title: Liver of Citrobacter rodentium infected mouse 3 (liver CFU positive)

description:
Cr_pos3
RawCounts_Cr.txt
Liver of Citrobacter rodentium infected mouse 1 (liver CFU positive)
Library 9

value: Citrobacter rodentium infected mouse (liver CFU positive)


status: Public on May 02 2023

type: SRA

city: Boston

first: Ting

last: Zhang

address: 181 longwood Ave

country: USA

institute: BWH

description:
Cr_pos1
RawCounts_Cr.txt
Liver of Citrobacter rodentium infected mouse 1 (liver CFU positive)
Library 7

library_source: transcriptomic

data_processing:
For data analysis, reads were trimmed using Trim Galore with automatic adapter sequence detection.
Then, trimmed reads were mapped to the mouse reference genome (mm10) and annotated using STAR v2.7.3a with default parameter.
The number of mapped reads to each gene were counted by featureCounts of the Subread package using mouse GENCODE annotation M25 (GRCm38.p6).
Assembly: mm10
Supplementary files format and content: Feature counts table

molecule: total RNA

organism: Mus musculus

source_name: liver

extract_protocol:
RNAs were extracted from the liver samples using Trizol (Thermo Fisher).
RNA-seq libraries were prepared by KAPA RNA Hyperprep Kit (Roche).

tag: tissue

value: liver

tag: treatment

tag: strain


title: 180822_P1-180822_P2-D10-AD002

city: La Jolla

first: Joseph

middle: R

last: Ecker

state: CA

address: 10010 North Torrey Pines Road

department: Genomic Analysis Laboratory

institute: HHMI-Salk-Institute

library_source: genomic

data_processing:
Reads trimed with Cutadapt for adapters and low quality bases (Phred score >= 20)
The resulting files are mapped to in-silico bisulfite converted reference genome using Bismark.
The raw BAM file will be filtered using samtools to filter reads by MAPQ > 10 and use Picard to remove PCR duplicates
Tab-delimited files (ALLC files) describing the quantity of unmethylated and methylated cytosine base calls will be generated with in-house developing package using the filtered BAM files
Assembly: mm10
Supplementary files format and content: tab-delimited files (ALLC files) describing the quantity of unmethylated and methylated cytosine base calls. Columns are: 1. Chromosome, 2. Position, 3. Strand, 4. Cytosin Context, 5. methylated base call, 6. total mapped base call, 7. not used, place holder column
Library strategy: snmC-seq2 & 3

molecule: genomic DNA

source_name: brain

growth_protocol: All experimental procedures using live animals were approved by the Salk Institute Animal Care and Use Committee. The knock-in mouse line, R26R-CAG-loxp-stop-loxp-Sun1-sfGFP-Myc (INTACT) was used for most experiments4 and they were maintained on a C57BL/6J background. 42-49 day old adult male and female INTACT mice were used for the retrograde labeling experiment.

extract_protocol:
Approximately two weeks after the AAVretro injection, brains were extracted from the 56-63 day old INTACT mice, immediately submerged in ice-cold slicing buffer (2.5mM KCl, 0.5mM CaCl2, 7mM MgCl2, 1.25mM NaH2PO4, 110mM sucrose, 10mM glucose and 25mM NaHCO3) that was bubbled with carbogen, and sliced into 0.6 mm coronal sections starting from the frontal pole. From each AAVretro-injected brain, the slices were kept in the ice-cold dissection buffer from which selected brain regions (Supplementary Table 1) were manually dissected under a fluorescent dissecting microscope (Olympus SZX16), following the Allen Mouse Common Coordinate Framework (CCF), Reference Atlas, Version 3 (2015) (Extended Data Fig. 1). The dissected brain tissues were transferred to prelabeled microcentrifuge tubes, immediately frozen in dry ice, and subsequently stored at -80oC.   For each dissected brain region, samples from 2 males and 2 females were pooled separately as biological replicates for nuclei preparation. The 2-mL glass tissue dounce homogenizer and pestles (Sigma-Aldrich D8938-1SET) were pre-chilled on ice. Nuclei were prepared using a modified protocol as reported by Lacar et al., 201622. In summary, the frozen brain tissues were transferred to the dounce homogenizer with 1 mL ice-cold NIM buffer (0.25M sucrose, 25mM KCl, 5mM MgCl2, 10mM Tris-HCl (pH7.4), 1mM DTT (Sigma 646563), 10μl of protease inhibitor (Sigma P8340)), with 0.1% Triton X-100 and 5μM Hoechst 33342 (Invitrogen H3570), and gently homogenized on ice with the pestle 10-15 times. The homogenate was transferred to pre-chilled microcentrifuge tubes and centrifuged at 1000 rcf for 8 min at 4oC to pellet the nuclei. The pellet was resuspended in 1 mL ice-cold NIM buffer, and again centrifuged at 1000 rcf for 8 min at 4oC. The pellet was then resuspended in 450 μL of ice-cold NSB buffer (0.25M sucrose, 5mM MgCl2, 10mM Tris-HCl (pH7.4), 1mM DTT, 9ul of Protease inhibitor), and filtered through 40μM cell strainer. The filtered nuclei suspension was incubated on ice for at least 30 minutes with 50μl of nuclease-free BSA for at least 10 minutes, then incubated with GFP antibody, Alexa Fluor 488 (Invitrogen, A-21311) and anti-NeuN antibody (EMD Millipore MAB377) conjugated with Alexa Fluor 647 (Invitrogen A20173). GFP+/NeuN+ single nuclei were isolated using fluorescence-activated nuclei sorting (FANS) on a BD Influx sorter with 100μm nozzle, and sorted into 384-well plates preloaded with 2μl of digestion buffer for snmC-seq (20 mL digestion buffer consists of 10 mL M-digestion buffer (2×, Zymo D5021-9), 1 ml Proteinase K (20 mg, Zymo D3001-2-20), 9 mL water, and 10 µL unmethylated lambda DNA (100 pg/µL, Promega, D1521)). The collected plates were incubated at 50oC for 20 minutes then stored at -20 oC.
The bisulfite conversion and library preparation were performed following the detailed snmC-seq2 protocol as previously described (Luo, C. et al., Nat. Commun. 2018, 9(1):3824.) or snmC-seq3 protocol described in (Liu, H. et al., Biorxiv. 2023:2023-04.). The libraries were sequenced on Illumina Novaseq 6000 using the S4 flow cell 2 x 150 bp mode.

treatment_protocol: To label neurons projecting to regions of interest, injections of rAAV2-retro-Cre (produced by Salk Vector Core or Vigene, 2x1012 to 1x1013 viral genomes/ml, produced with capsid from Addgene plasmid #81070 packaging pAAV-EF1a-Cre from Addgene plasmid #55636) were made into both hemispheres of the INTACT mice. Animals were anesthetized with either ketamine/xylazine or isoflurane, placed in a stereotaxic frame, and 0.1 to 0.5 microliters of AAV was injected by pressure into stereotaxic coordinates corresponding to the desired projection target.

value: brain

tag: source

value: THl

tag: target

value: SSp

tag: Sex

value: female


title: 180822_P1-180822_P2-D10-AD001

value: male


title: Liver of Citrobacter rodentium infected mouse 2 (liver CFU positive)

description:
Cr_pos2
RawCounts_Cr.txt
Liver of Citrobacter rodentium infected mouse 1 (liver CFU positive)
Library 8


title: Cecum adult

city: Hangzhou

first: Jiajin

last: Wu

state: Zhejiang

address: Yuhangtang road

country: China

institute: Zhejiang University

data_processing:
Sequences from NovaSeq 6000 system were  de-multiplexed and aligned using CellRanger version 3.1.0, available from 10x Genomics with default parameters.
Genome_build: ARS-USD1.2 cattle reference genome
Supplementary_files_format_and_content: Tab-delimited text files include barcodes, gene and count matrix for each Sample

organism: Bos taurus

source_name: Cecum

extract_protocol:
After removed the outer muscle layers, the adult (AD) duodenum, jejunum, cecum, and colon samples were chopped into 1 mm pieces and treated with different enzymes for different durations：the duodenum and cecum samples were dissociated using the Lamina Propria Dissociation Kit (Miltenyi biotec, Bergisch Gladbach, Germany) for 30 min at 37℃; the jejunum and colon samples were dissociated using the Multi Tissue Dissociation Kit 2 (Miltenyi biotec, Bergisch Gladbach, Germany) for 41 min at 37℃. Then, the digestion was stopped by adding 10% of FBS, followed by a filtration step with 40 μm sieve. Next, after centrifuged at 300 × g for 5 min at 4°C and washed with 1× DPBS containing 2% FBS and 0.2 mM EDTA, the samples were treated with Red Blood Cell Lysis Solution (Miltenyi Biotech, Bergisch Gladbach, Germany) for 2 min. The samples were washed with 1× DPBS containing 2% FBS and 0.2 mM EDTA, centrifuged at 300 × g for 5 min at 4°C, and re-suspended in 1× DPBS containing 10% FBS and 0.2 mM EDTA.  For the NB reticulum, omasum, abomasum, duodenum, jejunum, ileum, cecum, colon, and rectum samples, we firstly removed the outer muscle layers of samples, and then minced samples into 10 × 0.5 mm2 pieces. The samples were incubated with 20 mM EDTA for 30 min at 37℃, and then rinsed with DPBS and chopped into 1 mm pieces. Tissue pieces were incubated in a 37℃ water bath for 5 min with 0.25% Trypsin-EDTA (Gibco) and then the centrifuge tube containing tissue pieces was inserted into ice for 2 min, and the digestion was stopped by adding the pre-chilled HBSS. The supernatant was discarded after centrifuged at 300 × g for 2 min at 4℃, and samples were washed twice with cold HBSS. Samples were incubated with different dissociation enzymes for different durations. For NB reticulum, omasum, and abomasum samples, the multiple enzymes (1.5 mg/ml collagenase I, 1.5 mg/ml collagenase IV, 100 U/ml hyaluronidase, 1.5 mg/ml dispase, and 50 U/ml DNase I) were added and treated for 30 min at 37℃. For the NB duodenum, jejunum, ileum, cecum, colon, and rectum samples, the multiple enzymes in the Propria Dissociation Kit (Miltenyi biotec, Bergisch Gladbach, Germany) were added and treated for 30 min at 37℃. The 10% FBS was added to stop the digestion, followed by a filtration step through 70 μm and 30 μm SmartStrainer (Miltenyi biotec, Bergisch Gladbach, Germany). After centrifuged at 300 × g for 5 min at 4℃, samples were then re-suspended in 2 mL of HBSS. Then, samples were centrifuged at 300 × g for 5 min at 4℃ and washed twice with 1×PBS with 0.04% BSA. Finally, samples were centrifuged at 300 × g for 5 min at 4℃ and re-suspended in 1×PBS with 0.04% BSA.   For the NB liver sample, the isolated sample was transferred into cold DPBS, minced into 1mm pieces, and then incubated with 1.5mg/ml Collagenase I, 20U/ml papain, and 50U/ml DNase I for 20 min at 37℃. The dissociated cells were then passed through 70 μm and 30 μm SmartStrainer, centrifuged at 300 × g for 5 min at 4℃, and then re-suspended in 2 mL of cold HBSS. After centrifuged at 300 × g for 5 min, dissociated cells were washed twice with 1×PBS with 0.04% BSA at 4℃ and then re-suspended in 1×PBS with 0.04% BSA.  For the NB salivary gland sample, the isolated sample from parotid gland was transferred into cold 1×PBS. The blood and fat of the sample were removed, and then the sample was chopped into 0.5 mm pieces. After incubated with 1.5mg/ml collagenase II, 1.5mg/ml dispase, and 50U/ml DNase I for 20 min at 37℃, the sample was passed through 70 μm and 30 μm SmartStrainers, and then centrifuged at 300 × g for 5 min at 4℃. Dissociated cells were re-suspended in 300 μL of RPMI 1640 medium (Gibco), and 3mL of Red Blood Cell Lysis Solution wad added for 3 min. Dissociated cells were then centrifuged at 300 × g for 5 min at 4℃ and re-suspended in 2 mL of RPMI 1640 medium. After centrifuged at 300 × g for 5 min, dissociated cells were washed twice with 1×PBS with 0.04% BSA at 4℃ and then re-suspended in 1×PBS with 0.04% BSA.
The single-cell RNA-seq library preparation was performed using the Chromium Single Cell 3' Reagent Kits v3.

tag: breed

value: Holstein dairy cows

value: cecum

tag: developmental stage

value: adult


title: Duodenum adult

source_name: Duodenum

value: duodenum


description:
Cr_neg2
RawCounts_Cr.txt
Liver of Citrobacter rodentium infected mouse 2 (liver CFU negative)
Library 5

value: Citrobacter rodentium infected mouse (liver CFU negative)


title: Twi_ATAC_S2-WT_40uM_rep1

city: Madison

first: Melissa

middle: M

last: Harrison

email: mharrison3@wisc.edu, ykaur@wisc.edu, torreszelada@wisc.edu, mmfreund2@wisc.edu

state: WI

address: 420 Henry Mall

department: Biomolecular Chemistry

institute: University of Wisconsin Madison

description:
Twi_ATAC_S2-WT_40uM_small.bw
S2-Twi_ATACseq_peaks.narrowPeak

data_processing:
Raw ATAC-seq reads were trimmed to remove adapter sequences using NGmerge version 0.3.
Trimmed reads were aligned to the Drosophila melanogaster genome (version dm6) using bowtie2 with paramters -k 2 --very-sensitive --no-mixed --no-discordant -X 5000.
Reads with a mapping quality < 30 and reads aligning to the mitochondrial genome or scaffolds were discarded. Only fragments < 100bp were considered for downstream analysis.
Reads were combined across all replicates and peak calling was performed using MACS2 with parameters -f BAMPE --keep-dup all -g dm --call-summits.
featureCounts was used to count the number of reads aligning within 200bp (100bp upstream or downstream) of peak summits.
Differential accessibility analysis was performed using DESeq2. Regions with an adjusted p-value < 0.05 were considered differentially accessible.
bigWig files were generated using deepTools bamCoverage version 3.5.1 with parameters –binSize 10. bigWig files were z-score normalized by subtracting the genome-wide average of all bins from each bin value and dividing by the standard deviation of all bins.
Assembly: dm6
Supplementary files format and content: bigWig files: z-score normalized read depth for merged replicates. Only small fragments (< 100bp) are included.
Supplementary files format and content: narrowPeak files: MACS2 peaks called on combined replicates for all samples within each experiment.

