title: Bone marrow, 5,000 cells

city: Philadelphia

first: Zainul

last: Hasanali

address: 3620 Hamilton Walk, 230 John Morgan Building

institute: University of Pennsylvania

description: 5000 cells analyzed at equimolar ratio to 10k sample for double read depth

library_source: transcriptomic single cell

data_processing:
Parse Biosciences Pipeline v2.0
Assembly: hg38, mm10
Supplementary files format and content: matrix and csv files
Supplementary files format and content: Analysis performed by combining data from 10k and 5k: combined*

organism:
Homo sapiens
Mus musculus

source_name: bone marrow

extract_protocol: Parse Biosciences Evercode Whole Transcriptome Mini Kit

value: bone marrow


title: RENCA Parental rep 2

status: Public on Oct 14 2024

type: SRA

city: Buffalo

first: John

last: Ebos

state: NY

address: Elm & Carlton Streets

department: Department of Cancer Genetics and Genomics

country: USA

institute: Roswell Park Cancer Institute

library_source: transcriptomic

data_processing:
Pre-alignment quality control was performed using FASTQC v0.11.5.
Sequenced reads (fastq files) were alinged to the mouse genome GRCm38 M16 (Gencode) using STAR v2.6.0a.
Post-alignment quality control was performed using RSeQC v2.6.5.
Estimated counts at the gene level were quantified using RSEM v1.3.1.
Genome_build: GRCm38
Supplementary_files_format_and_content: tab-delimited text file include RSEM estimated counts at the gene level for all samples

molecule: polyA RNA

organism: Mus musculus

source_name: Kidney cancer cells

growth_protocol: Cells were maintained in RPMI supplemented with 5% FBS with 0.5μg/ml of isotype control or anti-PD-L1. Growth media was refreshed every 3-4 days.

extract_protocol:
Total RNA from individual samples was extracted using the RNeasy mini kit (QIAGEN; 74104). Genomic DNA was digested using DNase I (QIAGEN; 79254) as described in the on-column DNase digestion protocol from QIAGEN.
One microgram total RNA was used for library preparation using the TruSeq Stranded mRNA kit (Illumina Inc) following manufacturer's instructions.

tag: variant

value: RENCA

tag: treatment

value: Isotype Control

tag: dose

tag: treatment duration

value: >4 weeks


title: RENCA Parental met rep 1


title: Bone marrow, 10,000 cells

state: PA

description: 10,000 cells analyzed

molecule: total RNA

tag: tissue


title: H3.3CtrlH4K16R, Rep4

city: Chapel Hill

first: Greg

last: Matera

state: NC

address: 3352 Genome Sciences Building

institute: UNC Chapel Hill

data_processing:
Reads were quality trimmed using bbmap v38 bbduk qtrim=rl trimq=20
Reads were aligned using STAR v.2.7.2a to DM6.38
Raw count table was generated using subread v.2.0.3 using DM6.38 with the following parameters: -g gene_id -p -s 1
Assembly: DM6
Supplementary files format and content: Tab-delimited text file includes raw counts for each sample.

organism: Drosophila melanogaster

source_name: Whole L1, mixed sex

growth_protocol: Embryos from crosses kept at 25 degrees C, were laid on grape-juice agar plates and harvested for RNA collection at the L1 stage.

extract_protocol:
25-30 larvae were collected in 1ml Trizol. RNA was isolated from the aqueous phase using the Zymo Clean and Concentrator Kit (with DNASEI digestion).
Total RNA Seq libraries were prepared with Nugen Ovation Universal Drosophila kit

value: Whole L1

tag: Sex

value: pooled male and female

tag: genotype


title: H3.3CtrlH4K16R, Rep3


title: H3.3K36RH3.2K36R, Rep3


title: ATAC-seq_Oct4_KO_LateICM_3

city: Muenster

first: Yanlin

last: Hou

address: Röntgenstrasse 20

country: Germany

institute: Max Planck Institute for Molecular Biomedicine

description:
Oct4_KO_LateICM_3
Accessible chromatin regions

library_source: genomic

data_processing:
Sequence reads were trimmed for adapter sequences using SeqPurge and trimmed reads no shorter than 20 bases were aligned to the mm10 mouse reference genome using Bowtie2 with a maximum fragment size of 2000. Duplicated reads were removed using Picard MarkDuplicates (https://broadinstitute.github.io/picard/) and only reads uniquely mapped to the standard chromosomes except chrY and chrM (mitochondrial genome) with mapping quality of at least 30 were used for the following analysis. Reads on the positive and negative strands were shifted +4 bp and −5 bp, respectively. Peak calling was performed using MACS2 with the following parameters: --keep-dup all -g mm --nomodel --shift -50 --extsize 100 -B --SPMR. Peaks that overlap with the blacklisted regions (https://sites.google.com/site/anshulkundaje/projects/blacklists; https://sites.google.com/site/atacseqpublic/atac-seq-analysis-methods/mitochondrialblacklists-1) and satellite repeats (RepeatMasker) were removed. Significant peak summits (q-value ≤0.001) from biological replicates were merged for each sample group using BEDTools with a maximum distance between two summits to be merged of 200 bp. Only merged peak summits that overlap with summits from at least two replicates were retained. Reads per million (RPM)-normalized pileup signals in the bedGraph format were converted into bigWig files using the UCSC bedGraphToBigWig tool. The bigWig files for the average signal of biological replicates were generated using the UCSC bigWigMerge tool. The heatmaps of RPM-normalized pileup signals around the TSSs of GENCODE (vM23) protein-coding genes were generated using deepTools.
For count-based analysis of transposon insertion events, merged peak regions were generated by combining the adjacent peak summits of all embryo groups and selecting 200-bp regions around the centers of each merged summit. 5’ ends of shifted read alignment on both the positive and negative strands were considered as transposon insertion sites. The number of transposon insertions in each merged peak region were counted for each sample using BEDTools and normalized and transformed to log2 scale using the rlog function of the Bioconductor package DESeq2 and significantly differential peaks were identified with an adjusted p-value cutoff of 0.05. Peaks located within 100 bp from TSSs of GENCODE (vM23) genes were considered as TSS-proximal peaks. The presence of TF binding motifs in merged peaks was investigated using FIMO with the default cutoff (p-value <10-4).
Assembly: mm10
Supplementary files format and content: bigwig
Supplementary files format and content: rlog of read count

molecule: genomic DNA

source_name: E4.5 late ICM

extract_protocol:
All the embryos for ATAC-seq were collected from female mice superovulated with PMSG and hCG. Early and late morula samples were collected at E2.75 and E3.25, respectively. Early and late ICM samples were collected at E2.5 and cultured in KSOM with MEKi (PD0325901, 1 μM) until early and late blastocyst stages, respectively. ICM were isolated by immunosurgery.
20 to 40 morulae or ICMs were pooled and lysed in the lysis buffer (10 mM Tris·HCl, pH=7.4; 10 mM NaCl; 3 mM MgCl2; 0.15% Igepal CA-630; 0.1% Tween-20) for 10 min on ice. We added 0.1% Tween-20 to the lysis buffer as it greatly reduced reads mapped to mitochondrial DNA. Lyzed embryos were briefly washed, collected in 2.0 μl PBS containing 0.1 mg/ml polyvinyl alcohol (PVA) and transferred to 3.0 μl Tn5 transposome mixture (0.25 μl Tn5 transposome, 2.5 μl tagmentation buffer, Illumina, FC-121-1030; 0.25 μl H2O). The samples were incubated in 37 °C water bath for 30 min. Tagmentation was stopped by adding 2.0 μl 175 mM EDTA and incubated at 50 °C for 30 min. Excess EDTA was quenched by 2.0 μl 160 mM MgCl2. The libraries were amplified in the following reaction: 9.0 μl Transposed DNA, 10.0 μl NEBNext High-Fidelity 2×PCR Master Mix (New England Biolabs, M0541S), 0.25 μl 100 μM PCR Index 1, 0.25 μl 100 μM PCR Index 2 and 0.5 μl H2O. The sequences of index 1 and 2 are in Table EV2. The PCR program is as the following: 72 °C for 5 min; 98 °C for 30s; 16 thermocycles at 98 °C for 10 s, 63 °C for 30 s and 72 °C for 1 min; followed by 72 °C 5 min. Amplified libraries were purified twice with 1.2 × AMPure XP beads. The sequencing was performed on the NextSeq 500 system with pair-end 75bp.

value: E4.5 late ICM

tag: cell line

tag: cell type

value: 1 μm PD0325901, from E2.5 to ICM isolation


title: H3.3K36RH3.2HWT, Rep4


title: Set21, Rep3


title: H3.3K36RH3.2K36R, Rep4


title: H3.3CtrlH3.2HWT, Rep3


title: H3.3CtrlH4K16R, Rep2


title: H3.3K36RH3.2K36R, Rep1


title: H3.3CtrlH3.2HWT, Rep4


title: H3.3CtrlH3.2HWT, Rep2


title: Set21, Female, Rep3

data_processing:
Reads were quality trimmed using bbmap v38 bbduk qtrim=rl trimq=20
Reads were aligned using STAR v.2.7.2a to DM6.38
Raw count table was generated using subread v.2.0.3 using DM6.38 with the following parameters: -g gene_id -p -s 2
Assembly: DM6
Supplementary files format and content: Tab-delimited text file includes raw counts for each sample.

