogen, CA, USA).

treatment_protocol: Tregs are isolated by flow cytometry from PBMCs

tag: gender

value: male

tag: pasi score


title: Treg-normal control-3

description: circRNA expression

source_name: Peripheral blood, Treg,normal control


title: Treg-psoriasis-2

description: circRNA expression

value: female


title: Treg-normal control-2

description: circRNA expression


title: Treg-psoriasis-1

description: circRNA expression


title: myrAKT#2

tag: genotype/variation


title: MYCN#4


title: IL-17A-/- Knockout CD4+CD62L+ T cells replicate 2

description:
KO.2
replicate 2


title: Treg-normal control-1

description: circRNA expression


title: IL-17A+/+ wild-type CD4+CD62L+ T cells replicate 3

description:
WT.3
replicate 3

value: IL-17A+/+ wild-type


title: patch1

city: Cambridge

first: Joshua

last: Levin

address: 75 Ames St

department: Stanley Center

institute: Broad Institute

description:
nuclear RNA
TPM.patch.tsv
patchseq_170106_s1_S385_L001

data_processing:
RSEM v1.2.87 was run on raw fastq fileswith default parameters on alignments created by Bowtie on UCSC gene annotation
TPM estimates were tranformed to log space by taking log TPM +1
TopHat2 v2.0.10 was used to map fastq files to mm10 reference
RNASeq-QC was run on the TopHat2 bamfiles to get QC metrics
For TRN and patch-seq samples, cells were removed if they had less than 40% genome mapping rate or or less than 1500 detected genes
Clustering and DE analysis of TRN samples was perfomed with biSNE software (https://github.com/yinqingl/nucseq_analysis), including normalizing the log TPM data to a normalized log TPM
For non-TRN samples, nuclei expressing <1500 genes were removed
Seurat v2.2.0 was used for clustering of the non-TRN samples
Genome_build: mm10
Supplementary_files_format_and_content: Formatted expression matrix (tab deliminated text file) with cells as columns, genes as rows, and TPM values as entries

source_name: TRN

extract_protocol:
Sucrose gradient nuclei isolation followed by FACS sorting for Violet+, Violet+ GFP+ or Violet+ NeuN+ nuclei
Modified SMARTseq2 single nuclei RNA-Seq

tag: sample type

value: sNuc-seq

tag: used for patch-seq

value: Yes

tag: facs sorting signal

tag: plate type

value: Patch-Seq


title: patch3

description:
nuclear RNA
TPM.patch.tsv
patchseq_170106_s3_S409_L001


title: patch7

description:
nuclear RNA
TPM.patch.tsv
patchseq_170106_s7_S457_L001


title: patch2

description:
nuclear RNA
TPM.patch.tsv
patchseq_170106_s2_S397_L001


title: MYCN#1


title: IL-17A+/+ wild-type CD4+CD62L+ T cells replicate 2

description:
WT.2
replicate 2


title: IL-17A+/+ wild-type CD4+CD62L+ T cells replicate 1

description:
WT.1
replicate 1


title: siCHD7-2

city: ChengDu,SiChuan

first: Caojie

last: Liu

address: No.14,3Rd Section Of Ren Min Nan Rd.

data_processing:
Illumina Casava1.7 software used for basecalling.
Sequenced reads were trimmed for adaptor sequence, and masked for low-complexity or low-quality sequence, then mapped to mm8 whole genome using bowtie v0.12.2 with parameters -q -p 4 -e 100 -y -a -m 10 --best --strata
Reads Per Kilobase of exon per Megabase of library size (RPKM) were calculated using a protocol from Chepelev et al., Nucleic Acids Research, 2009. In short, exons from all isoforms of a gene were merged to create one meta-transcript. The number of reads falling in the exons of this meta-transcript were counted and normalized by the size of the meta-transcript and by the size of the library.
Genome_build: mm9
Supplementary_files_format_and_content: Matrix table with raw gene counts for every gene and every sample

source_name: human dental follicle cells

extract_protocol:
Human dental follicle was obtained from the unerupted third molar with undeveloped root. Patients aged 12-16 years undergoing the third molar extraction in West China Hospital of Stomatology, Sichuan University would meet the inclusion criteria. Patients with history of systemic disease, or undergoing maxillofacial surgery, whole body or partial radiotherapy, chemotherapy, periodontitis, oral mucosal disease or smoking were excluded. All the procedures are approved by Institutional Review Board and the informed consent of patients. After extraction, the dental follicle was instantly immersed into the phosphate buffer saline (PBS, Gibco) with 1× Penicillin-Streptomycin (Liquid, Gibco), i.e. PBS with 100 units/mL penicillin and 100 µg/mL streptomycin. The dental follicle was cut into size of 1mm3 pieces,  and digested with PBS with Type I Collagenase (3mg/ml, Gibco) and Dispase (3 mg/ml, Gibco) for 1 hour in 37℃ water bath with agitation.  The digested tissue suspension was cultured in 21cm2 petri dish in alpha minimum Eagle’s medium (α-MEM, HyClone) with 20% fetal bovine serum (FBS, Gibco) and 1× Penicillin-Streptomycin. Incubated at 37℃ with 5% CO2, the culture medium was changed every 2 days. When cell confluence rate reached 80%, hDFCs were passaged and subcultured in α-MEM with 10% FBS and 1× Penicillin-Streptomycin. hDFCs at passage 3 was applied for the subsequent research.  For osteogenic induction, hDFCs were cultured with osteogenic medium supplemented with 50 μM ascorbic acid (Sigma), 10 nM dexamethasone (Sigma) and 10 mM β-glycerophosphate (Sigma).
RNA libiaries were prepared for sequencing using standard Illumina protocols.

tag: strain

value: Home sapiens

tag: tissue

value: dental follicle

value: 16 years old


title: siCHD7-1

institute: West China Hospital of Stomatology, Sichuan University


title: patch6

description:
nuclear RNA
TPM.patch.tsv
patchseq_170106_s6_S445_L001


title: patch5

description:
nuclear RNA
TPM.patch.tsv
patchseq_170106_s5_S433_L001


title: patch4

description:
nuclear RNA
TPM.patch.tsv
patchseq_170106_s4_S421_L001


title: siCTRL-3


title: siCTRL-2


title: siCHD7-3


title: siCTRL-1