organism: Drosophila melanogaster

growth_protocol: Drosophila S2 cells were cultured at 27°C in Schneider's medium supplemented with 10% fetal bovine serum  and antibiotic/antimycotic.

extract_protocol:
2×10^5 cells were harvested for ATAC-seq by centrifugation at 600 × g for 3 minutes at RT. Cells were washed once in 100 µL 1X PBS and resuspended in 100 µL ATAC lysis buffer (10 mM Tris pH 7.5, 10 mM NaCl, 3 mM MgCl2,0.1% NP-40). Cells were centrifuged at 600 × g for 10 minutes at 4°C. Supernatant was removed and pellet was resuspended in 47.5 µL buffer TD (Illumina). 2.5 uL of Tagment DNA Enzyme (Illumina) was added and samples were incubated at 37°C for 30 minutes. Tagmented DNA was purified using MinElute Reaction Cleanup Kit (Qiagen) and eluted in 10 µL of buffer EB.
DNA was PCR amplified for 12 cycles with the following conditions:72°C for 5 minutes, 98°C for 30 seconds, then 12 cycles of 98°C for 10 seconds, 63°C for 30 seconds and 72°C for 1 minute. Amplified libraries were purified using a 1.2X ratio of Axygen paramagnetic beads.

treatment_protocol: Transcription factor expression was induced by adding CuSO4 to the cell culture media. For all induction experiments, cells were plated at 1×10^6 cells per mL. Experiments were performed with cells harvested 48 hours after induction.

tag: cell line

tag: genotype

value: 40uM CuSO4


description:
Cr_neg1
RawCounts_Cr.txt
Liver of Citrobacter rodentium infected mouse 1 (liver CFU negative)
Library 4


description:
Cr_Ctrl2
RawCounts_Cr.txt
Liver of uninfected mouse 2 (control group in Citrobacter rodentium experiment)
Library 2

value: uninfected mouse (control group in Citrobacter rodentium experiment)


title: 180822_P1-180822_P2-D1-AD008

value: ACA


title: 180822_P1-180822_P2-D1-AD007


title: 4_DDC7

city: Ghent

first: Lindsey

last: Devisscher

address: Corneel Heymanslaan 10

country: Belgium

institute: Ghent University

data_processing:
Extract UMIs, UMI-tools 1.1.1, command: umi_tools extract -I <in.fastq.gz> -S <out.fastq.gz>  --extract-method=regex  --bc-pattern='(?P<umi_1>.{{6}})(?P<discard_1>.{{4}}).*'
trim_galore --cores <threads> --stringency 3 --three_prime_clip_R1 3 -a " AGATCGGAAGAGC -a AAAAAAAAAAAA" --length 25 --output_dir <output_dir> <in.fastq.gz>
Mapping reads, STAR 2.7.9a, command: STAR --runThreadN 10 --genomeDir <index_star> --readFilesIn <in.fastq.gz> --outFilterType BySJout --outFilterMultimapNmax 20 --alignSJoverhangMin 8 --alignSJDBoverhangMin 1 --outFilterMismatchNmax 999 --outFilterMismatchNoverLmax 0.6 --alignIntronMin 20 --alignIntronMax 1000000 --alignMatesGapMax 1000000 --outSAMattributes NH HI NM MD --outSAMtype BAM SortedByCoordinate --outFileNamePrefix <sample_> --quantMode TranscriptomeSAM
Process UMIs, UMI-tools 1.1.1, command: umi_tools dedup -I <in.bam> --output-stats=deduplicated -S <out.bam>
Quantify expression with RSEM, RSEM 1.3.1, rsem-calculate-expression -p 60 --forward-prob 1 --no-bam-output --bam <in.bam> <rsem_index> <sample_name>
Genome_build: GRCm39, release 103
Supplementary_files_format_and_content: normalized counts (DESeq2)

source_name: Liver tissue

extract_protocol:
RNA was extracted from 20 mg mouse liver tissue preserved in RNAlater (ThermoFisher Scientific, Belgium) using the Aurum Total RNA Mini Kit (Bio-Rad Laboratories, Belgium), following manufacturer’s protocol. Quantification and purity of the isolated RNA were determined via spectrophotometric analysis with a Nanodrop spectrophotometer (ThermoFisher Scientific, Belgium).
Library construction was performed with the QuantSeq 3’ mRNA FWD kit (Lexogen) and UMI Second Strand Synthesis Module (Lexogen).

tag: disease state

value: Acute cholestasis

value: Liver


title: 5_Sham

value: Control, healthy


title: 6_DDC7


title: 4_Sham


title: 6_Sham


title: 180822_P1-180822_P2-D1-AD004

value: THp


title: 3_Sham


title: 180822_P1-180822_P2-D1-AD006

value: brain


title: 2_DDC7


title: 3_DDC7


description:
Cr_neg3
RawCounts_Cr.txt
Liver of Citrobacter rodentium infected mouse 3 (liver CFU negative)
Library 6

value: liver


title: 4_DDC28

value: Chronic cholestasis


title: 180822_P1-180822_P2-D1-AD001

value: THl

value: SSp


title: 180822_P1-180822_P2-C9-AD008

value: female


title: 180822_P1-180822_P2-D1-AD012


title: 5_DDC28


title: 3_DDC28


title: Omasum newborn

source_name: Omasum

value: Holstein dairy cows

value: omasum

value: newborn


title: 2_DDC28


description:
Cr_Ctrl3
RawCounts_Cr.txt
Liver of uninfected mouse 3 (control group in Citrobacter rodentium experiment)
Library 3


title: 180822_P1-180822_P2-C9-AD007

value: male


title: 180822_P1-180822_P2-D1-AD010


description:
Cr_Ctrl1
RawCounts_Cr.txt
Liver of uninfected mouse 1 (control group in Citrobacter rodentium experiment)
Library 1


title: 5_CsA7


title: 180822_P1-180822_P2-C9-AD006


title: 5_CsA28


title: 4_CsA7


title: 4_CTLD


title: 6_CTLD


title: 4_CsA28


title: 2_CsA28


title: 180822_P1-180822_P2-C9-AD001


title: 180822_P1-180822_P2-C9-AD002


title: 180822_P1-180822_P2-C9-AD004


title: 180822_P1-180822_P2-C8-AD010


title: 2_CTLD


title: 6_CTL


title: 4_CTL


title: 3_CTL


title: 180822_P1-180822_P2-C8-AD007


title: 1_CTL


description:
Lm_Ctrl1
RawCounts_Lm.txt
Liver of uninfected mouse 1 (control group in Listeria monocytogenes experiment)
Library 10

value: uninfected mouse (control group in Listeria monocytogenes experiment)


title: 6_CsA7


title: 5_BDL7


title: 180822_P1-180822_P2-C8-AD004


title: 3_BDL7


title: CD28 CAR, day 0, donor 18, biol rep 1, RNAseq

city: St. Louis

first: Nathan

last: Singh

state: MO

institute: Washington University School of Medicine

data_processing:
Basecalls and demultiplexing were performed with Illumina’s bcl2fastq2 software.
RNA-seq reads were then aligned and quantitated to the Ensembl release 101 primary assembly with an Illumina DRAGEN Bio-IT on-premise server running version 3.9.3-8 software.
Assembly: hg38
Supplementary files format and content: tab-delimited text file with gene counts

organism: Homo sapiens

source_name: Engineered T cells

extract_protocol:
Total RNA was extracted from purifiied CAR T cellsusing Qiazol (Qiagen) and recovered by RNA Clean and Concentrator spin columns (Zymo)
Samples were prepared according to library kit manufacturer’s protocol, indexed, pooled, and sequenced on an Illumina NovaSeq 6000

treatment_protocol: CD19-directed human CAR T cells containing either CD28 (19/28) or 41BB (19/BB) domains were combined with the CD19+ ALL cell line Nalm6. These co-cultures were established at an effector-to-target (E:T) ratio of 1:8 and monitored routinely for dynamic changes in quantity of CAR T cells and Nalm6. We maintained consistent CAR stimulation by adding additional Nalm6 to re-establish an E:T ratio of 1:8 every ~48 hours.

tag: individual

value: donor 18

value: Primary

tag: cell type

value: CD19-directed human CAR T cells containing CD28

value: Co-culture with Nalm6 cells

tag: time


title: 180822_P1-180822_P2-C8-AD002


title: 2_BDL7

value: Acute cholestasis

value: Liver


title: 3_CsA7


title: CD28 CAR, day 0, donor 18, biol rep 2, RNAseq


title: 1_CsA28


title: 6_BDL28


title: 1_BDL7


title: 180822_P1-180822_P2-C7-AD010

value: brain

value: THp

value: ACA


title: 4_BDL28

value: Chronic cholestasis


title: 5_BDL28


title: 2_BDL28


title: 180822_P1-180822_P2-C8-AD006

value: SSp


title: 3_CTLD

value: Control, healthy


title: 41BB CAR, day 0, donor 18, biol rep 1, RNAseq

value: CD19-directed human CAR T cells containing 41BB


title: 41BB CAR, day 0, donor 18, biol rep 2, RNAseq


title: 180822_P1-180822_P2-C7-AD008

value: THl

value: female


title: 180822_P1-180822_P2-C8-AD001

value: male


title: 180822_P1-180822_P2-C7-AD004


title: CD28 CAR, day 6, donor 18, biol rep 1, RNAseq


title: 180822_P1-180822_P2-C7-AD001


title: CD28 CAR, day 6, donor 18, biol rep 2, RNAseq


title: CD28 CAR, day 6, donor 18, biol rep 3 , RNAseq


title: 180822_P1-180822_P2-C6-AD012


title: Cdk13Δ_c14-1_minusDox72_rep1

city: Cambridge

first: Sara

middle: Jane

last: Dubbury

state: MA

address: 500 Main Street, 76-417

department: Koch Institute

institute: MIT

data_processing:
Raw RNA-seq reads were mapped using STAR aligner version 2.4.1d with the following parameters (Note: Boutz_Mm9_Master_05_06_13_mod_base1_1.juncs is a custom annotated master junctions file for mouse embryonic stem cells and can be made available upon request.):  STAR --runMode alignReads --runThreadN 8 --genomeDir UCSC_mm9 --sjdbFileChrStartEnd Boutz_Mm9_Master_05_06_13_mod_base1_1.juncs --twopassMode Basic --sjdbOverhang 74 --outReadsUnmapped Fastx --outSAMtype BAM Unsorted SortedByCoordinate --outFilterMultimapNmax 20 --outFilterMismatchNmax 999 --outFilterMismatchNoverLmax 0.04 --alignIntronMin 70 --alignIntronMax 500000 --alignMatesGapMax 500000 --alignSJoverhangMin 8 --alignSJDBoverhangMin 1 --outSAMstrandField intronMotif --outFilterType BySJout
We used custom Python scripts to derive a transcriptome annotation based on Mm9 (Mus_musculus_NCBI_build37.1) gene start and end boundaries, the location of polyadenylation sites as determined by 3’end sequencing (Almada et al, 2013), and the genomic locations of all mapped splice junctions from the RNA-seq data. Mapped splice junctions detected in all samples were combined and processed using custom Python scripts to filter out junctions representing < ~1% of transcripts. Alternative and constitutive intron classifications were performed using custom Python scripts, and are agnostic with regard to existing annotations other than known gene boundaries. If no overlapping introns exist for a given intron, it is assigned to the constitutive class. The subgroups containing overlapping introns are assigned a splicing classification if the start and end coordinates of all of the constituent introns fall into a pattern representing a known splice type (cassette, mutually exclusive, alternative 5' splice site, alternative 3' splice site). Annotation of the distal polyadenylation site isoform for each gene was based on the consensus isoform, i.e. the junctions defining each exon are the most frequently used junction detected among all of the combined samples. Each IPA site within a gene was then assigned to an additional transcript based on the consensus isoform but terminating at the IPA cleavage site. These gene annotations were then converted to DEXseq exon parts using DEXseq-associated script dexseq_prepare_annotation.py and the reads mapping to each exon part in each sample were counted using dexseq_count.py. Using the counts matrix thus derived for all biological replicates in each condition, DEXseq was used to identify changes in the relative abundance of each exon part as normalized to all exon parts within the gene, including those representing alternative splicing of internal introns as well as the IPAs and distal polyadenylation site isoform 3’ terminal exon. This gave a log2-fold change and FDR adjusted p value for each exon part as it differed between the Cdk13-expressing and Cdk13-depleted samples. IPA sites or distal polyadenylation site isoform 3’ terminal exons whose exon part exhibited an adjusted p value < 0.05 were considered statistically significant. Significantly changing alternative exons,IPA sites and distal polyadenylation site isoform 3’ terminal exons are listed in the Cdk13_48_72_All_DEXSeq_results.xslx processed data file.
Genome_build: mm9
Supplementary_files_format_and_content: Excel table containing annotated exon parts, quantification and determination of statistical significance for changes in exon part expression between Cdk13-expressing and -depleted cells.

molecule: polyA RNA

growth_protocol: V6.5 (C57Bl/6-129) mESCs and derived cell lines were cultured on 0.2% gelatin-coated tissue culture plates in ES media (HEPES-buffered DMEM (Thermo Fisher) supplemented with 15% FBS (Hyclone), 1000U/mL LIF (Millipore), non-essential amino acids, L-glutamine, BME, penicillin, and streptomycin). Cdk13∆ clones were maintained in 1ug/mL doxycycline (dox) (Sigma) in ES media (changed daily) to sustain complementing levels of Cdk13. To investigate Cdk13 loss, cells were washed at time zero with HBS and switched to ES media without dox for 48 or 72 hours.

extract_protocol:
RNA Sequencing: Total RNA was harvested using Trizol Reagent (Thermo Fischer) following the manufacturer's protocol and subsequently DNase treated with Turbo DNase (Thermo Fishcer) under standard reaction conditions. RNA quality was assessed by the Agilent 2100 BioAnalyzer and only samples with a RIN value >= 8.9 were used for library preparation.
RNA Sequencing: Libraries were made from 1ug of total RNA input using the TruSeq Stranded mRNA Library Prep Kit (Illumina RS-122-2102) with multiplexing barcodes, following the standard protocol with the following specifications: (1) 5 min RNA fragmentation time, (2) Superscript III (Thermo Fisher) was used for reverse transcription, (3) 15 cycles of PCR were used during the library amplification step, and (4) AMPure beads (Beckman Coulter) were used to size select/purify the library post PCR amplification instead of gel size selection.