source_name: Wandering L3 brain

growth_protocol: Larvae were seeded 50 per vial into molasses-cornmeal food, and raised at 25oC

extract_protocol:
Samples were collected in 1ml Trizol. RNA was isolated from the aqueous phase using the Zymo Clean and Concentrator Kit (with DNASEI digestion).
PolyA-selected libraries were prepared using the KAPA stranded mRNA kit

value: Wandering L3 brain

value: female


title: Set21, Female, Rep2


title: H3.3CtrlH3.2HWT, Rep1


title: Set21, Rep2


title: Set21, Rep4


title: yw, Rep2

value: yw


title: yw, Rep3


title: Set21, Rep1


title: yw, Male, Rep3

value: male


title: yw, Female, Rep2


title: yw, Male, Rep1


title: RNA from skin-infiltrating myeloid cells from Tyk2-/- mice_3

city: Vienna

first: Birgit

last: Strobl

address: Veterinärplatz 1

country: Austria

institute: Veterinary University Vienna

data_processing:
Sequencing reads were aligned to the GRCm39 reference genome using STAR (v2.7.3a) with Ensembl annotation (v105). Read counts were assigned to genomic features using featureCounts (v2.0.0).
Assembly: mm39 (GRCm39 Ensembl release 105).
Supplementary files format and content: Tab-delimited text file with raw counts for each sample.

source_name: skin

extract_protocol:
Total RNA was extracted using RNeasy Plus Micro Kit (74034; Qiagen) according manufacturer’s instructions. 3,8 - 12,7 ng of total RNA was used for the construction of sequencing libraries
For the preparation of NGS libraries from low-input samples we followed the Smart-seq2 protocol. The subsequent library preparation from the amplified cDNA was performed using the Nextera XT DNA library prep kit (Illumina). Library concentrations were quantified with the Qubit 2.0 Fluorometric Quantitation system (Life Technologies) and the size distribution was assessed using the 2100 Bioanalyzer instrument (Agilent). For sequencing, samples were diluted and pooled into NGS libraries in equimolar amounts.

treatment_protocol: 30 000 – 70 000 skin-infiltrating myeloid cells (CD45+CD3-CD19-NK1.1-) were sorted using BD FACS Melody into 350 uL RLT buffer supplemented with β-mercaptoethanol and stored at -80 °C prior to RNA isolation.

value: skin

tag: age

value: 9 weeks

tag: time-point

value: day 1 p.i


title: RNA from skin-infiltrating myeloid cells from Tyk2-/- mice_2


title: RNA from skin-infiltrating myeloid cells from Tyk2-K923E mice_3


title: RNA from skin-infiltrating myeloid cells from Tyk2-/- mice_1


title: yw, Rep4


title: Set21, Male, Rep3


title: Set21, Male, Rep2


title: H3.3K36RH3.2HWT, Male, Rep3


title: H3.3K36RH3.2HWT, Male, Rep2


title: H3.3CtrlH4K16R, Rep1


title: yw, Female, Rep3


title: Set21, Male, Rep1


title: Set21, Female, Rep1


title: RNA from skin-infiltrating myeloid cells from Tyk2-K923E mice_2


title: H3.3K36RH3.2HWT, Male, Rep1


title: H3.3K36RH3.2HWT, Female, Rep3


title: RNA from skin-infiltrating myeloid cells from WT mice_3

value: WT


title: H3.3K36RH3.2HWT, Female, Rep2


title: H3.3K36RH3.2HWT, Female, Rep1


title: RNA from skin-infiltrating myeloid cells from Tyk2-K923E mice_1


title: RNA from skin-infiltrating myeloid cells from WT mice_2


title: yw, Female, Rep1


title: H3.3CtrlH3.2HWT, Male, Rep2


title: H3.3CtrlH3.2HWT, Male, Rep3


title: H3.3CtrlH3.2HWT, Male, Rep1


title: RNA from skin-infiltrating myeloid cells from WT mice_1


title: H3.3CtrlH3.2HWT, Female, Rep3


title: H3.3CtrlH3.2HWT, Female, Rep1


title: yw, Male, Rep2


title: H3.3WTH3.2K36R, Male, Rep3


title: H3.3CtrlH3.2HWT, Female, Rep2


title: H3.3WTH3.2K36R, Male, Rep1


title: H3.3WTH3.2K36R, Male, Rep2


title: H3.3WTH3.2K36R, Female, Rep2


title: H3.3WTH3.2K36R, Female, Rep3


title: H3.3WTH3.2HWT, Male, Rep3


title: H3.3WTH3.2K36R, Female, Rep1


title: HeLa total RNA, Borohydride reduction, Marathon

city: New York

first: Samie

last: Jaffrey

state: New York

address: 1300 York Ave

department: Pharmacology

institute: Weill Cornell Medicine

description:
Duration: 0;temperature: N/A;buffer: N/A;ligand: N/A
18s Sequencing data 2

data_processing:
Sequencing data was processed using ShapeMapperV2.2 (https://github.com/Weeks-UNC/shapemapper2)
For HeLa 18s rRNA experiments, sequencing data was aligned to 12 unique 18S amplicons defined by the unique combinations of 4-nucleotide forward and 7-nucleotide reverse barcode sequences inserted during step 1 PCR. Sequencing data alignment was performed with ShapeMapperV2.2 with the following settings: --amplicon --output-parsed --output-aligned-reads --nproc 15 --output-counted-mutations --min-mutation-separation 0.
For Spinach, pUG, and AKT2 experiments, sequencing data was aligned to a single fasta sequence with ShapeMapperV2.2 with the following settings: --amplicon --dms --output-parsed - -min-mutation-separation 2. Deletions are ambiguously aligned and therefore ignored by invoking the --dms option.
Misincorporation signatures were calculated by aligning the data with ShapeMapperV2.2 with the following settings: --amplicon --output-counted-mutations --min-mutation-separation 0.
Counts of misincorporation types for each position within an RNA are tabulated by the --output-counted-mutations field. A minimum misincorporation separation distance of 0 was chosen to capture the full range of misincorporation types generated in BASH MaP and DMS MaP experiments.
ShapeMapper alignment with --output-parsed produces a .mut file which encodes the misincorporations within a sequencing read as a bitvector where 0 represents no misincorporation and 1 represents a misincorporation. This .mut file was then input into the program RingMapper which identifies positions with statistically high rates of co-occurring misincorporations7. The output of RingMapper was processed with the custom python script ring_pair_to_heatmap.py which filters all negative correlations and creates a square matrix which was then visualized in Prism GraphPad.
For M2 Spinach and pUG experiments, sequencing data was aligned to a single fasta sequence with ShapeMapperV2.2 with the following settings: --amplicon --dms --output-parsed --min- mutation-separation 2. The output parsed mutation file was then processed with the custom script mut_to_simple.py which retains only sequencing reads that cover the entire length of the RNA. Then the output .simple file was used to calculate a unique mutational profile for each point mutant with the script simple_to_M2_map.py. This script produces a square matrix with length equal to the length of the RNA and displays how often two mutations co-occur on the same sequencing read. To better visualize changes in nucleotide reactivity given a mutation at a certain position, Z-score normalization was performed with the script M2_Map_to_Zscores.py.
The normalized matrix was then visualized in Prism GraphPad as a heatmap.
RingMapper was run with default parameters and the output file was filtered to include only positive correlations between two G nucleotides. A second filter removed all correlations with Z-scores less than 2.0. Networks of structurally related nucleotides were visualized in Cytoscape with unique G bases represented as nodes and positive correlations represented as edges.
Minimum free energy RNA secondary structure modeling:
The baseline secondary structure model of Spinach was generated in mFold with default parameters. Population average DMS guided RNA secondary structure modeling was performed as described previously in the DANCE pipeline. Briefly, the python script DanceMapper.py was run on parsed mutation output files with the following settings for deriving population average data: --fit --maxc 1. Then, the population average reactivities were converted to free energy restraints for the RNA folding program RNAstructure through the foldClusters.py script. When processing BASH MaP data, the additional command --nog was used with foldClusters.py to ignore the reactivity data for G nucleotides. The resulting RNA secondary structures were visualized in RNA2D drawer. Mapping of per nucleotide DMS reactivity onto RNA2D drawer secondary models was performed through the custom python script color_reactivities_RNA2D_drawer.py. Conformation specific RNA secondary structure modeling was performed by increasing the maximum allowed conformations from 1 to 5 with -- maxc 5.
Single molecule probabilistic RNA secondary structure modeling:
A custom pipeline was developed to apply the DaVinci structural analysis method to BASH MaP56. This analysis method utilizes misincorporations to generate a folding constraint for each sequencing read. Folding constraints are then passed to ContraFold which folds each sequencing read as a unique RNA molecule. The complete analysis pipeline is as follows:
First, sequencing data is processed through ShapeMapper as described above with --amplicon --dms --output-parsed --min-mutation-separation 2 options enabled. The resulting .mut file is then filtered and converted to a .simple file with mut_to_simple.py. For DMS MaP experiments, the resulting .simple file is converted into a .bit file by running the simple_to_bit.py python script. For BASH MaP experiments, the resulting .simple file is instead converted into a .bit file with the script updated_simple_to_bit_rG42.py. This script requires an input fasta file and sets all upper-case G positions to be considered for base pairing. In addition, the script constrains all lowercase letters to be single stranded in the .bit file. If a sequencing read harbors a misincorporations at the position of one of the lowercase letters, then all lowercase letters in that sequencing read are instead considered for base pairing. The resulting .bit file is then passed as an argument to the fold-contrafold-uniq-bits-vectors2.py script. ContraFold was run either with normal settings or with --noncomplementary enabled. Typically, the fold-contrafold-uniq-bits-vectors2.py script is stopped after roughly 10,000 RNA molecules have been folded to save on computation time. The forgi vector encodings of the RNA secondary structures generated by ContraFold are output in the file forgi-vect-ser2.txt. Dimensional reduction is performed on the forgi vectors text file with the python script run-pca-on-forgi-vectors.py which reduces the vector space to 2 dimensions. Kmeans clustering is then performed on the 2-dimensional representation of RNA structures with the draw-kmeans-clusters.py script. Identification of a population average structure is done by choosing --num_clusters 1. Multiple conformational analysis is performed by varying the number of Kmeans clusters. The structure of the most representative of the Kmeans cluster is retrieved with the python script fetch_DaVinci_folded_copy.py and visualized with RNA 2D drawer.
Nucleotide reactivity normalization:
Mutation rates were normalized to reactivity values as previously described with the custom python script normalize_DAGGER_DANCE.py.
Tertiary folding constraint determination and implementation (DAGGER):
G’s likely to be engaged in tertiary interactions were identified by first selecting G’s in the bottom quartile for misincorporation rate. In situations when reactivity data was available for multiple conformations, G’s in separate conformations were treated as unique G’s and all unique G’s were pooled for reactivity normalization. From the bottom quartile of reactive G’s, G’s which displayed correlations to other lowly reactive G’s as identified by RingMapper were designated as engaged in a tertiary interaction.
The DaVinci analysis pipeline was modified to utilize tertiary constraints through setting tertiary G positions to lowercase G’s in the fasta input file for generation of .bit files by updated_simple_to_bit_rG42.py. The modified DaVinci analysis pipeline is reffered to as DAGGER.
All python scripts for analyzing data can be found at: https://github.com/SamieJaffreyLab/DAGGER_MaP
Assembly: custom fasta files for each RNA construct
Supplementary files format and content: Shapemapper output files include mutation rates for each nucelotide as output by ShapeMapper2. A complete description of the ShapeMapper output is available at (https://github.com/Weeks-UNC/shapemapper2). Mutate and MaP experiments processed files include a Z-score normalized mutate and map heatmap. See https://github.com/SamieJaffreyLab/DAGGER_MaP for detailed description of the heatmap file formats.
Library strategy: Amplicon sequencing

organism: Homo sapiens

source_name: cell line

growth_protocol: HeLa cells were grown in DMEM containing 10% FBS with 100 U ml−1penicillin and 100 µg ml−1streptomycin under standard culturing conditions (37°C with 5.0% CO2). SH-SY5Y cells were grown in a 1:1 ratio of DMEM to F12 media containing 10% FBS with 100 U ml−1penicillin and 100 µg ml−1streptomycin under standard culturing conditions (37°C with 5.0% CO2).

extract_protocol:
In vitro transcribed RNA was purified from the quenched DMS reaction with RNA Clean and Concentrator 5 kit and eluted in 10µL water. RNA was either directly processed or stored at -80°C
We noticed that DMS probing caused a large portion of adherent cells to become detached from the plate. We therefore decanted the quenched DMS probing media off the plate into a 15 mL falcon tube and pelleted the detached cells for 5 min at 4C and 1,000 x g. To isolate the total RNA, 2 mL Trizol reagent was added to the plate for the cells which had not become detached. Then, the 2 mL Trizol reagent was transferred to solubilize the pelleted cells which had become detached during DMS probing. Total RNA was then isolated according to manufacturer’s instructions.
Sequencing library preparation:
Reverse transcription
Superscript II: Reverse transcription with SSII was performed as follows. 200 ng of input RNA was mixed with 1µL 50µM random hexamer (HeLa total RNA) or 2µL of 1µM gene specific reverse transcription primer (in vitro transcribed RNAs) and annealed. Annealed RNA was added to reverse transcription master mix containing 6 mM final concentration of MnCl2. Reverse transcription reactions were incubated at 23°C for 15 min only when using random hexamers then 42°C for 3 hours.
Marathon RT: Reverse transcription with Marathon RT was performed with a modified protocol. First, 200 ng of input RNA was mixed with 2µL10 mM dNTP and 1µL 50µM random hexamer or 2µL of 1µM gene specific RT primer. The volume was brough up to 8µL and the RNA was annealed to the primer with the following thermocycler settings: 80°C for 1 min, 65°C for 5 min at which point the RNA was placed on ice for 2 min. Then, an RT master mix was created containing 4µL of 5x Marathon MaP buffer (250 mM Tris-HCl pH 7.5, 1 M NaCl), 4µLof 100% glycerol, 1µL of 100 mM DTT, 0.5µL of 20 mM MnCl2, 0.5µL of water. Next, 10µL of RT master mix was added to 8µL of annealed RNA followed by the addition of 2µL Marathon RT. Reverse transcription reactions were incubated at 23°C for 15 min only when using random hexamers then 42°C for 3 hours.
Reverse transcription reactions were inactivated by incubating at 75°C for 10 min. cDNA was purified with DNA clean and concentrator 5 columns by using 7 volumes of DNA binding buffer (140 µL) and eluted in 10µL water.
Sequencing libraries were generated via a 2 step PCR. Briefly, 1/5th of purified cDNA (2 µL) was amplified with step 1 forward and reverse primers. HeLa 18s rRNA samples included a 4-nucleotide barcode on the forward primer and a 7-nucleotide barcode on the reverse primer which enabled pooling of purified step 1 PCR products. AKT2 step 1 PCR primers were designed to amplify a ∼200 nucleotide segment of the AKT2 3’UTR. In vitro transcribed RNAs used non-barcoded step 1 forward and reverse primers which were specific to flanking sequences introduced into the RNA constructs. These flanking sequences are referred to as a Structure Cassette (SC). Step 1 PCR reactions were assembled on ice by adding to 2µL purified cDNA: 1.25µL of 10µM forward primer, 1.25µL of 10µM reverse primer, 9µL of water and 12.5µL of 2x Phusion HF master mix or 2x Phusion GC master mix for AKT2 step 1 PCR. Step 1 PCR reactions were run with the following thermocycler settings: 98°C for 2 min followed by 8-28 cycles of 98°C for 10 seconds, 65°C for 20 seconds, 72°C for 20 seconds, followed by a final extension of 2 min. PCR cycle number was optimized for each experiment separately. PCR reactions were purified with 1.8x AMPure XP beads according to manufactures instructions. Purified DNA was eluted in 25µL water.
Step 2 PCR was performed with NEB Next Multiplex Oligos for Illumina. Step 2 PCR reactions were composed of the following: 2-10 ng of purified step 1 DNA brough up to 12µL water, 4µL of i5 index primer or universal primer, 4µL i7 index primer, and 20µL of 2x Phusion HF or 2x Phusion GC for AKT2 library prep. Step 2 PCR reactions were run with the following thermocycler settings: 98°C for 2 min followed by 6-12 cycles of 98°C for 15 seconds, 65°C for 30 seconds, 72°C for 30 seconds, then 72°C for 5 min. PCR cycle number was optimized for each experiment separately. Step 2 PCR reactions were size selected with AMPure XP beads, eluted in 25µL of water, and quantified with Qubit 1x dsDNA HS kit. Libraries were pooled and sequenced on the NovaSeq 6000 PE 2x100, NovaSeq 6000 PE 2x150, MiSeq PE 2x150, Nextseq2000 P1 600 cycles, or MiSeq Micro 300 cycles.
Gene-specific BASH MaP:
Total RNA was treated with DNAse I for 30 min prior to BASH MaP treatment according to manufacturer’s instructions. Three replicates of 1 µg of total RNA was mixed with 1µL 50µM random hexamers and the volume was adjusted to 11 µL. RNA was annealed by heating to 85°C for 1 min, 65°C for 5 min, then placing on ice for 3 min. Reverse transcription was performed with SuperScript II as described previously8. Reverse transcription reactions were incubated at 23°C for 15 min and then 42°C for 3 hours. Reactions were inactivated by heating to 75°C for 10 min. Reverse transcription replicates were pooled and the cDNA was purified with the Zymo DNA Clean and Concentrator 5 kit and eluted in 10µL water.