tag: strain background

value: Embryonic Stem Cells

tag: clone number

tag: dox treatment

value: -Dox 72 hours

tag: barcode

value: ACTGAT

tag: rin value


title: Cdk13Δ_c14-1_minusDox48_rep1

treatment_protocol: Cdk13∆ clones were maintained in 1ug/mL doxycycline (dox) (Sigma) in ES media (changed daily) to sustain complementing levels of Cdk13. To investigate Cdk13 loss, cells were washed at time zero with HBS and switched to ES media without dox for 48 or 72 hours.

value: -Dox 48 hours

value: GTGGCC


title: Cdk13Δ_c11-8_minusDox48_rep1

value: TAGCTT


title: Cdk13Δ_c11-8_minusDox72_rep2

value: GAGTGG


title: Cdk13Δ_c14-1_minusDox72_rep2

value: ATTCCT


title: Cdk13Δ_c11-8_minusDox48_rep2

value: GGCTAC


title: Cdk13Δ_c14-1_minusDox48_rep2

value: GTTTCG


title: Cdk13Δ_c14-1_plusDox_rep2

value: +Dox

value: GATCAG


title: CD28 CAR, day 0, donor 18, biol rep 3 , RNAseq

value: donor 18

value: Primary

value: CD19-directed human CAR T cells containing CD28

tag: treatment

value: Co-culture with Nalm6 cells


title: Cdk13Δ_c14-1_plusDox_rep1

value: ACTTGA


title: Cdk13Δ_c11-8_plusDox_rep1

value: Embryonic Stem Cells

value: ATCACG


title: 180822_P1-180822_P2-C6-AD010

tag: tissue

value: brain

tag: source

value: THp

value: ACA

value: male


title: 41BB CAR, day 6, donor 18, biol rep 1, RNAseq

value: CD19-directed human CAR T cells containing 41BB


title: Cdk13Δ_c11-8_plusDox_rep2

value: TTAGGC


title: 41BB CAR, day 0, donor 18, biol rep 3 , RNAseq


title: Cdk13Δ_c11-8_minusDox72_rep1

value: -Dox 72 hours

value: CGTACG


title: 41BB CAR, day 6, donor 18, biol rep 2, RNAseq


title: 180822_P1-180822_P2-C6-AD006

tag: target

value: SSp

tag: Sex

value: female


title: 180822_P1-180822_P2-C7-AD006


title: StemCell CD11b positive selection enrichment KO_CP

city: Toronto

first: Arthur

last: Mortha

address: 1 King's College Circle

country: Canada

institute: University of Toronto

description: 10x genomics

library_source: transcriptomic single cell

data_processing:
The demultiplexing, barcoded processing, gene counting and aggregation were made using the Cell Ranger software v2.1.1 (https://support.10xgenomics.com/single-cell-gene-expression/software/pipelines/latest/what-is-cell-ranger)
Assembly: mm10
Supplementary files format and content: Tab-separated values files and matrix files

source_name: colon solitary isolated lymphoid tissues

extract_protocol: Library was performed according to the manufacter’s instructions (single cell 3’ v2 protocol, 10x Genomics). Briefly, GCs were resuspended in the master mix and loaded together with partitioning oil and gel beads into the chip to generate the gel bead-in-emulsion (GEM). The poly-A RNA from the cell lysate contained in every single GEM was retrotranscripted to cDNA, which contains an Ilumina R1 primer sequence, Unique Molecular Identifier (UMI) and the 10x Barcode. The pooled barcoded cDNA was then cleaned up with Silane DynaBeads, amplified by PCR and the apropiated sized fragments were selected with SPRIselect reagent for subsequent library construction. During the library construction Ilumina R2 primer sequence, paired-end constructs with P5 and P7 sequences and a sample index were added.

value: CD11b+ cells


title: StemCell CD11b positive selection enrichment WT_CP

description: 10x genomics

value: colon solitary isolated lymphoid tissues


title: StemCell CD11b positive selection enrichment KO_LP

description: 10x genomics

source_name: colon lamina propria

value: colon lamina propria


title: 180822_P1-180822_P2-C6-AD004


title: CD28 CAR, day 15, donor 18, biol rep 1, RNAseq

value: donor 18

value: Primary

value: CD19-directed human CAR T cells containing CD28

value: Co-culture with Nalm6 cells


title: 180822_P1-180822_P2-C6-AD002

value: brain

value: THl


title: StemCell CD11b positive selection enrichment WT_LP

description: 10x genomics


title: CD28 CAR, day 15, donor 18, biol rep 2, RNAseq


title: rep2, plate6, spaceH8

city: Stanford

first: Alan

last: Cheng

address: 240 Pasteur Drive

institute: Stanford University

data_processing:
Raw reads were pre-processed using FASTQC, Cutadapt, and PRINSEQ, followed by sequence alignment with the Tuxedo suite (Bowtie, Bowtie2, TopHat) and SAMtools, using default settings.
Transcript counts were quantified as fragments per kilobase of transcript per million mapped reads (FPKM) generated by TopHat/Cufflinks.
Assembly: mm10

source_name: otic vesicle and periotic tissue

extract_protocol:
Dissociated tdTomato+ cells were sorted into individual wells of 96-well plates using a FACSAria II instrument. Each well contained 5ul of lysis buffer from the Clontech SMARTer Ultra Low Input RNA kit V3, which was used for cDNA generation following manufacturer instructions.
Illumina libraries were constructed using the Nextera XT DNA Sample Preparation kit following manufacturer instructions.