treatment_protocol:
DMS modification of in vitro transcribed RNA
In vitro transcribed RNA was probed as follows. 200 ng – 1 µg of RNA was folded in a buffer designed to promote modification of A and C bases only (200 mM Bicine pH 7.75 at room temperature, 100 mM KCl) or a buffer which promotes the additional modification of m1G and m3U (200 mM Bicine pH 8.37 at room temperature, 100 mM KCl). RNA was refolded in buffer by heating to 95°C for 3 min then snap cooling on ice for 1 min. Then MgCl2 was added to a final concentration of 1-5 mM and the RNA was left at room temperature for 10 min. Neat DMS was diluted to 1.7M with anhydrous ethanol and added to the folded RNA. DMS reactions was performed for 10 min at room temperature or 6 min at 37°C. Spinach was probed at room temperature due to its thermal instability. pUG RNA was probed at 37°C. Reactions were quenched by the addition of 4 volumes of 20% beta-mercaptoethanol in water on ice. RNA was purified from the quenched DMS reaction with RNA Clean and Concentrator 5 kit and eluted in 10µL water. RNA was either directly processed or stored at -80°C.
In cell DMS modification and purification
DMS modification of SH-SY5Y cells was performed as follows. Cells were grown to 85% confluence in a 10 cm dish. Then, the media was exchanged for a DMS probing media which consisted of 1:1 DMEM to F12 medium with 10% FBS and 200 mM Bicine adjusted to pH 8.37 with NaOH at room temperature. Probing media was prewarmed to 37C and cell media was exchanged with 5.4 mL probing media for 3 min at 37°C. Then 600µL of prewarmed 1.7 M DMS or prewarmed 600µL ethanol was added to the cells and incubated for 6 min at 37°C. DMS reaction was stopped by the addition of 6 mL 20% 2-mercaptoethanol and cells were placed on ice. We noticed that DMS probing caused a large portion of adherent cells to become detached from the plate. We therefore decanted the quenched DMS probing media off the plate into a 15 mL falcon tube and pelleted the detached cells for 5 min at 4C and 1,000 x g. To isolate the total RNA, 2 mL Trizol reagent was added to the plate for the cells which had not become detached. Then, the 2 mL Trizol reagent was transferred to solubilize the pelleted cells which had become detached during DMS probing. Total RNA was then isolated according to manufacturer’s instructions.
M2 BASH MaP
Spinach and polyUG T7 templates were randomly mutagenized by performing 24 rounds of error-prone PCR. Mutagenized Spinach and polyUG were then in vitro transcribed, folded and DMS modified as described prior with minor modifications. MgCl2 was added to a final concentration of 5 mM and Spinach was probed in the presence of 5µM DFHBI-1T. polyUG RNA was probed in the presence of 2µM NMM.

value: cell line

value: HeLa

value: epithelial cell

value: 800 mM Potassium borohydride reduction followed by 4 hour depurination in 1M sodium acetate / acetic acid buffer pH 2.9 (BASH)


title: H3.3WTH3.2HWT, Male, Rep1


title: H3.3WTH3.2HWT, Male, Rep2


title: pUG, M2 BASH MaP, 170 mM SSII

description:
Duration: 6 min ;temperature: 37C;buffer: 200 mM Bicine pH 8.3, 150 mM KCl, 5 mM MgCl2;ligand: NMM 2 uM
M2 pUG DAGGER MaP

library_source: other

molecule: other

organism: synthetic construct

source_name: in vitro transcribed RNA

value: in vitro transcribed RNA

value: Pool of randomly mutagenized RNA produced through error-prone PCR of T7 template

value: DMS followed by BASH


title: HeLa total RNA, Borohydride reduction, SSII

description:
Duration: 0;temperature: N/A;buffer: N/A;ligand: N/A
18s Sequencing data 1


title: H3.3WTH3.2HWT, Female, Rep3


title: Mal C 0_C

type: RNA

city: Heidelberg

first: Emma

last: Phillips

address: In Neuenheimer Feld 580

department: Brain Tumor Translational Targets

institute: DKFZ

description:
SAMPLE 9
replicate 3

hyb_protocol: Standard Illumina Hybridization Protocol (Whole-Genome Gene Expression Direct Hybridization Assay Guide). 750 ng of cRNA were hybridized for 17 hr at 58°C on Illumina Human HT-12-Microarray.

scan_protocol: Gene Expression Microarrays were scanned using the Illumina iScan-Scanner according to Standard Illumina Scanning Protocol (Whole-Genome Gene Expression  Direct Hybridization Assay Guide)

data_processing: The data were normalized using quantile normalization with R using function normalize.quantiles from Bioconductor package preprocessCore

label: Biotin, Cy3-Streptavidin

source_name: NCH421k GSCs

label_protocol: The Illumina TotalPrep RNA Amplification Kit (life technologies) is used to generate single-stranded cRNA from input amounts of 200ng total RNA

growth_protocol: NCH421k GSCs were cultivated as floating neurospheres in serum-free DMEM/F-12 medium (Merck Millipore, F4815), supplemented with 20% BIT-admixture (Pelo Biotech, PB-SH-033-0000) and basal fibroblast growth factor (Biomol, 50361.50) and epidermal growth factor (Life Technologies, PHG0311) at 20 ng/ml each.

extract_protocol: RNA was extracted with RLT buffer and lysates were homogenized using QIAGEN QIAshredder columns. The QIAGEN RNeasy mini kit was used in accordance with the prescribed protocol

treatment_protocol: Cells were treated with 50 nM malformin C for 2, 4 and 6 hours, or DMSO for 6 hours.

tag: tumor stage

value: Glioblastoma

tag: tissue origin

value: Brain

value: GSC

value: control

tag: time

value: 0 hr


title: Mal C 6_C

description:
SAMPLE 12
replicate 3

value: Mal C

value: 6 hr


title: H3.3WTH3.2HWT, Female, Rep2

value: Wandering L3 brain

value: female


title: H3.3WTH3.2HWT, Female, Rep1


title: Spinach, M2 BASH MaP, 170 mM, SSII

description:
Duration: 8 min;temperature: 25C;buffer: 200 mM Bicine pH 8.3, 150 mM KCl, 5 mM MgCl2;ligand: DFHBI-1T 5 uM
M2 Spinach DAGGER MaP


title: Spinach, M2 DMS MaP, 170 mM, SSII

description:
Duration: 8 min;temperature: 25C;buffer: 200 mM Bicine pH 8.3, 150 mM KCl, 5 mM MgCl2;ligand: DFHBI-1T 5 uM
M2 Spinach DMS MaP

value: DMS


title: Mal C 6_B

description:
SAMPLE 8
replicate 2


title: pUG, M2 DMS MaP, 170 mM, SSII

description:
Duration: 6 min ;temperature: 37C;buffer: 200 mM Bicine pH 8.3, 150 mM KCl, 5 mM MgCl2;ligand: NMM 2 uM
M2 pUG DMS MaP


title: Mal C 6_A

description:
SAMPLE 4
replicate 1


title: Mal C 0_B

description:
SAMPLE 5
replicate 2


title: Mal C 0_A

description:
SAMPLE 1
replicate 1


title: Mal C 4_C

description:
SAMPLE 11
replicate 3

value: 4 hr


title: Mal C 2_B

description:
SAMPLE 6
replicate 2

value: 2 hr


title: Mal C 4_B

description:
SAMPLE 7
replicate 2


title: Mal C 2_A

description:
SAMPLE 2
replicate 1


title: Mal C 2_C

description:
SAMPLE 10
replicate 3


title: Mal C 4_A

description:
SAMPLE 3
replicate 1


title: WTT_2

city: Taipei

first: Yijuang

last: Chern

address: 128 Sec. 2, Academia Rd.

department: Institute of Biomedical Sciences

country: Taiwan

institute: Academia Sinica

data_processing:
The small RNA library construction, NGS deep sequencing and bioinformatic analysis were accomplished by Welgene Biotech. Co., Ltd. (Taipei, ROC).
Sequencing data was processed using the Illumina software BCL2FASTQ v2.20.
Trimmomatic was implemented to clip the 3′ adaptor sequence, trim or remove the reads according to the quality score and discard trimmed reads shorter than 18 nucleotides.
Qualified reads after filtering low-quality data were analyzed using miRDeep2 for aligning reads to the reference genome downloaded from UCSC.
miRNAs are detected by miRDeep2. Expression values are normalized by library size and multiplied by a factor of 1E6, which corresponds technically to counts per million mapped miRNA reads (RPM). The library size is the total number of reads mapping to miRNA precursors.
The selection criteria for DE miRNAs were FC > 1.25 and p < 0.05.
Genome_build: GRCm38
Supplementary_files_format_and_content: miRNA_WTT_vs_WTC.xlsx: Excel file includes miRNA counts and normalized miRNA for WTT vs. WTC.
Supplementary_files_format_and_content: miRNA_R6_2T_vs_R6_2C.xlsx: Excel file includes miRNA counts and normalized miRNA for R6_2T vs. R6_2C.
Supplementary_files_format_and_content: R6_2T_vs_R6_2C_differentially_expressed_miRNA.txt: Tab-delimited text file includes values for R6/2C vs. R6/2T differentially expressed miRNAs. miRNAs are detected by miRDeep2. Expression values are normalized by library size and multiplied by a factor of 1E6, which corresponds technically to counts per million mapped miRNA reads (RPM). The library size is the total number of reads mapping to miRNA precursors.

source_name: Striatum

growth_protocol: R6/2 mice (B6CBA-Tg(HDexon1)62Gpb/3J) were obtained from Jackson Laboratory (Bar Harbor, ME, USA) and mated with female control mice (B6CBAFI/J). Mice were housed at the Institute of Biomedical Sciences (IBMS) Animal Care Facility (Academia Sinica, Taipei, Taiwan) under a standard 12 hr light/dark cycle. Animal experiments were performed under approved protocols by the Academia Sinica Institutional Animal Care and Utilization Committee (Taipei, Taiwan).

extract_protocol:
RNA was extracted by TRIzol reagent or miRNeasy mini kit (Qiagen). DNase digestion was performed by the DNA-free™ Kit (Life Technologies, Carlsbad, CA, USA) or the RNase-free DNase set (Qiagen) following the manufacturer’s instructions. The concentration and purity of RNA were measured at 260, 280 and 230 nm by a NanoDrop (Thermo Fisher Scientific, Waltham, MA, USA).
Sample libraries were prepared using the QIAseq miRNA Library Kit (QIAGEN) according to the manufacturer's protocols. Adaptors are ligated sequentially to the 3' and 5' ends of miRNAs. Subsequently, universal cDNA synthesis with UMI assignment, cDNA cleanup, library amplification and library cleanup are performed. Libraries were sequenced on an Illumina instrument (75-cycle single-end read, 75SE).

treatment_protocol: For the bilateral intrastriatal delivery of AAV expressing control or TRAX shRNA, mice were anesthetized with ketamine/xylazine HCl (87.56 and 7.5 ug/kg, respectively, via an intraperitoneal injection) and injected with the indicated AAV (2 µl per injection spot, 2.5*10^9 vector genome/µl) with a 10 ml syringe (Hamilton, Reno, NV, USA) at a rate of 0.5 ul/min. The AAV was injected at the desired position: AP+0.5, L±2, DV-2.5 and -3.5 mm relative to bregma and the dura surface. Then the mouse brains were removed from the mice directly for dissection of striatum.

tag: strain

value: WT

tag: shRNA

value: shTRAX

value: Striatum

value: 10.5 weeks


title: WTT_3


title: pUG, 0 mM, SSII

description:
Duration: 6 min ;temperature: 37C;buffer: 200 mM Bicine pH 8.3, 150 mM KCl, 5 mM MgCl2;ligand: NMM 2 uM
Poly UG_untreated

value: in vitro transcribed RNA

value: EtOH no DMS control


title: Spinach, BASH MaP, 10.625 mM, SSII

description:
Duration: 10 min;temperature: 25;buffer: 200 mM Bicine pH 7.75, 100 mM KCl, 5 mM MgCl2;ligand: DFHBI-1T 5 uM
Spinach initial DM11

value: DMS followed by BASH


title: R6_2T_3


title: Spinach, BASH MaP, 0 mM, SSII

description:
Duration: 10 min;temperature: 25;buffer: 200 mM Bicine pH 7.75, 100 mM KCl, 5 mM MgCl2;ligand: DFHBI-1T 5 uM
Spinach initial DM12

value: EtOH no DMS control followed by BASH


title: pUG, 170 mM, SSII

description:
Duration: 6 min ;temperature: 37C;buffer: 200 mM Bicine pH 8.3, 150 mM KCl, 5 mM MgCl2;ligand: NMM 2 uM
Poly UG_DMS


title: Spinach, BASH MaP, 42.5 mM, SSII

description:
Duration: 10 min;temperature: 25;buffer: 200 mM Bicine pH 7.75, 100 mM KCl, 5 mM MgCl2;ligand: DFHBI-1T 5 uM
Spinach initial DM9


title: WTC_1

value: Control


title: WTC_2


title: Spinach, BASH MaP, 21.25 mM, SSII

description:
Duration: 10 min;temperature: 25;buffer: 200 mM Bicine pH 7.75, 100 mM KCl, 5 mM MgCl2;ligand: DFHBI-1T 5 uM
Spinach initial DM10


title: WTC_3


title: Spinach, BASH MaP, 85 mM, SSII

description:
Duration: 10 min;temperature: 25;buffer: 200 mM Bicine pH 7.75, 100 mM KCl, 5 mM MgCl2;ligand: DFHBI-1T 5 uM
Spinach initial DM8


title: R6_2C_3


title: R6_2C_2


title: Spinach, DMS MaP, 21.25 mM, SSII

description:
Duration: 10 min;temperature: 25;buffer: 200 mM Bicine pH 7.75, 100 mM KCl, 5 mM MgCl2;ligand: DFHBI-1T 5 uM
Spinach initial DM4


title: R6_2C_1


title: Spinach, DMS MaP, 10.625 mM, SSII

description:
Duration: 10 min;temperature: 25;buffer: 200 mM Bicine pH 7.75, 100 mM KCl, 5 mM MgCl2;ligand: DFHBI-1T 5 uM
Spinach initial DM5


title: Spinach, BASH MaP, 170 mM, SSII

description:
Duration: 10 min;temperature: 25;buffer: 200 mM Bicine pH 7.75, 100 mM KCl, 5 mM MgCl2;ligand: DFHBI-1T 5 uM
Spinach initial DM7


title: R6_2T_1


title: MHCC97H_P65

city: Shanghai

first: Peiyi

last: Xie

address: Fenglin Road No.180

country: China

institute: Zhongshan Hospital, Fudan University

data_processing:
Peak calling was performed by Model based Analysis of ChIP-seq (MACS2 version 2.1.0.20151222) on UseGalaxy (usegalaxy.org) server. The cutoff value for peak detection was set to 0.05. BamCoverage (version 2.2.3) from deeptools was used to generate BigWig files, which were visualized by integrative genomics viewer (IGV version 2.5.3). The input peaks were subtracted from the p65 binding peaks and visualized in IGV.
Assembly: mm9
Supplementary files format and content: bed

extract_protocol:
For ChIP-seq, DNA samples were sonicated in a Biorupter Pico (Diagenode) to acquire nearly 300 bp fragments. Next, the MicroPlex Library Preparation Kit (Diagenode) was employed for library preparation, followed by sequencing through the HiSeq 2000 Illumina platform of the NIG. The quality of the ChIP-seq raw data files were evaluated through FASTQ quality check (FASTQC). FastQ files were evaluated in the public server usegalaxy.org where the sequence reads were aligned to the mouse reference genome (mm9) using Bowtie2 (version 0.4) with default parameters.
For ChIP analysis, MHCC97H cells were fixed employing 1% formaldehyde for 30 min, followed by adding 1.25 mol/L glycine for 5 minutes to end the reaction.

value: human heptocellular carcinoma

tag: antibody


title: Spinach, DMS MaP, 0 mM, SSII

description:
Duration: 10 min;temperature: 25;buffer: 200 mM Bicine pH 7.75, 100 mM KCl, 5 mM MgCl2;ligand: DFHBI-1T 5 uM
Spinach initial DM6


title: MHCC97H_Input

value: input


title: Spinach, DMS MaP, 85 mM, SSII

description:
Duration: 10 min;temperature: 25;buffer: 200 mM Bicine pH 7.75, 100 mM KCl, 5 mM MgCl2;ligand: DFHBI-1T 5 uM
Spinach initial DM2


title: RNA-Seq of Campeiostachys nutans: salt-leaf-2

city: Yangling

first: Juanjuan

last: Fu

state: Shaanxi

address: No.22 Xinong Road

institute: Northwest A&F University

data_processing:
The adapter sequences were removed, and raw reads containing ambiguous bases ‘N’ > 5 % or those what were low-quality (more than 20 % Q ≤ 10 bases) were re_x0002_moved.
The Inchworm, Chrysalis, and Butterfly modules of Trinity software (version: v2.0.6) was used for de novo assembly of the clean reads
The Inchworm pro_x0002_gram assembled the reads into the unique sequences of transcripts;
Chrysalis program clusters the Inchorm Contigs into clusters and constructs complete de Bruijn graphs for each cluster.
Butterfly program con_x0002_structed the transcripts. Then use Tgicl v2.0.6 clustered transcripts to unigenes, which involves sequence splicing and redundancy removal to acquire non-redundant uni_x0002_genes.
Assembly: De novo transcriptome (Trinity.gene.fasta)

organism: Campeiostachys nutans

source_name: leaf

extract_protocol:
Total RNA was extracted using a Trizol reagent according to the manufacturer’s protocol (Takara Bio Inc., Otsu, Japan). The integrity and purity of the RNA was determined by using the Agilent 2100 RNA 6000 Kit and electrophoresis on 1 % agarose gel.
De novo transcriptome sequencing of the samples was performed by Illumina NovaSeq 6000 instrument at the Lc. Biotech Co., Ltd. (Hangzhou, China, http://www.lc-bio.com/) following the manufacturer’s procedure.

value: leaf

value: 28-day-old C. nutans seedlings treated without 200 mM salt treatment for 5 d

value: wt

value: salt


title: RNA-Seq of Campeiostachys nutans: salt-leaf-1


title: Hippocampus from AppNL-G-F; Trem2 KO, 3 mo, AKTK184

city: Columbus

first: Jie

last: Gao

state: OH

address: 460 W. 12th Ave

department: The Department of Neuroscience

institute: The Ohio State University

data_processing:
Demultiplexed FASTQ files of bulk RNA sequencing data were aligned to the mouse genome (Mus_musculus.GRCm39) using STAR (version 2.7.10a).
Adapters were trimmed using flexbar (version 3.5.0.).
Reads mapped to genomic features were counted using featureCounts (version 2.0.3).
The count matrix was imported in R (version 4.2.0) for analysis. Gene expression signatures of different experimental conditions were compared using DeSeq2 (version 1.36.0), using the Wald test for hypothesis testing.
Assembly: Mus_musculus.GRCm39
Supplementary files format and content: tab-delimited text file includes raw counts for each sample

source_name: Hippocampus

extract_protocol:
Total RNA was extracted using Quick-RNA miniprep (R1055, Zymo Research). RNA quality was evaluated by TapeStation using high Sensitivity RNA ScreenTape (5067-5579, Agilent)
RNA seq libraries were prepared using SMART-Seq mRNA LP Kit (Takara Bio) following the manufacturer's instructions