value: otic vesicle and periotic tissue

tag: developmental stage

value: tdTomato+


title: rep2, plate5, spaceH8


title: rep1, plate3, spaceH9


title: 180822_P1-180822_P2-C5-AD010

value: THp

value: ACA

value: male


title: CD28 CAR, day 15, donor 18, biol rep 3 , RNAseq

tag: cell line


title: 180822_P1-180822_P2-C6-AD001


title: 41BB CAR, day 15, donor 18, biol rep 1, RNAseq

value: CD19-directed human CAR T cells containing 41BB


title: 180822_P1-180822_P2-C5-AD008


title: rep2, plate4, spaceH9

tag: genotype


title: rep2, plate5, spaceH9


title: rep1, plate1, spaceH9


title: rep2, plate6, spaceH7


title: rep1, plate2, spaceH8


title: rep1, plate3, spaceH8


title: 41BB CAR, day 6, donor 18, biol rep 3 , RNAseq


title: rep2, plate5, spaceH7


title: rep1, plate3, spaceH7


title: rep1, plate2, spaceH7


title: 180822_P1-180822_P2-C5-AD004

value: SSp


title: 180822_P1-180822_P2-C5-AD007


title: 180822_P1-180822_P2-C5-AD006

value: female


title: CD28 CAR, day 6, donor 30, biol rep 1, RNAseq

value: donor 30


title: rep2, plate5, spaceH6


title: rep2, plate6, spaceH6


title: rep1, plate3, spaceH6


title: 180822_P1-180822_P2-C5-AD001


title: CD28 CAR, day 6, donor 30, biol rep 2, RNAseq


title: 41BB CAR, day 15, donor 18, biol rep 3 , RNAseq


title: rep2, plate6, spaceH5


title: 41BB CAR, day 15, donor 18, biol rep 2, RNAseq


title: rep2, plate5, spaceH5


title: CD28 CAR, day 6, donor 30, biol rep 3 , RNAseq


title: 180822_P1-180822_P2-C3-AD010


title: rep1, plate3, spaceH5


title: rep1, plate1, spaceH5


title: rep2, plate5, spaceH4


title: 180822_P1-180822_P2-C3-AD012


title: 41BB CAR, day 6, donor 30, biol rep 3, RNAseq


title: 180822_P1-180822_P2-C3-AD008


title: 180822_P1-180822_P2-C3-AD007


title: 41BB CAR, day 6, donor 30, biol rep 2, RNAseq


title: 41BB CAR, day 13, donor 30, biol rep 2, RNAseq


title: rep1, plate2, spaceH4


title: rep2, plate4, spaceH4


title: rep1, plate3, spaceH4


title: rep1, plate1, spaceH4


title: 41BB CAR, day 6, donor 30, biol rep 1, RNAseq


title: 180822_P1-180822_P2-C3-AD006


title: 180822_P1-180822_P2-C3-AD004


title: rep2, plate6, spaceH4


title: 41BB CAR, day 13, donor 30, biol rep 3 , RNAseq


title: rep2, plate6, spaceH3


title: CD28 CAR, day 8, donor 31, biol rep 1, RNAseq

value: donor 31


title: rep2, plate5, spaceH3


title: CD28 CAR, day 8, donor 31, biol rep 2, RNAseq


title: 180822_P1-180822_P2-C3-AD001


title: rep2, plate4, spaceH5


title: rep2, plate4, spaceH3


title: rep1, plate2, spaceH5


title: rep1, plate3, spaceH3


title: 180822_P1-180822_P2-C3-AD002


title: rep1, plate1, spaceH3


title: CD28 CAR, day 8, donor 31, biol rep 3 , RNAseq


title: rep1, plate2, spaceH3


title: 180822_P1-180822_P2-C2-AD012


title: rep2, plate6, spaceH2


title: 180822_P1-180822_P2-C2-AD010


title: rep2, plate5, spaceH2


title: 41BB CAR, day 8, donor 31, biol rep 1, RNAseq


title: 41BB CAR, day 8, donor 31, biol rep 2, RNAseq


title: rep1, plate3, spaceH2


title: 180822_P1-180822_P2-C2-AD008


title: rep1, plate2, spaceH2


title: rep2, plate4, spaceH12


title: 41BB CAR, day 8, donor 31, biol rep 3, RNAseq


title: rep2, plate4, spaceH2


title: 180822_P1-180822_P2-C2-AD007


title: rep2, plate6, spaceH12


title: 180822_P1-180822_P2-C2-AD006


title: rep2, plate5, spaceH12


title: 180822_P1-180822_P2-C2-AD002


title: 41BB CAR, day 17, donor 31, biol rep 1, RNAseq


title: rep1, plate1, spaceH12


title: rep2, plate5, spaceH11


title: rep1, plate3, spaceH11


title: 41BB CAR, day 17, donor 31, biol rep 2, RNAseq


title: rep2, plate4, spaceH11


title: CD28 CAR, day 17, donor 31, biol rep 1, RNAseq


title: 41BB CAR, day 17, donor 31, biol rep 3 , RNAseq


title: 180822_P1-180822_P2-C2-AD001


title: rep1, plate2, spaceH11


title: 180822_P1-180822_P2-C12-AD010


title: 180822_P1-180822_P2-C12-AD012


title: rep2, plate5, spaceH10


title: rep1, plate2, spaceH12


title: 180822_P1-180822_P2-C12-AD008


title: CD28 CAR, day 17, donor 31, biol rep 2, RNAseq


title: rep2, plate4, spaceH10


title: rep1, plate2, spaceH10


title: CD28 CAR, day 17, donor 31, biol rep 3, RNAseq


title: rep1, plate1, spaceH10


title: rep2, plate6, spaceH1


title: 180822_P1-180822_P2-C12-AD006


title: rep2, plate6, spaceH11


title: CD28 CAR, day 17, donor 30, biol rep 2, RNAseq


title: CD28 CAR, day 17, donor 30, biol rep 1, RNAseq


title: rep2, plate5, spaceH1


title: 180822_P1-180822_P2-C12-AD007


title: 180822_P1-180822_P2-C12-AD004


title: rep2, plate4, spaceH1


title: rep1, plate3, spaceH1


title: CD28 CAR, day 6, donor 32, biol rep 1, RNAseq

value: donor 32


title: rep1, plate1, spaceH1


title: 180822_P1-180822_P2-C12-AD002


title: CD28 CAR, day 17, donor 30, biol rep 3 , RNAseq


title: rep2, plate6, spaceG9


title: rep2, plate4, spaceG9


title: 180822_P1-180822_P2-C11-AD010


title: rep1, plate3, spaceG9


title: 180822_P1-180822_P2-C11-AD012


title: rep2, plate5, spaceG9


title: CD28 CAR, day 6, donor 32, biol rep 3 , RNAseq


title: rep1, plate2, spaceG9


title: 180822_P1-180822_P2-C11-AD008


title: rep1, plate1, spaceG9


title: rep1, plate3, spaceH10


title: 41BB CAR, day 6, donor 32, biol rep 1, RNAseq


title: rep2, plate6, spaceG8


title: rep2, plate5, spaceG8


title: rep2, plate4, spaceG8


title: 41BB CAR, day 6, donor 32, biol rep 2, RNAseq


title: rep1, plate1, spaceG8


title: rep2, plate6, spaceG7


title: rep1, plate2, spaceH1


title: 180822_P1-180822_P2-C11-AD004


title: CD28 CAR, day 6, donor 32, biol rep 2, RNAseq


title: rep2, plate5, spaceG7


title: CD28 CAR, day 17, donor 32, biol rep 1, RNAseq


title: 180822_P1-180822_P2-C11-AD002


title: 180822_P1-180822_P2-C11-AD006


title: CD28 CAR, day 17, donor 32, biol rep 2, RNAseq


title: rep2, plate4, spaceG7


title: 180822_P1-180822_P2-C10-AD008


title: 180822_P1-180822_P2-C10-AD007


title: rep1, plate1, spaceG7


title: 180822_P1-180822_P2-C11-AD007


title: rep1, plate3, spaceG7


title: 180822_P1-180822_P2-C10-AD006


title: rep2, plate6, spaceG6


title: rep2, plate6, spaceG5


title: rep1, plate3, spaceG8


title: CD28 CAR, day 17, donor 32, biol rep 3, RNAseq


title: 41BB CAR, day 13, donor 32, biol rep 2, RNAseq


title: 41BB CAR, day 13, donor 32, biol rep 1, RNAseq


title: rep2, plate4, spaceG6


title: 41BB CAR, day 6, donor 32, biol rep 3 , RNAseq


title: rep2, plate4, spaceG5


title: 41BB CAR, day 13, donor 32, biol rep 3 , RNAseq


title: rep1, plate3, spaceG5


title: rep2, plate5, spaceG5


title: rep1, plate2, spaceG5


title: CD28 CAR, day 7, donor 33, biol rep 1, RNAseq

value: donor 33


title: rep1, plate1, spaceG5


title: rep2, plate6, spaceG4


title: 180822_P1-180822_P2-C10-AD001


title: 180822_P1-180822_P2-C10-AD002


title: CD28 CAR, day 7, donor 33, biol rep 2, RNAseq

value: Primary


title: rep2, plate5, spaceG4


title: 180822_P1-180822_P2-C1-AD012


title: rep1, plate2, spaceG6


title: rep2, plate4, spaceG4


title: CD28 CAR, day 7, donor 33, biol rep 3 , RNAseq

value: CD19-directed human CAR T cells containing CD28


title: rep1, plate3, spaceG4


title: 180822_P1-180822_P2-C1-AD010


title: rep1, plate1, spaceG4


title: 180822_P1-180822_P2-C1-AD008


title: rep2, plate6, spaceG3


title: 180822_P1-180822_P2-C10-AD004


title: rep1, plate3, spaceG3


title: rep2, plate5, spaceG3


title: 41BB CAR, day 7, donor 33, biol rep 3, RNAseq

value: Co-culture with Nalm6 cells


title: rep2, plate4, spaceG3


title: 41BB CAR, day 7, donor 33, biol rep 1, RNAseq


title: 180822_P1-180822_P2-C1-AD007

value: brain


title: 41BB CAR, day 7, donor 33, biol rep 2, RNAseq


title: rep1, plate2, spaceG3


title: 180822_P1-180822_P2-C1-AD006


title: rep1, plate1, spaceG3


title: 180822_P1-180822_P2-C1-AD004


title: rep2, plate5, spaceG2


title: CD28 CAR, day 14, donor 33, biol rep 1, RNAseq


title: CD28 CAR, day 14, donor 33, biol rep 2, RNAseq


title: rep2, plate6, spaceG2


title: rep2, plate4, spaceG2


title: 180822_P1-180822_P2-C1-AD001

value: THl


title: rep1, plate3, spaceG2

value: otic vesicle and periotic tissue


title: rep1, plate2, spaceG2

value: tdTomato+


title: 41BB CAR, day 14, donor 33, biol rep 1, RNAseq


title: CD28 CAR, day 14, donor 33, biol rep 3, RNAseq


title: rep1, plate1, spaceG2


title: 180822_P1-180822_P2-B9-AD012

value: THp


title: rep2, plate6, spaceG12


title: rep1, plate3, spaceG12


title: rep1, plate2, spaceG12


title: 41BB CAR, day 14, donor 33, biol rep 3, RNAseq


title: 41BB CAR, day 14, donor 33, biol rep 2, RNAseq


title: rep1, plate3, spaceG11


title: rep1, plate1, spaceG12


title: 180822_P1-180822_P2-B9-AD007

value: ACA


title: 180822_P1-180822_P2-B9-AD008


title: rep1, plate2, spaceG11


title: 180822_P1-180822_P2-B9-AD010

value: male


title: PREP0259_KAduB14669_SI-TT-C5v1_VDJ_TCR_pooled

first: Kwasi

last: Adu-Berchie

state: Massachusetts

address: 3 Blackfan Circle

institute: Harvard University

description: VDJ sequencing

data_processing:
Data processing was performed using the 10x Cell Ranger Software v 7.0.0
Supplementary files format and content: GRCh38-3.0.0
Supplementary files format and content: Tab-separated values files and matrix files

source_name: Primary cells

extract_protocol:
To isolate T cells from collagen gels, collagen matrices were mechanically chopped into small pieces and digested in 100U/ml collagenase type IV (ThermoFisher Scientific #17104019) at 37ºC for 1 hour in digestion media (RPMI 1640 + 10% FBS), with intermittent mechanical disruption through 1ml pipette tips. Collagenase was then quenched with MACS buffer (PBS, 0.5% BSA, 2mM EDTA), after which cells were then passed through 30µM strainers (Miltenyi #130-098-458) and washed twice in PBS. Live cells were enriched using a dead cell removal kit (Miltenyi #130-090-101) according to the manufacturer's protocol and resuspended in PBS+2% BSA
TCR sequencing was performed using the chromium single-cell RNA-seq 5’ (10x) with Feature Barcoding by the Bauer Core Facility at Harvard University using the NovaSeq SP

value: Primary cells

value: T cells


title: 180822_P1-180822_P2-B9-AD006

value: SSp

value: female


title: rep2, plate4, spaceG11


title: rep1, plate1, spaceG11


title: PREP0259_KAduB14669_SI-TT-D5v1_GEX_TCR_pooled

description: Gene expression


title: rep2, plate6, spaceG10


title: 180822_P1-180822_P2-B9-AD004


title: rep2, plate5, spaceG10


title: 180822_P1-180822_P2-B9-AD002


title: 2D_Control_D3

description: Activated T cells cultured on plate plastic for 3 days

data_processing:
Data processing was performed using the 10x Cell Ranger Software v 3.0.1
Supplementary files format and content: GRCh38-3.0.0
Supplementary files format and content: Tab-separated values files and matrix files

extract_protocol:
To isolate T cells from collagen gels, collagen matrices were mechanically chopped into small pieces and digested in 100U/ml collagenase type IV (ThermoFisher Scientific #17104019) at 37ºC for 1 hour in digestion media (RPMI 1640 + 10% FBS), with intermittent mechanical disruption through 1ml pipette tips. Collagenase was then quenched with MACS buffer (PBS, 0.5% BSA, 2mM EDTA), after which cells were then passed through 30µM strainers (Miltenyi #130-098-458) and washed twice in PBS. Live cells were enriched using a dead cell removal kit (Miltenyi #130-090-101) according to the manufacturer's protocol and resuspended in PBS+2% BSA
Library construction was performed by the Bauer Core Facility at Harvard University according to the manufacturer's protocol (10x Chromium Single Cell 3’ v3 platform)

value: Plate Culture


title: 180822_P1-180822_P2-B9-AD001


title: rep1, plate3, spaceG10


title: rep1, plate2, spaceG10


title: rep1, plate1, spaceG10


title: Crosslinked_Collagen_D3

description: Activated T cells cultured in slow relaxing matrices for 3 days

value: Slow Relaxing Matrix


title: rep2, plate4, spaceG12


title: PREP0259_KAduB14669_SI-NA-A1v1_SP_TCR_pooled

description: Cell hashing


title: rep2, plate5, spaceG1


title: 180822_P1-180822_P2-B8-AD008


title: 180822_P1-180822_P2-B8-AD007


title: rep1, plate1, spaceG1


title: rep2, plate6, spaceF9


title: rep1, plate2, spaceG1


title: rep2, plate5, spaceF9


title: Differentiated HSPCs day 9 erythroid, GATA1 library, rep32, scRNA-seq

first: Jorge Diego

last: Martin Rufino

address: 415 Main Street

institute: Harvard University / Broad Institute of MIT and Harvard / Boston Children's Hospital

description:
10x Genomics
perturb_BE_seq_GATA1_rep32_filtered_feature_bc_matrix.h5
perturb_BE_seq_GATA1_gex_rep32

data_processing:
The demultiplexing, barcoded processing, gene counting and aggregation were made using the Cell Ranger software v6.0.1 (https://support.10xgenomics.com/single-cell-gene-expression/software/pipelines/latest/what-is-cell-ranger).
Assembly: hg38
Supplementary files format and content: h5 files, contains gene expression matrices and cropseq transcript matrices, and barcoded antibody matrices (if applicable)

source_name: Hematopoietic system

extract_protocol:
Following final FACS enrichment to remove debris and purify transduced cells, 5-mL PBS-BSA coated tubes described above were spun down at 400 x g for 5 minutes at 4ºC. The entire supernatant was carefully removed and 1mL of PBS_0.05% BSA was added. Cells were then counted twice using an automated Countess 2 cell counter (Thermo Fisher, I-CACC2). Cell viability as determined by Trypan Blue was always >95%. Cells were again washed with 5mL PBS_0.05% BSA and resuspended at a final concentration of 1,000 cells / µL and kept on ice.
Single-cells were immediately processed using v3.1 Chemistry Dual Index kits according to manufacturer’s instructions (10X Genomics), using 20µL of cell suspension and 23.2 µL of water on the cell suspension loading step. Libraries were sequenced using a 28 base pair (Read 1), 10 base pair (Index 1), 10 base pair (Index 2), 90 base pair (Read 2) configuration on Nextseq 500, Nextseq 2000 or Novaseq S2/S4 instruments.

value: Differentiated hematopoietic stem and progenitor cells

value: Hematopoietic system

tag: library type

value: mRNA

tag: antibodies/tags


title: Differentiated HSPCs day 9 erythroid, GATA1 library, rep31, scRNA-seq

description:
10x Genomics
perturb_BE_seq_GATA1_rep31_filtered_feature_bc_matrix.h5
perturb_BE_seq_GATA1_gex_rep31

tag: cell type


title: rep2, plate4, spaceF9


title: 180822_P1-180822_P2-B8-AD006


title: rep2, plate4, spaceG10


title: rep1, plate2, spaceF9


title: 180822_P1-180822_P2-B8-AD004


title: rep2, plate6, spaceF8


title: rep2, plate5, spaceF8


title: Differentiated HSPCs day 9 erythroid, GATA1 library, rep30, scRNA-seq

description:
10x Genomics
perturb_BE_seq_GATA1_rep30_filtered_feature_bc_matrix.h5
perturb_BE_seq_GATA1_gex_rep30


title: Differentiated HSPCs day 9 erythroid, GATA1 library, rep29, scRNA-seq

description:
10x Genomics
perturb_BE_seq_GATA1_rep29_filtered_feature_bc_matrix.h5
perturb_BE_seq_GATA1_gex_rep29


title: rep2, plate4, spaceF8


title: 180822_P1-180822_P2-B8-AD010


title: rep1, plate3, spaceF8


title: rep2, plate6, spaceG1


title: rep1, plate2, spaceF8


title: NonCrosslinked_Collagen_D3

description: Activated T cells cultured in fast relaxing matrices for 3 days

value: Fast Relaxing Matrix


title: 180822_P1-180822_P2-B7-AD008

value: brain


title: D0_Control

description: Dynabead activated T cells before matrix culture

value: Pre-Gel control


title: rep1, plate1, spaceF8

value: otic vesicle and periotic tissue


title: rep2, plate6, spaceF7

value: tdTomato+


title: rep2, plate5, spaceF7


title: Differentiated HSPCs day 9 erythroid, GATA1 library, rep26, scRNA-seq

description:
10x Genomics
perturb_BE_seq_GATA1_rep26_filtered_feature_bc_matrix.h5
perturb_BE_seq_GATA1_gex_rep26


title: rep2, plate4, spaceF7


title: rep1, plate2, spaceF7


title: Differentiated HSPCs day 9 erythroid, GATA1 library, rep25, scRNA-seq

description:
10x Genomics
perturb_BE_seq_GATA1_rep25_filtered_feature_bc_matrix.h5
perturb_BE_seq_GATA1_gex_rep25


title: 180822_P1-180822_P2-B7-AD006

value: THp


title: rep1, plate1, spaceF7


title: 180822_P1-180822_P2-B8-AD002

value: THl


title: 180822_P1-180822_P2-B7-AD004

value: male


title: rep2, plate6, spaceF6


title: rep1, plate1, spaceF9


title: Differentiated HSPCs day 7 erythroid, GATA1 library, rep24, scRNA-seq

description:
10x Genomics
perturb_BE_seq_GATA1_rep24_filtered_feature_bc_matrix.h5
perturb_BE_seq_GATA1_gex_rep24


title: 180822_P1-180822_P2-B7-AD002


title: Differentiated HSPCs day 9 erythroid, GATA1 library, rep28, scRNA-seq

description:
10x Genomics
perturb_BE_seq_GATA1_rep28_filtered_feature_bc_matrix.h5
perturb_BE_seq_GATA1_gex_rep28


title: 180822_P1-180822_P2-B8-AD001


title: rep2, plate4, spaceF6


title: Differentiated HSPCs day 7 erythroid, GATA1 library, rep23, scRNA-seq

description:
10x Genomics
perturb_BE_seq_GATA1_rep23_filtered_feature_bc_matrix.h5
perturb_BE_seq_GATA1_gex_rep23


title: 180822_P1-180822_P2-B7-AD001


title: 180822_P1-180822_P2-B6-AD012

value: ACA


title: rep2, plate5, spaceF5


title: rep2, plate5, spaceF6


title: Differentiated HSPCs day 7 erythroid, GATA1 library, rep22, scRNA-seq

description:
10x Genomics
perturb_BE_seq_GATA1_rep22_filtered_feature_bc_matrix.h5
perturb_BE_seq_GATA1_gex_rep22


title: Differentiated HSPCs day 9 erythroid, GATA1 library, rep27, scRNA-seq

description:
10x Genomics
perturb_BE_seq_GATA1_rep27_filtered_feature_bc_matrix.h5
perturb_BE_seq_GATA1_gex_rep27


title: 180822_P1-180822_P2-B6-AD010


title: rep2, plate6, spaceF5


title: rep1, plate3, spaceF5


title: Differentiated HSPCs day 7 erythroid, GATA1 library, rep21, scRNA-seq

description:
10x Genomics
perturb_BE_seq_GATA1_rep21_filtered_feature_bc_matrix.h5
perturb_BE_seq_GATA1_gex_rep21


title: rep2, plate4, spaceF5


title: rep2, plate6, spaceF4


title: rep1, plate2, spaceF5


title: Differentiated HSPCs day 7 erythroid, GATA1 library, rep20, scRNA-seq

description:
10x Genomics
perturb_BE_seq_GATA1_rep20_filtered_feature_bc_matrix.h5
perturb_BE_seq_GATA1_gex_rep20


title: 180822_P1-180822_P2-B6-AD008


title: rep2, plate5, spaceF4


title: rep2, plate4, spaceF4


title: Differentiated HSPCs day 7 erythroid, GATA1 library, rep19, scRNA-seq

description:
10x Genomics
perturb_BE_seq_GATA1_rep19_filtered_feature_bc_matrix.h5
perturb_BE_seq_GATA1_gex_rep19


title: rep1, plate3, spaceF4


title: rep1, plate2, spaceF4


title: rep1, plate1, spaceF4


title: rep1, plate2, spaceF6


title: Differentiated HSPCs day 7 erythroid, GATA1 library, rep17, scRNA-seq

description:
10x Genomics
perturb_BE_seq_GATA1_rep17_filtered_feature_bc_matrix.h5
perturb_BE_seq_GATA1_gex_rep17


title: rep2, plate6, spaceF3


title: 180822_P1-180822_P2-B6-AD002


title: 180822_P1-180822_P2-B5-AD007


title: Differentiated HSPCs day 4 erythroid, GATA1 library, rep15, scRNA-seq

description:
10x Genomics
perturb_BE_seq_GATA1_rep15_filtered_feature_bc_matrix.h5
perturb_BE_seq_GATA1_gex_rep15


title: Differentiated HSPCs day 4 erythroid, GATA1 library, rep14, scRNA-seq

description:
10x Genomics
perturb_BE_seq_GATA1_rep14_filtered_feature_bc_matrix.h5
perturb_BE_seq_GATA1_gex_rep14