value: Hippocampus

value: AppNL-G-F; Trem2 KO

tag: gender

value: male

value: 3 mo


title: R6_2T_2

value: shTRAX

value: Striatum


title: Hippocampus from AppNL-G-F; Trem2 KO, 3 mo, AKTK185


title: Hippocampus from AppNL-G-F; Trem2 KO, 9 mo, AKTK3677

value: 9 mo


title: Hippocampus from AppNL-G-F; Trem2 KO, 9 mo, AKTK3679


title: Hippocampus from AppNL-G-F; Trem2 KO, 3 mo, AKTK183


title: RNA-Seq of Campeiostachys nutans: control-leaf-2

tag: treatment

value: control


title: Hippocampus from AppNL-G-F, 9 mo, AK4277

value: AppNL-G-F


title: Hippocampus from AppNL-G-F; Trem2 KO, 9 mo, AKTK3676


title: Hippocampus from AppNL-G-F, 9 mo, AK4276


title: RNA-Seq of Campeiostachys nutans: control-leaf-1


title: Spinach, DMS MaP, 42.5 mM, SSII

description:
Duration: 10 min;temperature: 25;buffer: 200 mM Bicine pH 7.75, 100 mM KCl, 5 mM MgCl2;ligand: DFHBI-1T 5 uM
Spinach initial DM3

value: in vitro transcribed RNA

value: WT

value: DMS


title: Hippocampus from AppNL-G-F, 3 mo, AK130


title: ECA722 x  N2, F1, rep3

city: Atlanta

first: Avery

middle: Davis

last: Bell

state: GA

address: North Avenue

department: School of Biological Sciences

institute: Georgia Institute of Technology

data_processing:
RNA-seq data were quantified against strain-specific transcriptomes, either just for one strain or pseudo-diploid for the reference strain and N2. These 'personalized' transcriptomes were generated by inserting variants from CaeNDR (20210121 release for Salmon and emase processing; 20220216 for Kallisto processing) into the C. elegans reference genome using g2gtools (v0.1.31).
we generated a salmon (v1.4) index for the diploid transcriptome using salmon index with options -k 31 --keepDuplicates (no decoy, all other parameters default)
FASTQ files were quality controlled with FastQC and Illumina adapter sequences were trimmed using Trimmomatic (v0.39) with parameters ILLUMINACLIP: TruSeq3-PE-2.fa:1:30:12:2:True
Salmon (v1.4) quantification with equivalence class outputs saved was performed against the pseudo-diploid transcript’s index with salmon quant -l ISR --dumpeq --fldMean <sample-specific mean> --fldSD <sample-specific SD> --rangeFactorizationBins 4 --seqBias --gcBias
Salmon outputs were converted to .bin inputs for emase-zero using alntools salmon2ec (v0.1.1)
emase-zero (v0.3.1) was run on this input using parameters --model 4 -t 0.0001 -i 999
For kallisto abundance estimates, first, pseudo-diploid and strain-specific transcriptomes were created using script create_personalized_transcriptome.py from the Ornaments code suite (initial version)
Kallisto indexes of strain specific transcriptomes were created with command kallisto index (v0.50.1)
Kallisto abundance estimates were generated from strain-specific transcriptome index using command kallisto quant with parameter --rf-stranded
To generate the count matrix, kallisto abundances were read into DESeq2 (v1.42.0) using tximport (v1.30.0), followed by DESeq2's length and library size normalization and variance stabilizing transformation (vst)
Assembly: ce11 (ws276)
Supplementary files format and content: Salmon quants.sf output files for RNA quantification of diploid transcripts (transcripts are named <name>_<parental strain haplotype>). One tab-delimited text file per sample
Supplementary files format and content: Salmon auxillary info eq_classes.txt output file for quantification on diploid transcriptome. One text file per sample
Supplementary files format and content: emase-zero per-gene counts output. One tab-delimited text file per sample
Supplementary files format and content: Kallisto abundance.tsv output with RNA abundance metrics for each transcript (from strain-specific transcriptome). One tab-delimited text file per sample
Supplementary files format and content: Genes x samples matrix of estimated counts from kallisto abundances after DESeq2 tximport, length and library size normalization, and variance stabilizing (vst) transformation. Tab delimited text file with row names and column names.

organism: Caenorhabditis elegans

source_name: whole animal

growth_protocol: We first bleach synchronized all parental strains to ensure that the parents that would be mated were of similar developmental stage. After allowing these worms to grow for two days, we generated mating plates by placing 60-80 N2fog-2 L4 pseudo-hermaphrodites onto each of five 6cm plates with small bacteria spots, and added 40 L4 males of the appropriate strain to each plate. We concurrently moved 80 individual L4 hermaphrodites to each of three 6cm plates for each parental strain (N2 and seven wild strains) to simultaneously generate the parental strains used for sequencing from self-matings while the F1 crosses were generated from cross-matings. After allowing mating for 48 hours, we collected and synchronized the offspring for the crosses and self-matings by collecting all parental worms and embryos from the bacterial lawn, treating with bleach, and allowing embryos to develop into L1 larvae and arrest over 30 hours in liquid buffer. After 30 hours, L1s were transferred directly to the bacterial lawn of 6cm plates at a density of ~400 L1s per plate. After allowing the worms to develop for ~36 hours, we removed males from the F1 plates as soon as they were detectable, and screened the parental plates for any spontaneously-generated males, which were also removed. Plates used for RNA sequencing (at least 3 per strain) had all of their males removed as L4s or young adults. Worms were harvested as day 1 reproductively mature young adults, specifically when most worms were gravid with embryos and laid embryos were visible on the plates.

extract_protocol:
RNA was extracted from worms stored in TRIzol (Invitrogen #15596026) following standard procedure (following  He, 2011) using a TRIzol (Invitrogen #15596026) chloroform (Fisher #C298-500) extraction and RNeasy columns (Qiagen #74104). This extraction was performed in 3 batches of 15 over two consecutive days, with one replicate from each strain included in each batch. RNA was stored at -80°C for ~1 week prior to library generation.
Following RNA quality checks (all RINs 9.8 or greater), mRNA was enriched from 1μg RNA with the NEBNext Poly(A) mRNA magnetic isolation module (NEB #E7490) and sequencing libraries generated using the NEBNext Ultra II directional RNA library preparation kit (NEB #E7760) with 8 cycles of PCR

value: whole animal

value: wildtype

value: ECA722 x N2 F1

value: Hermaphrodite

tag: developmental stage

value: day 1 adult

tag: generation


title: Hippocampus from AppNL-G-F, 9 mo, AK4275

value: Hippocampus


title: QX1211 x  N2, F1, rep3

tag: tissue


title: Hippocampus from AppNL-G-F, 3 mo, AK129


title: CB4856 x  N2, F1, rep3

tag: Sex


title: Splenocytes BbIsPDIA6 Rep2

city: Hangzhou

first: Xiaotian

last: Tang

state: Zhejiang

address: Agro-Bio-Environ Building, Zhejiang University

department: Entomology

institute: Zhejiang University

data_processing:
Partek Flow
RNAseq data were trimmed and aligned to the mouse genome ( from Ensembl), with associated annotation file using STAR (v2.7.3a).
The aligned reads were quantified to Ensembl transcripts using the Partek E/M algorithm and the subsequent steps were performed on gene-level annotation followed by total count normalization.
The counts generated were processed by DESeq2.
Assembly: mm39
Supplementary files format and content: tab-delimited text files include baseMean for each gene

source_name: Spleen

extract_protocol:
RNA was isolated using RNeasy Mini kit according to the manufacturer’s instructions (Qiagen, #74104).
RNA libraries were prepared for sequencing using standard Illumina protocols

treatment_protocol: The murine splenocytes were stimulated by the Lyme disease agent Borrelia burgdorferi, with or without IsPDIA6, for 6 hours.

value: Spleen

value: Splenocytes

value: Borrelia and IsPDIA6


title: RNA-Seq of Campeiostachys nutans: salt-leaf-3

value: leaf

value: 28-day-old C. nutans seedlings treated without 200 mM salt treatment for 5 d

tag: genotype

value: wt

value: salt


title: RNA-Seq of Campeiostachys nutans: control-leaf-3

value: control


title: Hippocampus from AppNL-G-F, 3 mo, AK132

value: AppNL-G-F

value: male

value: 3 mo


title: EG4348 x N2, F1, rep3

value: whole animal


title: DL238 x  N2, F1, rep3

value: wildtype

value: DL238 x N2 F1


title: ECA701 x  N2, F1, rep3

value: ECA701 x N2 F1

value: Hermaphrodite


title: Splenocytes BbIsPDIA6 Rep1


title: Splenocytes Bb Rep1

value: Borrelia


title: ECA722, parental, rep3

value: day 1 adult

value: parental


title: WM989 cisplatin treated, scRNA-seq.

first: Dylan

middle: L

last: Schaff

address: 421 Curie Blvd, BRB II/II rm 445

department: Bioengineering

description: 10X Genomics

data_processing:
Data was processed using CellRanger software 5.0.1, aligning to the hg38 assembly.
While generating count matrices, we also linked seqeuncing reads from our clonal barcodes to the 10X cell barcodes.
We uploaded the outputs from the filtered feature-barcode matrix
Assembly: hg38
Supplementary files format and content: *.matrix.mtx: Sparse count matrix
Supplementary files format and content: *.features.tsv: Tab-separated list of features (genes or clone) and identifiers for type of feature.
Supplementary files format and content: *.barcodes.tsv: 10X cell barcodes

source_name: Melanoma

extract_protocol:
Cells were trypsinized and washed with PBS before following the 10X Genomics 3' v3.1 protocol to generate single cell emulsions with beads. We loaded cells following instructions to achieve 10,000 cells per condition.
We used the 10X Genomics 3' v3.1 protocol for library construction. In brief, single cells were encapsulated into gel bead-in-emulsions (GEMs) using the 10X genomics chromium controller. RNA with a polyA tail was reverse transcribed into cDNA. cDNA from all cells in a sample was cleaned using Dynabeads, the cDNA was amplified, and the samples were size selected using AMPure XP beads. Full-length cDNA was fragmented and sequencing adapters and indices were ligated. A final amplification was performed before being cleaned with AMPure XP beads a final time. The final libraries were then sequenced.

value: Melanoma

tag: cell line

value: BRAF V600E

value: Cisplatin


title: Splenocytes Bb Rep3

value: Spleen

tag: cell type

value: Splenocytes


title: Splenocytes Bb Rep2


title: N2, parental, rep3

value: N2 with fog-2 deletion


title: ECA722 x  N2, F1, rep2

value: ECA722 x N2 F1


title: WM989 doxorubicin treated, scRNA-seq

description: 10X Genomics

value: Doxorubicin


title: XZ1516, parental, rep3


title: CB4856, parental, rep3


title: DL238, parental, rep3


title: ECA701, parental, rep3


title: WM989 CoCl2 treated, scRNA-seq.

description: 10X Genomics


title: QX1211, parental, rep3


title: XZ1516 x  N2, F1, rep3


title: WM989 acid treated, scRNA-seq

description: 10X Genomics

value: Acidic media


title: WM989 trametinib treated, scRNA-seq

description: 10X Genomics

value: Trametinib


title: XZ1516 x  N2, F1, rep2

value: whole animal

value: wildtype


title: EG4348, parental, rep3


title: ECA701 x  N2, F1, rep2

value: ECA701 x N2 F1


title: WTT_1

value: WT

value: shTRAX

value: Striatum

tag: age

value: 10.5 weeks


title: Spinach, DMS - Reduction + Depurination, 170 mM, SSII

description:
Duration: 6 min ;temperature: 37C;buffer: 200 mM Bicine pH 8.3, 150 mM KCl, 2.5 mM MgCl2;ligand: N/A
Spinach M2G4 2

value: in vitro transcribed RNA

value: DMS - KBH4 Reduction + 4 hour Depurination


title: Spinach, + Reduction + Depurination, 0 mM, SSII

description:
Duration: 0 min ;temperature: 37C;buffer: 200 mM Bicine pH 8.3, 150 mM KCl, 2.5 mM MgCl2;ligand: N/A
Spinach M2G4 3

value: EtOH no DMS control + 4 hour 800 mM KBH4 Reduction + 4 hour Depurination


title: CB4856 x  N2, F1, rep2

value: Hermaphrodite

value: day 1 adult


title: Spinach, DMS + Reduction + Depurination, 170 mM, SSII

description:
Duration: 6 min ;temperature: 37C;buffer: 200 mM Bicine pH 8.3, 150 mM KCl, 2.5 mM MgCl2;ligand: N/A
Spinach M2G4 1

value: DMS + 4 hour 800 mM KBH4 Reduction + 4 hour Depurination


title: DL238 x  N2, F1, rep2

value: DL238 x N2 F1


title: Untreated WM989 sample 3, scRNA-seq

description: 10X Genomics

value: Melanoma

value: BRAF V600E

value: Untreated


title: QX1211 x  N2, F1, rep2

value: whole animal


title: Spinach, - Reduction + Depurination, 0 mM, SSII

description:
Duration: 0 min ;temperature: 37C;buffer: 200 mM Bicine pH 8.3, 150 mM KCl, 2.5 mM MgCl2;ligand: N/A
Spinach M2G4 4

value: EtOH no DMS control - KBH4 Reduction + 4 hour Depurination


title: EG4348 x N2, F1, rep2

value: wildtype


title: Untreated WM989 sample 2, scRNA-seq

description: 10X Genomics


title: WM989 dabrafenib treated, scRNA-seq

description: 10X Genomics

value: Dabrafenib


title: ECA722, parental, rep2

value: parental


title: XZ1516, parental, rep2


title: Total ribosomes profiling (Reh1 Δ6), rep1

city: Austin

first: Can

last: Cenik

state: TX

address: Moffett Molecular Biology Building 2.122 2500 Speedway

department: Molecular Biosciences

institute: The University of Texas at Austin

data_processing:
Ribosome profiling data were processed using RiboFlow. We extracted the first 12 nucleotides from the 5’ end of the reads using UMI-tools with the following parameters: 'umi_tools extract -p "^(?P<umi_1>.{12})(?P<discard_1>.{4}).+$" --extract-method=regex'. The four nucleotides downstream of the UMIs are discarded as they are incorporated during the reverse transcription step. We clipped the 3’ adapter AAAAAAAAAACAAAAAAAAAA using cutadapt with the parameters “-a AAAAAAAAAACAAAAAAAAAA --overlap=4 --trimmed-only --maximum-length=56 --minimum-length=31 --quality-cutoff=28”. After UMI extraction and adapter trimming, reads were aligned to ribosomal and transfer RNAs using Bowtie2 with the parameters "-L 15 --no-unal --norc". The unaligned reads were mapped to a manually-curated transcriptome with the parameters "-L 15 --norc --no-unal". We retained alignments with mapping quality greater than 2 followed by deduplication using UMI-tools. As the last step, .ribo files are created using RiboPy Assembly: A curated transcriptome of primary transcripts was used for alignment. Reference files for use with RiboFlow are located at https://github.com/ribosomeprofiling/yeast_reference Supplementary files format and content: .ribo HDF5-formatted files contain ribosome occupancy at nucleotide resolution for lengths from 21 to 40. See https://ribopy.readthedocs.io/en/latest/ribo_file_format.html for a detailed description of the ribo format.
Ribosome profiling data were processed using RiboFlow. We extracted the first 12 nucleotides from the 5’ end of the reads using UMI-tools with the following parameters: 'umi_tools extract -p "^(?P<umi_1>.{12})(?P<discard_1>.{4}).+$" --extract-method=regex'. The four nucleotides downstream of the UMIs are discarded as they are incorporated during the reverse transcription step.
We clipped the 3’ adapter AAAAAAAAAACAAAAAAAAAA using cutadapt with the parameters “-a AAAAAAAAAACAAAAAAAAAA --overlap=4 --trimmed-only --maximum-length=56 --minimum-length=31 --quality-cutoff=28”.
After UMI extraction and adapter trimming, reads were aligned to ribosomal and transfer RNAs using Bowtie2 with the parameters "-L 15 --no-unal --norc". The unaligned reads were mapped to a manually-curated transcriptome with the parameters "-L 15 --norc --no-unal". We retained alignments with mapping quality greater than 2 followed by deduplication using UMI-tools. As the last step, .ribo files are created using RiboPy
Assembly: A curated transcriptome of primary transcripts was used for alignment. Reference files for use with RiboFlow are located at https://github.com/ribosomeprofiling/yeast_reference
Assembly: Supplementary files format and content: .ribo HDF5-formatted files contain ribosome occupancy at nucleotide resolution for lengths from 21 to 40. See https://ribopy.readthedocs.io/en/latest/ribo_file_format.html for a detailed description of the ribo format.
Supplementary files format and content: Ribo files
Supplementary files format and content: Ribo files from -35 to 60 nt
Library strategy: Ribo-seq

organism: Saccharomyces cerevisiae

source_name: yeast

growth_protocol: Cultures of AJY4408 with plasmid pAJ4761 were grown to OD600 of 0.3 in 1.5 L of Leu-Glucose media by continuous overnight growth. β-estradiol was added to a final concentration of 1 uM and growth was continued for an additional 90 min.

extract_protocol:
Cells were washed once in IP buffer (Tris HCl 20 mM pH 7.5, 100 mM KCl, 10 mM MgCl2, 100 ug/ml CHX, Pierce Protease Inhibitor Cocktail, EDTA-free (1 tablet/10 ml), 1 mM PMSF, 1 uM each leupeptin and pepstatin and 5 mM beta-mercaptoethanol (BME)) and then resuspended in a volume of IP buffer equal to that of the cell pellet. Extracts were prepared by agitation with glass beads, Igepal was added to 0.1% extracts were clarified by centrifugation at 4°C for 15 minutes at 18,000 g. 20 mg of RNA (~2 ml sample) was digested with 62.5 units of RNase I (Epicentre) at 4°C for 1 hr. 50ul was removed for total Ribo-seq sample. 25 ul bed volume washed anti-FLAG magnetic beads (Sigma Milipore) was added to the remaining sample and samples were rotated for 1 hr at 4°C. Beads were washed 4 times with IP buffer supplemented with 0.1% Igepal and transferred to a clean tube during the last wash. Reh1-bound ribosomes were eluted in 100 ul of IP buffer supplemented with 0.1% Igepal and 150 ug/ml 3xFLAG peptide (Sigma Milipore) for 15 min at 4°C. 12.5 units RNase I was added to the eluate and digestion was carried out at room temperature for 45 min. RNAse I digestion reaction was stopped by addition of QIAzol followed by RNA extraction by chloroform and ethanol precipitation. For total RIbo-seq samples, 6.25 units RNase I (Epicentre) was added to 5ul of sample in a total volume of 50 ul in IP buffer supplemented with 0.1% Igepal. Digestion was carried out at room temperature for 45 min. Digested lysates were layered onto 50 ul 1M sucrose cushions prepared in IP buffer and ribosomes were pelleted by centrifugation at 70,000rpm in a TLA 100 rotor (Beckman Coulter) for 30 min at 4°C.
RNA was extracted from the ribosome pellets with the addition of 700 μl QIAzol followed by chloroform and ethanol precipitation. Isolated RNAs were size-selected using a 15% polyacrylamide TBE-UREA gel (Invitrogen) and 28-35 nt fragments were excised and extracted by crushing the gel fragment in 400 µL of RNA extraction buffer (300 mM NaOAc [pH 5.5], 5 mM MgCl2). The sample was passed through a Spin X filter (Corning 8160) and the flow through was ethanol precipitated (2.5 X volume) with 1.5 µL of Glycoblue (Invitrogen AM9516).
Ribosome profiling libraries were prepared as previously described43 using the D-Plex Small RNA-Seq Kit (Diagenode). The cDNA was amplified for ten cycles. Each library was cleaned with PCR purification kit (Qiagen) and eluted with 30 μL of RNase-free water. To completely remove the primer-dimers, size-selection was done in a 3% agarose pre- cast gel (Sage Science) with the BluePippin system (Sage Science) using 180-235 nt range (tight settings, target:208 bp). The resulting size-selected libraries were analyzed with the Agilent High Sensitivity DNA Kit (Agilent). Each library was mixed in equimolar proportion for sequencing with the NovaSeq 6000 SP PE.