title: rep2, plate4, spaceF3


title: rep1, plate2, spaceF3


title: 180822_P1-180822_P2-B5-AD006


title: rep2, plate6, spaceF2


title: 180822_P1-180822_P2-B5-AD002


title: Differentiated HSPCs day 4 erythroid, GATA1 library, rep13, scRNA-seq

description:
10x Genomics
perturb_BE_seq_GATA1_rep13_filtered_feature_bc_matrix.h5
perturb_BE_seq_GATA1_gex_rep13


title: rep1, plate1, spaceF3


title: 180822_P1-180822_P2-B6-AD007


title: rep2, plate5, spaceF2


title: rep1, plate3, spaceF2


title: 180822_P1-180822_P2-B6-AD001


title: Differentiated HSPCs day 7 erythroid, GATA1 library, rep18, scRNA-seq

description:
10x Genomics
perturb_BE_seq_GATA1_rep18_filtered_feature_bc_matrix.h5
perturb_BE_seq_GATA1_gex_rep18


title: 180822_P1-180822_P2-B5-AD001


title: rep1, plate2, spaceF2


title: Differentiated HSPCs day 4 erythroid, GATA1 library, rep11, scRNA-seq

description:
10x Genomics
perturb_BE_seq_GATA1_rep11_filtered_feature_bc_matrix.h5
perturb_BE_seq_GATA1_gex_rep11


title: 180822_P1-180822_P2-B4-AD012


title: 180822_P1-180822_P2-B4-AD010


title: rep1, plate1, spaceF2


title: Differentiated HSPCs day 4 erythroid, GATA1 library, rep10, scRNA-seq

description:
10x Genomics
perturb_BE_seq_GATA1_rep10_filtered_feature_bc_matrix.h5
perturb_BE_seq_GATA1_gex_rep10


title: rep2, plate5, spaceF3


title: rep2, plate5, spaceF12


title: rep2, plate6, spaceF12


title: rep2, plate4, spaceF12


title: Differentiated HSPCs day 4 erythroid, GATA1 library, rep9, scRNA-seq

description:
10x Genomics
perturb_BE_seq_GATA1_rep9_filtered_feature_bc_matrix.h5
perturb_BE_seq_GATA1_gex_rep9


title: rep1, plate2, spaceF12


title: rep1, plate3, spaceF12


title: rep1, plate1, spaceF12


title: Differentiated HSPCs day 2 erythroid, GATA1 library, rep8, scRNA-seq

description:
10x Genomics
perturb_BE_seq_GATA1_rep8_filtered_feature_bc_matrix.h5
perturb_BE_seq_GATA1_gex_rep8


title: 180822_P1-180822_P2-B4-AD008


title: rep2, plate6, spaceF11


title: rep2, plate5, spaceF11


title: Differentiated HSPCs day 2 erythroid, GATA1 library, rep7, scRNA-seq

description:
10x Genomics
perturb_BE_seq_GATA1_rep7_filtered_feature_bc_matrix.h5
perturb_BE_seq_GATA1_gex_rep7


title: rep1, plate3, spaceF11


title: rep2, plate4, spaceF11


title: Differentiated HSPCs day 4 erythroid, GATA1 library, rep12, scRNA-seq

description:
10x Genomics
perturb_BE_seq_GATA1_rep12_filtered_feature_bc_matrix.h5
perturb_BE_seq_GATA1_gex_rep12


title: Differentiated HSPCs day 2 erythroid, GATA1 library, rep6, scRNA-seq

description:
10x Genomics
perturb_BE_seq_GATA1_rep6_filtered_feature_bc_matrix.h5
perturb_BE_seq_GATA1_gex_rep6


title: 180822_P1-180822_P2-B4-AD004


title: 180822_P1-180822_P2-B4-AD006


title: rep1, plate2, spaceF11


title: rep2, plate6, spaceF10


title: 180822_P1-180822_P2-B4-AD002


title: Differentiated HSPCs day 2 erythroid, GATA1 library, rep5, scRNA-seq

description:
10x Genomics
perturb_BE_seq_GATA1_rep5_filtered_feature_bc_matrix.h5
perturb_BE_seq_GATA1_gex_rep5


title: Differentiated HSPCs day 2 erythroid, GATA1 library, rep4, scRNA-seq

description:
10x Genomics
perturb_BE_seq_GATA1_rep4_filtered_feature_bc_matrix.h5
perturb_BE_seq_GATA1_gex_rep4


title: rep2, plate5, spaceF10


title: 180822_P1-180822_P2-B3-AD010


title: rep1, plate2, spaceF10


title: rep1, plate1, spaceF10


title: rep2, plate4, spaceF10


title: Differentiated HSPCs day 2 erythroid, GATA1 library, rep3, scRNA-seq

description:
10x Genomics
perturb_BE_seq_GATA1_rep3_filtered_feature_bc_matrix.h5
perturb_BE_seq_GATA1_gex_rep3


title: Differentiated HSPCs day 2 erythroid, GATA1 library, rep2, scRNA-seq

description:
10x Genomics
perturb_BE_seq_GATA1_rep2_filtered_feature_bc_matrix.h5
perturb_BE_seq_GATA1_gex_rep2


title: rep2, plate6, spaceF1


title: 180822_P1-180822_P2-B4-AD001


title: rep1, plate3, spaceF10


title: rep1, plate2, spaceF1


title: Differentiated HSPCs day 2 erythroid, GATA1 library, rep1, scRNA-seq

description:
10x Genomics
perturb_BE_seq_GATA1_rep1_filtered_feature_bc_matrix.h5
perturb_BE_seq_GATA1_gex_rep1


title: 180822_P1-180822_P2-B3-AD007


title: rep1, plate1, spaceF1


title: rep2, plate4, spaceF1


title: rep2, plate5, spaceE9


title: 180822_P1-180822_P2-B3-AD006


title: rep1, plate3, spaceE9


title: 180822_P1-180822_P2-B3-AD004


title: rep1, plate1, spaceE9


title: rep1, plate2, spaceE9


title: 180822_P1-180822_P2-B3-AD001


title: rep2, plate6, spaceE8


title: rep2, plate5, spaceE8


title: rep2, plate4, spaceE8


title: 180822_P1-180822_P2-B2-AD010


title: 180822_P1-180822_P2-B4-AD007


title: U2F PCCB-G2-NC5

city: Chang sha

first: Wendiao

state: Hunan

address: No. 932, Lushan South Road

institute: School of Life  Sciences, Central South University

data_processing:
RNA-seq reads were filtered to get clean reads using FastQC (v0.20.0)
The clean reads were aligned to the human genome GRCh38.p13 using STAR (v2.7.9a)
Transcript Per Million was calculated by RSEM (1.3.3)
Differential gene expression analysis was done using DEseq2 package (v1.34.0) in R (v4.1.1)
P value was adjusted using the Benjamini–Hochberg method
Assembly: hg38
Supplementary files format and content: csv, gene expression for all samples

source_name: U2F organoid

growth_protocol: hFOs were cultured using the STEMdiff Dorsal Forebrain Organoid Differentiation Kit (STEMCELL Technologies, 08620) based on the manufacturer’s instructions with some modifications.

extract_protocol:
Qiagen miRNeasy Mini Kit was used to extract total RNA
Sequencing libraries were prepared for RNA-seq using standard Illumina protocols

treatment_protocol: We used CRISPRi to knock down PCCB expression in U2F hiPSCs. The established PCCB knockdown hiPSCs were then used to generate hFOs. On day 60 of organoid culture,  total RNA was extracted for RNA-seq. analysis.

value: hiPSCs

value: control

value: human forebrain organoids (hFOs)


title: U2F PCCB-G2-1

value: PCCB knockdown


title: U2F PCCB-G2-2


title: U2F PCCB-G2-3


title: rep1, plate3, spaceE8


title: U2F PCCB-G2-NC4


title: rep1, plate2, spaceE8


title: 180822_P1-180822_P2-B2-AD008

value: female


title: rep2, plate6, spaceE7


title: 180822_P1-180822_P2-B2-AD007


title: rep1, plate1, spaceE8


title: U2F PCCB-G2-NC3


title: 180822_P1-180822_P2-B2-AD006

value: SSp


title: rep2, plate5, spaceE7


title: rep1, plate2, spaceE7


title: U2F PCCB-G2-NC2


title: rep2, plate4, spaceE7


title: 180822_P1-180822_P2-B2-AD012


title: rep1, plate1, spaceE7


title: rep2, plate6, spaceE6


title: 180822_P1-180822_P2-B2-AD002


title: U2F PCCB-G1-5


title: 180822_P1-180822_P2-B2-AD004


title: U2F PCCB-G2-NC1


title: rep1, plate3, spaceE6


title: 180822_P1-180822_P2-B2-AD001


title: rep1, plate2, spaceE6


title: rep2, plate5, spaceE6


title: U2F PCCB-G1-4


title: rep1, plate1, spaceE6


title: rep2, plate6, spaceE5


title: rep2, plate5, spaceE5


title: U2F PCCB-G1-3


title: 180822_P1-180822_P2-B12-AD012


title: rep2, plate4, spaceE5


title: rep1, plate3, spaceE5


title: 180822_P1-180822_P2-B12-AD010


title: rep1, plate2, spaceE5


title: U2F PCCB-G1-1


title: 180822_P1-180822_P2-B12-AD007


title: U2F PCCB-G1-NC5


title: rep2, plate6, spaceE4


title: rep2, plate5, spaceE4


title: rep1, plate1, spaceE5


title: 180822_P1-180822_P2-B12-AD006


title: U2F PCCB-G1-NC4


title: rep1, plate3, spaceE4


title: rep2, plate4, spaceE4


title: rep1, plate2, spaceE4


title: U2F PCCB-G1-NC3


title: rep1, plate1, spaceE4


title: 180822_P1-180822_P2-B12-AD004


title: rep2, plate6, spaceE3


title: U2F PCCB-G1-NC2


title: rep2, plate5, spaceE3


title: 180822_P1-180822_P2-B12-AD002


title: rep2, plate4, spaceE3


title: U2F PCCB-G1-NC1


title: rep1, plate3, spaceE3


title: 180822_P1-180822_P2-B12-AD001


title: 180822_P1-180822_P2-B12-AD008


title: rep1, plate2, spaceE3


title: 180822_P1-180822_P2-B11-AD012


title: rep1, plate1, spaceE3


title: U2F PCCB-G1-2


title: rep2, plate5, spaceE2


title: rep2, plate6, spaceE2


title: rep1, plate3, spaceE2


title: rep2, plate4, spaceE2


title: rep1, plate1, spaceE2


title: 180822_P1-180822_P2-B11-AD007


title: rep2, plate6, spaceE12


title: 180822_P1-180822_P2-B11-AD004


title: rep2, plate5, spaceE12


title: rep2, plate4, spaceE12


title: rep1, plate3, spaceE12


title: 180822_P1-180822_P2-B11-AD002


title: rep1, plate1, spaceE12


title: 180822_P1-180822_P2-B11-AD001


title: rep2, plate5, spaceE11


title: rep1, plate3, spaceE11


title: rep2, plate4, spaceE11


title: 180822_P1-180822_P2-B11-AD010


title: 180822_P1-180822_P2-B11-AD008


title: 180822_P1-180822_P2-B10-AD007


title: rep1, plate2, spaceE11


title: 180822_P1-180822_P2-B10-AD010


title: rep2, plate5, spaceE10


title: rep1, plate3, spaceE10


title: rep2, plate6, spaceE10


title: rep1, plate1, spaceE10


title: rep2, plate4, spaceE10


title: 180822_P1-180822_P2-B10-AD006


title: rep1, plate2, spaceE10


title: rep1, plate2, spaceE2


title: rep2, plate6, spaceE1


title: rep1, plate3, spaceE1


title: rep2, plate6, spaceD9


title: rep2, plate5, spaceD9


title: 180822_P1-180822_P2-B10-AD004


title: rep2, plate4, spaceE1


title: rep1, plate2, spaceD9


title: 180822_P1-180822_P2-B10-AD008


title: 180822_P1-180822_P2-B10-AD012


title: rep2, plate6, spaceD8


title: rep1, plate1, spaceD9


title: 180822_P1-180822_P2-B1-AD010


title: 180822_P1-180822_P2-B10-AD002


title: 180822_P1-180822_P2-B1-AD007


title: 180822_P1-180822_P2-B1-AD006


title: rep2, plate5, spaceD8


title: rep1, plate2, spaceD8


title: rep1, plate3, spaceD8


title: 180822_P1-180822_P2-B1-AD004


title: rep2, plate4, spaceD8


title: rep1, plate1, spaceD8


title: rep2, plate4, spaceD7


title: 180822_P1-180822_P2-B1-AD012


title: 180822_P1-180822_P2-B1-AD008


title: rep1, plate3, spaceD7


title: rep1, plate2, spaceD7


title: 180822_P1-180822_P2-B1-AD002


title: rep1, plate1, spaceE1


title: rep1, plate1, spaceD7


title: rep2, plate5, spaceE1


title: rep2, plate6, spaceD6


title: 180822_P1-180822_P2-B1-AD001


title: rep2, plate4, spaceD9


title: rep2, plate4, spaceD6


title: rep2, plate5, spaceD6


title: rep1, plate3, spaceD6


title: rep1, plate2, spaceD6


title: 180822_P1-180822_P2-A9-AD007


title: 180822_P1-180822_P2-A9-AD010


title: rep1, plate1, spaceD6


title: rep2, plate6, spaceD5


title: 180822_P1-180822_P2-A9-AD004


title: 180822_P1-180822_P2-A9-AD001


title: 180822_P1-180822_P2-A8-AD010


title: 180822_P1-180822_P2-A9-AD006


title: 180822_P1-180822_P2-A8-AD008


title: rep2, plate6, spaceD4


title: rep1, plate1, spaceD5


title: rep2, plate4, spaceD5


title: rep1, plate2, spaceD5


title: rep2, plate5, spaceD5


title: rep2, plate5, spaceD4


title: rep1, plate3, spaceD5


title: rep2, plate4, spaceD4


title: rep1, plate3, spaceD4


title: 180822_P1-180822_P2-A8-AD007


title: rep1, plate2, spaceD4


title: 180822_P1-180822_P2-A8-AD006


title: rep2, plate6, spaceD3


title: rep1, plate1, spaceD4


title: rep2, plate5, spaceD3


title: rep2, plate4, spaceD3


title: 180822_P1-180822_P2-A8-AD004


title: 180822_P1-180822_P2-A8-AD002


title: rep1, plate2, spaceD3


title: 180822_P1-180822_P2-A8-AD001


title: rep1, plate3, spaceD3


title: rep2, plate6, spaceD2


title: 180822_P1-180822_P2-A7-AD012


title: 180822_P1-180822_P2-A7-AD010


title: rep2, plate4, spaceD2


title: rep1, plate2, spaceD2


title: rep2, plate6, spaceD12


title: rep2, plate5, spaceD12


title: rep2, plate4, spaceD12


title: rep2, plate6, spaceD11


title: rep2, plate4, spaceD11


title: rep1, plate1, spaceD12


title: rep1, plate1, spaceD11


title: rep1, plate2, spaceD11


title: rep2, plate5, spaceD11


title: 180822_P1-180822_P2-A6-AD012


title: 180822_P1-180822_P2-A7-AD006


title: 180822_P1-180822_P2-A7-AD004


title: rep2, plate6, spaceD10


title: rep2, plate5, spaceD10


title: rep2, plate4, spaceD10


title: 180822_P1-180822_P2-A7-AD002


title: rep1, plate1, spaceD3


title: rep1, plate1, spaceD10


title: 180822_P1-180822_P2-A6-AD010


title: rep2, plate4, spaceD1


title: rep2, plate5, spaceD2


title: 180822_P1-180822_P2-A6-AD007


title: rep1, plate3, spaceD1


title: 180822_P1-180822_P2-A7-AD008


title: rep1, plate2, spaceC9


title: 180822_P1-180822_P2-A6-AD004


title: rep1, plate3, spaceC9


title: 180822_P1-180822_P2-A6-AD006


title: rep2, plate6, spaceC8


title: 180822_P1-180822_P2-A5-AD008


title: rep1, plate1, spaceC9


title: rep2, plate4, spaceC8


title: rep2, plate5, spaceC8


title: rep1, plate2, spaceC8


title: 180822_P1-180822_P2-A5-AD006


title: rep1, plate2, spaceD10


title: rep1, plate1, spaceC8


title: rep2, plate6, spaceC7


title: rep2, plate5, spaceC7


title: 180822_P1-180822_P2-A5-AD004


title: 180822_P1-180822_P2-A5-AD002


title: rep1, plate3, spaceC7


title: rep2, plate4, spaceC7


title: 180822_P1-180822_P2-A6-AD008


title: rep1, plate2, spaceC7


title: rep2, plate6, spaceC6


title: rep2, plate4, spaceC9


title: rep1, plate1, spaceC7


title: rep2, plate5, spaceC6


title: rep2, plate4, spaceC6


title: rep1, plate3, spaceC6


title: 180822_P1-180822_P2-A5-AD007


title: rep1, plate3, spaceC8


title: rep1, plate2, spaceC6


title: 180822_P1-180822_P2-A4-AD006


title: rep2, plate4, spaceC5


title: rep1, plate1, spaceC6


title: 180822_P1-180822_P2-A4-AD008


title: 180822_P1-180822_P2-A4-AD004


title: rep2, plate6, spaceC5


title: rep1, plate3, spaceC5


title: rep1, plate2, spaceC5


title: 180822_P1-180822_P2-A4-AD001


title: rep1, plate3, spaceC4


title: rep2, plate5, spaceC4


title: 180822_P1-180822_P2-A4-AD002


title: 180822_P1-180822_P2-A4-AD012


title: rep1, plate1, spaceC4


title: 180822_P1-180822_P2-A4-AD010


title: rep1, plate2, spaceC4


title: 180822_P1-180822_P2-A3-AD012


title: rep2, plate5, spaceC3


title: 180822_P1-180822_P2-A3-AD010


title: rep2, plate6, spaceC3


title: 180822_P1-180822_P2-A4-AD007


title: rep1, plate3, spaceC3


title: rep1, plate1, spaceC3


title: rep2, plate4, spaceC3


title: 180822_P1-180822_P2-A3-AD008


title: rep2, plate5, spaceC2


title: 180822_P1-180822_P2-A3-AD007


title: rep2, plate6, spaceC4


title: rep2, plate4, spaceC2


title: rep1, plate1, spaceC2


title: 180822_P1-180822_P2-A2-AD010


title: rep1, plate2, spaceC2


title: rep2, plate6, spaceC12


title: rep2, plate5, spaceC12


title: 180822_P1-180822_P2-A2-AD008


title: rep1, plate3, spaceC12


title: rep2, plate4, spaceC12


title: rep1, plate2, spaceC12


title: 180822_P1-180822_P2-A2-AD007


title: rep2, plate5, spaceC11


title: rep2, plate4, spaceC11


title: 180822_P1-180822_P2-A2-AD006


title: rep1, plate3, spaceC11


title: 180822_P1-180822_P2-A2-AD004


title: rep1, plate2, spaceC3


title: rep1, plate2, spaceC11


title: rep1, plate1, spaceC11


title: rep2, plate6, spaceC10


title: 180822_P1-180822_P2-A3-AD004


title: 180822_P1-180822_P2-A2-AD002


title: rep2, plate5, spaceC10


title: rep2, plate6, spaceC2


title: rep1, plate3, spaceC10


title: 180822_P1-180822_P2-A2-AD001


title: 180822_P1-180822_P2-A12-AD012


title: rep1, plate2, spaceC10


title: rep1, plate1, spaceC10


title: 180822_P1-180822_P2-A12-AD010


title: rep2, plate6, spaceC1


title: 180822_P1-180822_P2-A3-AD001


title: 180822_P1-180822_P2-A12-AD008


title: rep2, plate4, spaceC1


title: rep1, plate3, spaceC1


title: rep2, plate6, spaceB9


title: rep1, plate2, spaceC1


title: 180822_P1-180822_P2-A12-AD007


title: rep2, plate5, spaceB9


title: rep2, plate4, spaceB9


title: rep1, plate3, spaceB9


title: 180822_P1-180822_P2-A12-AD004


title: rep1, plate1, spaceB9


title: 180822_P1-180822_P2-A12-AD002


title: rep2, plate6, spaceB8


title: rep2, plate5, spaceB8


title: rep2, plate4, spaceB8


title: rep1, plate3, spaceB8


title: 180822_P1-180822_P2-A12-AD001


title: rep1, plate2, spaceB8


title: 180822_P1-180822_P2-A11-AD012


title: rep2, plate4, spaceB7


title: 180822_P1-180822_P2-A11-AD007


title: 180822_P1-180822_P2-A11-AD006


title: rep1, plate1, spaceB7


title: 180822_P1-180822_P2-A11-AD004


title: rep1, plate3, spaceB7


title: 180822_P1-180822_P2-A11-AD002


title: rep2, plate6, spaceB5


title: 180822_P1-180822_P2-A11-AD001


title: rep1, plate2, spaceB6


title: rep1, plate1, spaceB6


title: rep2, plate6, spaceB6


title: rep1, plate1, spaceC1


title: 180822_P1-180822_P2-A10-AD008


title: rep1, plate2, spaceB7


title: rep1, plate3, spaceB6


title: 180822_P1-180822_P2-A10-AD010


title: rep2, plate5, spaceB6


title: 180822_P1-180822_P2-A10-AD007


title: rep2, plate5, spaceB5


title: rep1, plate3, spaceB5


title: rep2, plate6, spaceB4


title: rep2, plate5, spaceB4


title: rep1, plate2, spaceB5


title: 180822_P1-180822_P2-A10-AD006


title: CD45+ immune cells, hMBP, scRNAseq

city: Leipzig

first: Ruth

middle: Martha

last: Stassart

address: Liebigstraße 19. Haus C

department: Paul-Flechsig-Institute of Brain Research

country: Germany

institute: University Clinic Leipzig

description: 10x Genomics

data_processing:
Raw Illumina bcl files were demultiplexed using the mkfastq command in bcl2fastq v.2.20. Gene expression matrices were generated with the count command in Cell Ranger v.3.1.0 using the default mm10 mouse genome.
For processed Seurat object using R v.4.1.2 and Seurat v. 4.1.1, Cells were filtered to contain more than 1000 UMIs, 500 genes, and less than 5% mitochondrial gene reads. Data were log-normalized,the 2000 most highly variable genes selected, and PCA was performed. The first 30 dimensions were selected for downstream analysis. Dimensionality reduction was performed using UMAP with n.neighbors = 30, min.dist = 0.3. Louvain clustering was performed with resolution = 0.8. Iterative quality control was performed by calculating marker genes for every cluster to remove contaminating cell populations. Doublets were removed using scDblFinder v.1.8.0 and scran v.1.22.1. Cell cycle scores were calculated regressed out during data scaling. Final clusters were annotated based on canonical marker genes. For detailed report see: https://github.com/nikofleischer/myelin-axonal_damage
Assembly: mm10
Supplementary files format and content: matrix files: raw output of Cell Ranger pipeline in .h5 format, processed Seurat object: both samples combined, dimensionality reductions and annotations

source_name: lumbosacral spinal cord white matter in EAE

extract_protocol:
Mice were transcardially perfused with ice-cold HBSS and the white matter of the lumbosacral spinal cord was dissected and dissociated using the adult brain dissociation kit (ABDK, Miltenyi Biotech) combined with manual mechanic dissociation and the translation and transcription inhibitors actinomysin D, triptolide, and asinomycin. Cells were strained through a 70 um cell strainer and debris isolated using the debris removal solution in the ABDK kit. Then, the cells were labelled with CD45+ microbeads (Miltenyi Biotech) and isolated through a magnetic column. 16K CD45+ cells were loaded into the 10x chip to achieve 10K cell recovery.
Library was performed according to the manufacter’s instructions (single cell 3’ v3.1 dual index protocol, 10x Genomics). Briefly, CD45+ cells were resuspended in the master mix and loaded together with partitioning oil and gel beads into the chip to generate the gel bead-in-emulsion (GEM). The poly-A RNA from the cell lysate contained in every single GEM was retrotranscripted to cDNA, which contains an Ilumina R1 primer sequence, Unique Molecular Identifier (UMI) and the 10x Barcode. The pooled barcoded cDNA was then cleaned up with Silane DynaBeads, amplified by PCR and the apropiated sized fragments were selected with SPRIselect reagent for subsequent library construction. During the library construction Ilumina R2 primer sequence, paired-end constructs with P5 and P7 sequences and a sample index were added.

treatment_protocol: Experimental autoimmune encephalomyelitis was induced by injecting an emulsion of Complete Freund’s Adjuvant (CFA) made from Mycobacterium tuberculosis (BD Bioscience, 231141) with a total of 200µg MOG35-55 (AnaSpec, AS-60130-1) per mouse subcutaneously into all 4 flanks. 400 ng pertussis toxin (Sigma, P7208) were administered intraperitoneally after immunization and 48 hours later. Mice were scored daily and the tissue was collected at peak EAE; at 4 days post onset.

value: CD45+ cells

value: hMBP

tag: strain

tag: age

value: 6 months old


title: 180822_P1-180822_P2-A10-AD004

value: brain


title: rep2, plate4, spaceB4


title: 180822_P1-180822_P2-A10-AD002


title: CD45+ immune cells, WT, scRNAseq

description: 10x Genomics

value: lumbosacral spinal cord white matter in EAE

value: WT


title: rep2, plate4, spaceB6

value: otic vesicle and periotic tissue

value: tdTomato+


title: 180822_P1-180822_P2-A10-AD001

value: THl


title: rep1, plate3, spaceB4


title: 180822_P1-180822_P2-A1-AD010

value: THp

value: male


title: rep2, plate4, spaceB5


title: rep2, plate6, spaceB3


title: rep2, plate5, spaceB3


title: 180822_P1-180822_P2-A1-AD012

value: ACA


title: untagged_1

city: Namur

first: Carlo

last: Yague-Sanz

address: 61 rue de Bruxelles

department: Medecine

institute: Namur University

data_processing:
Demultiplexing: Reads were assigned to experimental samples using their 5' barcode sequences, and barcodes were kept. Barcode is always first segment of the raw (.fastq) file name.
Collapsing reads: To remove PCR duplicates, reads were collapsed usinf fastx_collapser, part of FASTX Toolkit 0.0.14
Debrarcoding: The 5' barcodes were removed using pyBarcodeFilter.py v3.0 script from pyCRAC package.
Filtering reads: Reads were trimmed using flexbar v3.4.0 to remove the 3 -linker sequence (TGGAATTCTCGGGTGCCAAGGC) and were additionaly filtered and only 3'linker-containing reads were included to further analyses.
Reads alignment: Reads were aligned to the S. pombe genome sequence  by novoalign version 2.07.00.
Additional processing: novoalign SAM reads were modified using custom made script to keep only the 5' or 3' ends of reads. SAM files were converted to BAM format using samtools v1.9 and BigWig files were generated using bamCoverage v3.1.3.
Assembly: Genome_build: ASM294.2
Supplementary files format and content: Supplementary_files_format_and_content: Genomic coordinates of the 3' ends of mapped reads in BigWig format

organism: Schizosaccharomyces pombe

source_name: vegetative cells

growth_protocol: Cells were grown at 30°C in EMM medium supplemented with thiamine to allow Seb1 depletion until mid-log exponential phase (OD600nm ~0.5-0.8).

extract_protocol:
Crosslinked samples were lyzed in TNMC100 (50 mM Tris-HCl pH 7.5, 150 mM NaCl, 0.1% NP-40, 5 mM MgCl2, 10 mM CaCl2, 5 mM β-mercaptoethanol, 50U of DNase RQ1 and a protease-inhibitor cocktail) with zirconia beads in a 50 mL conical. Cleared lysates were incubated with IgG Sepharose and washed three times with TMN600 (50 mM Tris-HCl pH 7.5, 600 mM NaCl, 0.1% NP-40, 1.5 mM MgCl2) and two times TMN100 (50 mM Tris-HCl pH 7.5, 100 mM NaCl, 0.1% NP-40, 5 mM MgCl2). Protein:RNA complexes were eluted with HaloTEV and the supernantant was subjected to nickel affinity purification in denaturing conditions.
After extensive washing of the nickel beads, the addition of 3’ linkers, 5’ end radiolabeling, and 5’ linker ligation was performed on beads. Protein:RNA complexes were then eluted, precipitated, separated on gels, and transferred to nitrocellulose membranes. After detection of labeled RNAs by autoradiography, the appropriate region was excised and treated with Proteinase K. The RNA was then isolated, reverse transcribed and PCR amplified as described  (Turowki et al., 2020). Following gel extraction of PCR amplicons corresponding to 140-200 bp, libraries were measured by Qbit and analyzed by Illumina sequencing.
CRAC (Cross-linking and Analysis of cDNA)

treatment_protocol: Cells were cross-linked in culture media using megatron UVC cross-linker for 100 seconds as described (Turowski et al., 2020. DOI:10.1016/j.molcel.2020.06.002)

tag: strain name


title: Rpa2-HTP seb1d_2

value: h+ his7-366 leu1-32 ura4-d18 ade6-m210 pFB45::URA4 Rpa2-HTP::KanMx6 P41nmt1-seb1::natMx6


title: untagged_2


title: Rpa2-HTP seb1d_1


title: 180822_P1-180822_P2-A1-AD008


title: rep2, plate4, spaceB3


title: rep2, plate5, spaceC9


title: rep1, plate3, spaceB3


title: 180822_P1-180822_P2-A1-AD007


title: Rpa2-HTP WT_2

value: h+ his7-366 leu1-32 ura4-d18 ade6-m210 pFB45::URA4 Rpa2-HTP::KanMx6


title: 180822_P1-180822_P2-A1-AD006


title: Rpa2-HTP WT_1


title: 180822_P1-180822_P2-A1-AD004


title: rep1, plate2, spaceB3


title: HCC1954-Par 2

city: New York

first: Siting

last: Gan

state: NY

address: 417 East 68th Street

department: SKI

institute: MSKCC

description:
HCC1954_NormalizedCounts.xlsx
HCC1954_RawCounts.xlsx

data_processing:
FASTQ raw sequences were mapped by STAR RNA-seq aligner using human genome reference GRCh38 and mouse genome reference GRCm38
Uniquely mapped reads were assigned to annotated genes by HTSeq to measure read counts
Counts were normalized by library size to be subject to differential gene expression analysis
Assembly: GRCh38 and GRCm38
Supplementary files format and content: Processed data files are formated for normalized gene counts
Supplementary files format and content: Gene names are listed in the first column

growth_protocol: Cells were grown in either DMEM or RPMI based media supplemented with 10% FBS, 2mM L-glutamine, 100IU.mL Pen/strep, and 1 µg/mL amphotericin B to a confluence of 80%

extract_protocol:
Cells were extracted as a pellet and lysed using the QIAshredder columns in Buffer RLT from QIAgen supplemented with 1:100 β-Mercaptoethanol. RNA isolation was performed usng the QIAgen RNeasy mini kit.
cDNA libraries were constructed using either TruSeq RNA Sample Prep Kit v2 or NEBNext Ultra RNA Library Prep Kit and sequenced on a HiSeq 2500 system at a depth of 25-50 million reads per sample

value: breast cancer cell line

value: parental


title: rep1, plate1, spaceB3


title: HCC1954-Par 1


title: 180822_P1-180822_P2-A1-AD002

value: SSp

value: female


title: 180822_P1-180822_P2-A1-AD001


title: rep2, plate6, spaceB2


title: HCC1954-BrM 2b 2

value: brain metastatic derivative


title: 180801_CEMBA_mm_P56_P63_RS2_180801_P7_180801_P8_C2_AD004

value: STR


title: rep1, plate1, spaceB5


title: HCC1954-BrM 2b 1


title: 180801_CEMBA_mm_P56_P63_RS2_180801_P7_180801_P8_C2_AD001

value: brain


title: rep2, plate4, spaceB2


title: MDA231 IP 4

data_processing:
Xenome classified the reads as either human or mouse genome
FASTQ raw sequences were mapped by STAR RNA-seq aligner using human genome reference GRCh38 and mouse genome reference GRCm38
Uniquely mapped reads were assigned to annotated genes by HTSeq to measure read counts
Counts were normalized by library size to be subject to differential gene expression analysis
Assembly: GRCh38 and GRCm38

source_name: Brain

extract_protocol:
Mice were injected with 250mg/kg 5-fluorocytosine and after 12 hours brains were harvested and homogenized. mRNA was extracted by 4 × 250 uL/brain Oligo (dT)25 magnetic beads and immunoprecipitated with 2.5 ug/brain anti-BrdU antibody
SMARTer PCR cDNA kit was used to construct libraries using the immunoprecipitated mRNA

value: Brain


title: MDA231 IP 3


title: 180801_CEMBA_mm_P56_P63_RS2_180801_P7_180801_P8_C1_AD012

value: SC

value: P


title: rep1, plate2, spaceB2

value: otic vesicle and periotic tissue

value: tdTomato+


title: 180801_CEMBA_mm_P56_P63_RS2_180801_P7_180801_P8_C1_AD010


title: 180801_CEMBA_mm_P56_P63_RS2_180801_P7_180801_P8_C1_AD008


title: rep1, plate1, spaceB2


title: rep2, plate6, spaceB12


title: 180801_CEMBA_mm_P56_P63_RS2_180801_P7_180801_P8_C1_AD007


title: MDA231 IP 1


title: MDA231 input 4


title: HCC1954-Par 3


title: rep1, plate3, spaceB12


title: MDA231 input 2


title: MDA231 input 3


title: rep2, plate5, spaceB12


title: 180801_CEMBA_mm_P56_P63_RS2_180801_P7_180801_P8_C1_AD002

value: THl


title: rep1, plate2, spaceB12


title: MDA231 input 1


title: rep2, plate5, spaceB2


title: 180801_CEMBA_mm_P56_P63_RS2_180801_P7_180801_P8_C10_AD012


title: 180801_CEMBA_mm_P56_P63_RS2_180801_P7_180801_P8_C10_AD010


title: rep1, plate1, spaceB12


title: rep2, plate6, spaceB11


title: rep2, plate5, spaceB11


title: 180801_CEMBA_mm_P56_P63_RS2_180801_P7_180801_P8_C10_AD007

value: male


title: 180801_CEMBA_mm_P56_P63_RS2_180801_P7_180801_P8_C10_AD008


title: HCC1954 IP 3


title: HCC1954 IP 2


title: 180801_CEMBA_mm_P56_P63_RS2_180801_P7_180801_P8_C10_AD004

value: THp


title: HCC1954 IP 4


title: rep2, plate4, spaceB11


title: HCC1954 IP 1


title: rep1, plate3, spaceB11


title: HCC1954 input 4


title: rep2, plate4, spaceB12


title: rep1, plate2, spaceB11


title: HCC1954 input 3


title: 180801_CEMBA_mm_P56_P63_RS2_180801_P7_180801_P8_B9_AD010


title: rep2, plate6, spaceB10


title: HCC1954 input 2


title: rep1, plate1, spaceB11


title: 180801_CEMBA_mm_P56_P63_RS2_180801_P7_180801_P8_C11_AD006


title: 180801_CEMBA_mm_P56_P63_RS2_180801_P7_180801_P8_B9_AD008


title: HCC1954 input 1


title: rep2, plate5, spaceB10


title: 180801_CEMBA_mm_P56_P63_RS2_180801_P7_180801_P8_B9_AD006


title: 180801_CEMBA_mm_P56_P63_RS2_180801_P7_180801_P8_B9_AD002


title: rep2, plate4, spaceB10


title: rep1, plate3, spaceB10


title: rep1, plate1, spaceB10


title: ATF4mKO_22mo_3

city: Iowa City

first: Matthew

last: Miller

state: IA

address: 375 Newton Road

institute: University of Iowa

data_processing:
Sequencing reads were trimmed using Trimmomatic and mapped to the mm10 reference genome using RNA-STAR v2.7.8a using the online platform Galaxy
Reads were summarized by gene (mm10 entrez identifiers) using featureCounts v2.0.1
Assembly: mm10
Supplementary files format and content: Tab delimited text file containing matrix table of raw gene counts for every gene per sample

source_name: Tibialis anterior muscle

extract_protocol:
Tibialis anterior muscles were collected, flash frozen on liquid nitrogen, and RNA was harvested using Trizol reagent (Invitrogen) and purified using the RNeasy kit and RNase-free DNase Set (Qiagen). Samples were quantified using the Trinean DropSense 16 and RNA quality was assessed using the Agilent BioAnalyzer, with all samples showing an RNA integrity Number (RIN) greater than 8.
Libraries were prepared using the Illumina TruSeq Stranded mRNA Sample Prep Kit.

value: Tibialis anterior muscle

value: ATF4 flox/flox; MCK-Cre

value: 22 months


title: ATF4mKO_22mo_1


title: 180801_CEMBA_mm_P56_P63_RS2_180801_P7_180801_P8_C10_AD001

data_processing:
Reads trimed with Cutadapt for adapters and low quality bases (Phred score >= 20)
The resulting files are mapped to in-silico bisulfite converted reference genome using Bismark.
The raw BAM file will be filtered using samtools to filter reads by MAPQ > 10 and use Picard to remove PCR duplicates
Tab-delimited files (ALLC files) describing the quantity of unmethylated and methylated cytosine base calls will be generated with in-house developing package using the filtered BAM files
Assembly: mm10
Supplementary files format and content: tab-delimited files (ALLC files) describing the quantity of unmethylated and methylated cytosine base calls. Columns are: 1. Chromosome, 2. Position, 3. Strand, 4. Cytosin Context, 5. methylated base call, 6. total mapped base call, 7. not used, place holder column
Library strategy: snmC-seq2 & 3

extract_protocol:
Approximately two weeks after the AAVretro injection, brains were extracted from the 56-63 day old INTACT mice, immediately submerged in ice-cold slicing buffer (2.5mM KCl, 0.5mM CaCl2, 7mM MgCl2, 1.25mM NaH2PO4, 110mM sucrose, 10mM glucose and 25mM NaHCO3) that was bubbled with carbogen, and sliced into 0.6 mm coronal sections starting from the frontal pole. From each AAVretro-injected brain, the slices were kept in the ice-cold dissection buffer from which selected brain regions (Supplementary Table 1) were manually dissected under a fluorescent dissecting microscope (Olympus SZX16), following the Allen Mouse Common Coordinate Framework (CCF), Reference Atlas, Version 3 (2015) (Extended Data Fig. 1). The dissected brain tissues were transferred to prelabeled microcentrifuge tubes, immediately frozen in dry ice, and subsequently stored at -80oC.   For each dissected brain region, samples from 2 males and 2 females were pooled separately as biological replicates for nuclei preparation. The 2-mL glass tissue dounce homogenizer and pestles (Sigma-Aldrich D8938-1SET) were pre-chilled on ice. Nuclei were prepared using a modified protocol as reported by Lacar et al., 201622. In summary, the frozen brain tissues were transferred to the dounce homogenizer with 1 mL ice-cold NIM buffer (0.25M sucrose, 25mM KCl, 5mM MgCl2, 10mM Tris-HCl (pH7.4), 1mM DTT (Sigma 646563), 10μl of protease inhibitor (Sigma P8340)), with 0.1% Triton X-100 and 5μM Hoechst 33342 (Invitrogen H3570), and gently homogenized on ice with the pestle 10-15 times. The homogenate was transferred to pre-chilled microcentrifuge tubes and centrifuged at 1000 rcf for 8 min at 4oC to pellet the nuclei. The pellet was resuspended in 1 mL ice-cold NIM buffer, and again centrifuged at 1000 rcf for 8 min at 4oC. The pellet was then resuspended in 450 μL of ice-cold NSB buffer (0.25M sucrose, 5mM MgCl2, 10mM Tris-HCl (pH7.4), 1mM DTT, 9ul of Protease inhibitor), and filtered through 40μM cell strainer. The filtered nuclei suspension was incubated on ice for at least 30 minutes with 50μl of nuclease-free BSA for at least 10 minutes, then incubated with GFP antibody, Alexa Fluor 488 (Invitrogen, A-21311) and anti-NeuN antibody (EMD Millipore MAB377) conjugated with Alexa Fluor 647 (Invitrogen A20173). GFP+/NeuN+ single nuclei were isolated using fluorescence-activated nuclei sorting (FANS) on a BD Influx sorter with 100μm nozzle, and sorted into 384-well plates preloaded with 2μl of digestion buffer for snmC-seq (20 mL digestion buffer consists of 10 mL M-digestion buffer (2×, Zymo D5021-9), 1 ml Proteinase K (20 mg, Zymo D3001-2-20), 9 mL water, and 10 µL unmethylated lambda DNA (100 pg/µL, Promega, D1521)). The collected plates were incubated at 50oC for 20 minutes then stored at -20 oC.
The bisulfite conversion and library preparation were performed following the detailed snmC-seq2 protocol as previously described (Luo, C. et al., Nat. Commun. 2018, 9(1):3824.) or snmC-seq3 protocol described in (Liu, H. et al., Biorxiv. 2023:2023-04.). The libraries were sequenced on Illumina Novaseq 6000 using the S4 flow cell 2 x 150 bp mode.


title: 180801_CEMBA_mm_P56_P63_RS2_180801_P7_180801_P8_B9_AD012


title: ATF4mKO_22mo_2


title: 180801_CEMBA_mm_P56_P63_RS2_180801_P7_180801_P8_B8_AD010


title: rep2, plate5, spaceB1


title: 180801_CEMBA_mm_P56_P63_RS2_180801_P7_180801_P8_B8_AD008


title: Control_22mo_4

value: ATF4 flox/flox


title: rep2, plate6, spaceB1


title: Control_22mo_3


title: 180801_CEMBA_mm_P56_P63_RS2_180801_P7_180801_P8_B8_AD007


title: Control_22mo_2


title: rep1, plate3, spaceB1


title: rep2, plate4, spaceB1


title: 180801_CEMBA_mm_P56_P63_RS2_180801_P7_180801_P8_B8_AD004


title: Control_22mo_1


title: ATF4mKO_22mo_4


title: rep1, plate2, spaceB1


title: 180801_CEMBA_mm_P56_P63_RS2_180801_P7_180801_P8_B8_AD002


title: ATF4mKO_6mo_4

value: 6 months


title: 180801_CEMBA_mm_P56_P63_RS2_180801_P7_180801_P8_B9_AD001


title: rep1, plate1, spaceB1


title: 180801_CEMBA_mm_P56_P63_RS2_180801_P7_180801_P8_B8_AD012

value: female


title: 180801_CEMBA_mm_P56_P63_RS2_180801_P7_180801_P8_B7_AD012


title: ATF4mKO_6mo_3


title: rep2, plate6, spaceA9


title: ATF4mKO_6mo_2


title: ATF4mKO_6mo_1


title: rep2, plate5, spaceA9


title: 180801_CEMBA_mm_P56_P63_RS2_180801_P7_180801_P8_B7_AD007


title: 180801_CEMBA_mm_P56_P63_RS2_180801_P7_180801_P8_B7_AD010


title: Control_6mo_4


title: 180801_CEMBA_mm_P56_P63_RS2_180801_P7_180801_P8_B7_AD006


title: Control_6mo_3


title: 180801_CEMBA_mm_P56_P63_RS2_180801_P7_180801_P8_B7_AD008


title: rep1, plate3, spaceA9


title: Control_6mo_2


title: rep1, plate1, spaceA9


title: rep2, plate4, spaceA9


title: Control_6mo_1


title: rep1, plate2, spaceA9


title: 180801_CEMBA_mm_P56_P63_RS2_180801_P7_180801_P8_B7_AD002


title: rep2, plate5, spaceA8


title: 180801_CEMBA_mm_P56_P63_RS2_180801_P7_180801_P8_B7_AD004


title: rep2, plate4, spaceA8


title: 180801_CEMBA_mm_P56_P63_RS2_180801_P7_180801_P8_B7_AD001


title: 180801_CEMBA_mm_P56_P63_RS2_180801_P7_180801_P8_B6_AD012


title: rep1, plate3, spaceA8


title: 180801_CEMBA_mm_P56_P63_RS2_180801_P7_180801_P8_B6_AD010


title: FWA-1

city: Nanchang

first: Reqing

last: He

state: Jiangxi

address: No.999 Xuefu Avenue, Honggutan District

department: College of Life Science

institute: Nanchang University

data_processing:
Adaptor and low-quality sequences from raw sequencing data were removed using bbduk.sh (v38.90) with following parameters: hdist=1 mink=11 ktrim=r qtrim=rl trimq=20 minlen=25 trimpolyg=20.
Clean reads were mapped to the Arabidopsis reference genome (TAIR10) using Bismark (v0.19.1) with default settings. Context-dependent methylations were identified and extracted using “bismark_methylation_extractor” of Bismark package.
Assembly: TAIR10
Supplementary files format and content: Bedgraph file, including the methlylation density information.

organism: Arabidopsis thaliana

source_name: seedling

growth_protocol: All the plants used in this study were in the Columbia-0 ecotype (Col-0) and were grown under long day conditions (16 hours light, 8 hours dark).

extract_protocol:
DNA was extracted using a CTAB-based method
100 ng of DNA were used to make libraries using the Scale Methyl-DNA Lib Prep Kit (Illumina)

value: seedling

value: FWA-dLbCpf1-22aa-1 (+)


title: FWA-2

value: FWA-dLbCpf1-22aa-1 (-)


title: 180801_CEMBA_mm_P56_P63_RS2_180801_P7_180801_P8_B6_AD008

source_name: brain

value: brain


title: Col-1


title: rep1, plate2, spaceA8


title: rep1, plate1, spaceA8


title: CACTA-1

value: CACTA1-dAsCPF1-22a-3 (+)


title: 180801_CEMBA_mm_P56_P63_RS2_180801_P7_180801_P8_B6_AD007

value: SC

value: P


title: rep2, plate6, spaceA7

value: otic vesicle and periotic tissue

value: tdTomato+


title: 180801_CEMBA_mm_P56_P63_RS2_180801_P7_180801_P8_B6_AD006

value: STR


title: 180801_CEMBA_mm_P56_P63_RS2_180801_P7_180801_P8_B6_AD001

value: THl

value: male


title: rep2, plate5, spaceA7


title: rep1, plate3, spaceA7


title: 180801_CEMBA_mm_P56_P63_RS2_180801_P7_180801_P8_B5_AD012


title: rep2, plate4, spaceA7


title: rep1, plate1, spaceA7


title: 180801_CEMBA_mm_P56_P63_RS2_180801_P7_180801_P8_B5_AD010


title: 180801_CEMBA_mm_P56_P63_RS2_180801_P7_180801_P8_B5_AD008


title: 180801_CEMBA_mm_P56_P63_RS2_180801_P7_180801_P8_B5_AD007


title: CACTA-2

value: CACTA1-dAsCPF1-22a-3 (-)


title: rep2, plate6, spaceA6


title: rep1, plate3, spaceA6


title: rep2, plate5, spaceA6


title: 180801_CEMBA_mm_P56_P63_RS2_180801_P7_180801_P8_B5_AD006

value: THp


title: 180801_CEMBA_mm_P56_P63_RS2_180801_P7_180801_P8_B5_AD002


title: 180801_CEMBA_mm_P56_P63_RS2_180801_P7_180801_P8_B5_AD001


title: rep2, plate6, spaceA5


title: rep1, plate1, spaceA6


title: 180801_CEMBA_mm_P56_P63_RS2_180801_P7_180801_P8_B4_AD012


title: 180801_CEMBA_mm_P56_P63_RS2_180801_P7_180801_P8_B4_AD010


title: rep2, plate5, spaceA5


title: 180801_CEMBA_mm_P56_P63_RS2_180801_P7_180801_P8_B5_AD004


title: 180801_CEMBA_mm_P56_P63_RS2_180801_P7_180801_P8_B4_AD008


title: rep2, plate4, spaceA5


title: 180801_CEMBA_mm_P56_P63_RS2_180801_P7_180801_P8_B4_AD007


title: rep1, plate2, spaceA5


title: KG1 Xenograft Tumor IMiD-activated MSC Treated_S10

type: RNA

city: Navi Mumbai

first: Jyoti

middle: Anand

last: Kode

state: Maharashtra

address: Kharghar

department: Kode Lab

country: India

institute: Advanced Centre for Treatment, Research & Education in Cancer (ACTREC), Tata Memorial Centre

description: IMiD treated Bilineage leukemia MSC- treated subcutaneous KG1 tumor

hyb_protocol: Fragmentation of labeled cRNA and hybridization were done using the Gene Expression Hybridization kit of (Agilent Technologies, In situ Hybridization kit, Part Number 5190-0404). Hybridization was carried out in Agilent’s Surehyb Chambers at 65º C for 16 hours. The hybridized slides were washed using Agilent Gene Expression wash buffers (Agilent Technologies, Part Number 5188-5327)

scan_protocol: Scanned using Agilent Microarray Scanner (Agilent Technologies, Part Number G2600D).

data_processing: Data extraction from Images was done using Feature Extraction software  and Normalization of the data was done in GeneSpring GX  : 75th percentile shift  method and gene based analysis was performed

source_name: KG1 human-mouse xenograft

label_protocol: The samples for Gene expression were labeled using Agilent Quick-Amp labeling Kit (p/n5190-0442). The total RNA were reverse transcribed at 40°C using oligo dT primer tagged to a T7 polymerase promoter and converted to double stranded cDNA. Synthesized double stranded cDNA were used as template for cRNA generation. cRNA was generated by in vitro transcription and the dye Cy3 CTP(Agilent) was incorporated during this step. The cDNA synthesis and in vitro transcription steps were carried out at 40°C. Labeled cRNA was cleaned up using Qiagen RNeasy columns (Qiagen, Cat No: 74106) and quality assessed for yields and specific activity using the Nanodrop ND-1000.

growth_protocol: BM from Bi-lineage leukemia (ETP-ALL) patient was treated with Stem Cell Enrichment cocktail and MSC were enriched by separation on FH gradient. Enriched cells were seeded in culture flasks with IMDM media supplemented with 10% FBS and 1% antibiotics and glutamine. Adherent fibroblastoid cells were expanded upto passage 4 and 5. At passage P3 or P4, MSC cells were trypsinized, washed with PBS and cell pellet was used to treat KG1 xenograft tumor. Conditioned media from confluent culture cells was stored separately and stored in aliquots at -20°C. For immunomodulation set, BM-MSC cells were treated for 24 hours with IMiD reagent containing anti-TLR3 antibody + TLR4 ligand LPS + CXCR4 antagonist peptide (Gift from Kyoto University, Japan).

extract_protocol: RNA was extracted using Qiagen’s RNeasy minikit as per the manufacturer’s protocol

treatment_protocol: AML cell line KG1 was used to generate subcutaneous xenograft tumor in NOD-SCID mice. Once tumor was generated, it was passaged in another set of mice to stabilize it. This donor mice were used to prepare tumor bearing mice (8-10 week old mice of average 20gm body weight). Once tumor size gains 2-3 mm size, Adriamycin group mice were treated i.v. with ADR 2.5 mg/kg body weight on 1,5 and 9th days. MSC-treated group was treated with BM MSC 0.5 million cells intratumorally in 25µl on day 0 and day 15. MSC Conditioned media-treated group was treated with 200 µl supernatants every alternate day, intratumorally. MSC-IMiD treated group was treated with BM MSC (priorly stimulated with IMiD) 0.5 million cells intratumorally in 25µl on day 0 and day 15. Mice were observed for behavior, toxicity and reduction in tumor growth up to 30 days. Subcutaneously grown KG1 derived tumors were isolated harvested from mice and piece from representative mice were stored in RNA later for gene expression analysis.

value: Human-Mouse xenograft

tag: donor

value: Mouse # 1729

tag: tumor details

value: Tumor size:length 11.68 mm, width:11.39 mm; Relative Tumor Volume at 31 day: 15.77


title: MSC_S9

description: AML10 MSC P4

source_name: MSC cultured from Bone Marrow

growth_protocol: Bone marrow (BM) from cancer patients (AML/ ETP-ALL/ Lymphoma) was divided in two parts. One part from BM (AML/ ETP-ALL) was processed using ficoll-hypaque (FH) density gradient centrifugation to obtain leukemic blast population. Leukemic cells obtained were stored in RNAlater at -20°C. Second part of BM (AML/ ETP-ALL/ Lymphoma) was treated with Stem Cell Enrichment cocktail and MSC were enriched by separation on FH gradient. Enriched cells were seeded in culture flasks with DMEM media supplemented with 10% FBS and 1% antibiotics and glutamine. Adherent fibroblastoid cells were expanded upto passage P5.

treatment_protocol: Leukemic cells were washed in PBS and pelleted. The cell pellet was suspended in RNA 200-500ul RNAlater and stored at -200C. AML cell line was cultured and store in RNAlater. MSC culture at 80% confluency was trypisinized, washed with PBS and cell pellet was stored in 200-500µl of RNAlater at -20°C.

value: Bone marrow

value: AML patient

tag: cell details

value: Cultured AML MSC at passage P4


title: KG1 Xenograft Tumor MSC culture supt Treated_S10

description: Bilineage leukemia 02 MSC supernatant treated subcutaneous KG1 tumor

value: Mouse # 1790

value: Tumor size:length 11.19 mm, width:10.48 mm; Relative Tumor Volume at 31 day:9.13


title: KG1 Xenograft Tumor MSC Treated_S10

description: Bilineage leukemia 02 MSC treated subcutaneous KG1 tumor

value: Mouse # 1764

value: Tumor size:length 12.87 mm, width:12.31 mm; Relative Tumor Volume at 31 day:9.01


title: rep1, plate2, spaceA6

value: otic vesicle and periotic tissue


title: rep1, plate2, spaceB4

value: tdTomato+


title: KG1 Xenograft Tumor Control Untreated_S10

description: Subcutaneously grown KG1 tumor in NOD-SCID mice

value: Mouse # 1763

value: Tumor size:length 15.97 mm, width:18.8 mm; Relative Tumor Volume at 31 day: 21.52


title: MSC_S6

description: Lymphoma 01 Uninvolved BM MSC P4

value: Lymphoma patient

value: Cultured uninvolved BM MSC at passage P4


title: KG1 Xenograft Tumor ADR Treated_S10

description: Adriamycin treated Subcutaneous KG1 tumor

value: Mouse # 1746

value: Tumor size:length 13.78 mm, width:12.78 mm; Relative Tumor Volume at 31 day:12.29


title: rep1, plate1, spaceA5


title: 180801_CEMBA_mm_P56_P63_RS2_180801_P7_180801_P8_B4_AD006

first: Joseph

institute: HHMI-Salk-Institute

value: brain

value: THp

value: STR

value: female


title: BMMNC_S8

description: Bilineage Leukemia (ETP-ALL) 02 blasts

source_name: Bone marrow derived mononuclear cells

value: Bone marrow

value: Bi-lineage leukemia patient

value: Bi-lineage Leukema blasts


title: MSC_S2

description: AML07 MSC P3

value: AML patient

value: Cultured AML MSC at passage P3


title: KG1_S7

description: KG1 cell line

source_name: AML Cell line

value: AML Cell line


title: BMMNC_S3

description: AML08 Leukemic blasts

value: AML Leukemic blasts


title: rep2, plate6, spaceA4

value: otic vesicle and periotic tissue

value: tdTomato+


title: rep2, plate5, spaceA4


title: 180801_CEMBA_mm_P56_P63_RS2_180801_P7_180801_P8_B4_AD002

value: THl


title: 41BB CAR, day 15, scRNAseq

data_processing:
The demultiplexing, barcoded processing, gene counting and aggregation were made using the Cell Ranger software v2.1.1 (https://support.10xgenomics.com/single-cell-gene-expression/software/pipelines/latest/what-is-cell-ranger)
Downstream single-cell analysis was performed using Seurat package v4.0.5 within the R programming environment v4.1.2.16 Lower bound for the number of genes in individual cells was chosen based on binary logarithm distribution and was set to 9.8. Cells with more than 7,500 genes were filtered out.
The percentage of mitochondrial counts was calculated for every cell, and only cells with mitochondrial percentage less than 10% were used in further analysis. Filtered matrices were normalized using a scaling factor of 10,000 and centered.
Assembly: hg38
Supplementary files format and content: Tab-separated values files and matrix files

source_name: Engineered T cells

extract_protocol:
CAR T cells from chronic stimulation cultures were isolated using flow-based sorting.
Cells were processed using the 10x Genomics Chromium Single Cell V(D)J Reagent Kits (10x Genomics, PN-1000006, PN-1000020, PN-120262) to generate single-cell emulsions for barcoding, reverse transcription and cDNA amplification. Immediately following these steps, 10x barcoded fragments were pooled and attached to standard Illumina adaptors to generate a barcoded single-cell RNA library.

value: CD19-directed human CAR T cells containing 41BB

tag: treatment

value: Co-culture with Nalm6 cells

tag: time

value: day 15


title: 180801_CEMBA_mm_P56_P63_RS2_180801_P7_180801_P8_B3_AD010

tag: tissue

value: brain

tag: source

value: SC

value: P


title: 180801_CEMBA_mm_P56_P63_RS2_180801_P7_180801_P8_B3_AD008

tag: target

tag: Sex

value: male


title: rep1, plate2, spaceA4

tag: developmental stage


title: BMMNC_S1

description: AML06 Leukemic blasts

value: Bone marrow


title: 41BB CAR, day 7, scRNAseq

value: day 7


title: CD28 CAR, day 7, scRNAseq

value: CD19-directed human CAR T cells containing CD28


title: rep1, plate1, spaceA4

tag: genotype