treatment_protocol: Cycloheximide (CHX) was added to a final concentration of 100 µg/ml and cultures were shaken for 10 minutes at 30°C, harvested and frozen at -80°

value: yeast


title: Untreated WM989 sample 1, scRNA-seq

description: 10X Genomics


title: DL238, parental, rep2


title: Total ribosomes profiling (Reh1 Δ6), rep2


title: Brunello library sequencing to validate representation of all guide RNAs

city: Dallas

first: Nicholas

middle: K

last: Conrad

state: Texas

address: 6000 Harry Hines Blvd

department: Microbiology

institute: UT Southwestern Medical Center

data_processing:
The fastq files were subjected to quality check using fastqc (version 0.11.9, http://www.bioinformatics.babraham.ac.uk/projects/fastqc) and adaptors trimmed using Cutadapt version 3.3 (Martin M. et al. 2011).
The trimmed fastq files were mapped to Brunello library sequence (Doench et al. 2016; Addgene #73179), read counts generated, and median normalized using MAGeCK-VISPR version 0.5.6 (Li W. et al. 2015) disallowing any mismatches.
Positively and negatively selected sgRNA and genes were identified using the default parameters of MAGeCK-VISPR.
Assembly: n/a; seqences aligned to Brunello library (Doench et al. 2016; Addgene #73179)
Supplementary files format and content: rra.gene_summary.txt - text file; gene summary output from MAGeCK-VISPR
Supplementary files format and content: rra.sgrna_summary.txt - text file; sgRNA summary output from MAGeCK-VISPR
Supplementary files format and content: library_counts.txt - text file; read counts corresponding to each guide RNA in the Brunello library from the 'library' sample
Library strategy: CRISPR knockout screen

extract_protocol:
DNA was extracted using phenol:chloroform:iso-amyl alcohol method. See Scarborough et al., 2021 for details.
Library amplification was completed following a two-step variation of the BROAD institute protocol (BROAD Institute PCR of sgRNAs for Illumina sequencing). The two step variation of the BROAD institute protocol consists of an initial amplification step using primers flanking the P5 and P7 primers used for sequencing.

treatment_protocol: Cells were treated with 1 μM TG for 24 hours (replenished every 12 hours) on day 9


title: Spinach, DMS MaP, 170 mM, SSII

description:
Duration: 10 min;temperature: 25;buffer: 200 mM Bicine pH 7.75, 100 mM KCl, 5 mM MgCl2;ligand: DFHBI-1T 5 uM
Spinach initial DM1

value: WT

value: DMS


title: TG-treated CRISPR screen - sorted - biological replicate 3

value: 1 μM TG for 24 hours


title: TG-treated CRISPR screen - input - biological replicate 3


title: EG4348, parental, rep2


title: N2, parental, rep2

value: N2 with fog-2 deletion


title: QX1211, parental, rep2

value: Hermaphrodite


title: ECA722 x  N2, F1, rep1

value: ECA722 x N2 F1

value: day 1 adult


title: CB4856, parental, rep2


title: Δ6Reh1-bound ribosomes profiling, rep2


title: TG-treated CRISPR screen - sorted - biological replicate 2


title: DL238 x  N2, F1, rep1

value: DL238 x N2 F1


title: XZ1516 x  N2, F1, rep1


title: ECA701 x  N2, F1, rep1

value: ECA701 x N2 F1


title: CB4856 x  N2, F1, rep1


title: TG-treated CRISPR screen - sorted - biological replicate 1


title: Reh1-bound ribosomes profiling, rep2


title: ECA701, parental, rep2


title: Reh1-bound ribosomes profiling, rep1


title: ECA722, parental, rep1


title: Total ribosomes profiling (Reh1), rep2


title: N6203HS96h

city: Shenzhen

first: Kun

last: Sun

state: Guangdong

address: Rm A1921, Guangming International Innovation Center,

institute: Shenzhen Bay Laboratory

data_processing:
Raw RNA-seq reads were first preprocessed using Ktrim software to remove sequencing adaptors and low-quality cycles; PCR duplicates (i.e., reads with identical sequences) and ribosomal RNAs were then removed using in-house programs and the remaining reads were aligned to the human genome (build NCBI GRCh38) using STAR software; expression quantification were performed using featureCounts software against RefSeq gene annotation.
Differential expression analysis were performed using DESeq2 software; genes with an expression change larger than 1.5-fold and adjusted p-value smaller than 0.05 were considered as differentially expressed genes (DEGs). Functional annotation of the DEGs were performed using DAVID webserver.
Assembly: NCBI GRCh38
Supplementary files format and content: Raw read counts for all genes

extract_protocol:
Total RNA was collected from the engineered cells at 24, 48, and 96 h after HS using the RNA microprep kit (Zymo Research, R1050) and sent for bulk RNA-seq (Novogene)
Library was constructed by Novogene following standard protocol


title: Δ6Reh1-bound ribosomes profiling, rep1


title: Total ribosomes profiling (Reh1), rep1


title: TG-treated CRISPR screen - input - biological replicate 2


title: XZ1516, parental, rep1


title: QX1211 x  N2, F1, rep1


title: N6203CT96h


title: EG4348 x N2, F1, rep1


title: QX1211, parental, rep1


title: CB4856, parental, rep1


title: N6215HS96h


title: EG4348, parental, rep1


title: D1_Teff_Stimulated_AAVS1_1_KO_RNA

city: San Francisco

first: Alexander

last: Marson

state: CA

address: 1650 Owens St

institute: UCSF Gladstone Institutes

data_processing:
RNAseq data was processed using the pipeline described in Freimer et al 2022. In brief, fastq adapter trimming was performed with cutadapt (v2.10).
Low-quality bases were trimmed with seqtk (v0.5.0).
Reads were then aligned with STAR62 (v 2.7.10a) and mapped to GRCh38.
UMI counting and deduplication was performed with umi_tools (v1.0.1) and gene counts were generated from the deduplicated reads using featureCounts (subread v2.0.1) using Gencode v41 basic transcriptome annotation.
Quality control metrics were generated for each sample with Fastqc (v0.11.9), rseqc (v3.0.1), and Multiqc (v1.9).
Assembly: GRCh38
Supplementary files format and content: deduplicated count matrix for all files

source_name: Lymphocyte

growth_protocol: CD4+ regulatory and effector T cells were isolated from fresh Peripheral Blood Leukopaks (STEMCELL Technologies, #70500) from healthy human donors, after institutional review board–approved informed written consent (STEMCELL Technologies). The contents of the Leukopaks were washed twice with a 1X volume of EasySep buffer (DPBS, 2% fetal Bovine Serum (FBS), 1mM pH 8.0 EDTA) using centrifugation. The washed cells were resuspended at 200E6 cells/mL in EasySep buffer and isolated with the EasySep™ Human CD4+CD127lowCD25+ Regulatory T Cell Isolation Kit (STEMCELL Technologies, #18063), according to the manufacturer’s protocol. Following isolation with the kit, Tregs were stained Alexa Fluor® 647 anti-human CD25 Antibody (Biolegend, #302618), PE anti-Human CD127 (Beckon Dickinson, #557938), and Pacific Blue™ anti-human CD4 Antibody (Biolegend, #344620) and isolated using FACS to ensure a pure population without contaminating effector cells. After sorting pure CD4+CD127lowCD25+ Regulatory T Cells, the cells were seeded at 1x106 cells/mL in XVIVO-15 (Lonza, #02-053Q) supplemented with 55 uM 2-mercaptoethanol, 4 mM N-acetyl L-cysteine, and 200 U/mL IL-2 (Amerisource Bergen, #10101641). Teffs were seeded at 1x106 cells/mL in RPMI-1640 supplemented with 10% FCS, 2 mM L-Glutamine (Fisher Scientific #25030081), 10 mM HEPES (Sigma, #H0887-100ML), 1X MEM Non-essential Amino Acids (Fisher, #11140050), 1 mM Sodium Pyruvate (Fisher Scientific #11360070), 100 U/mL Penicillin-Streptomycin (Sigma, #P4333-100ML), and 50 U/mL IL-2 (Amerisource Bergen, #10101641). Both cell subsets were then stimulated with ImmunoCult™ Human CD3/CD28/CD2 T Cell Activator (STEMCELL Technologies, #10990) at 25 uL/mL for Tregs and 6.25 uL/mL for Teff. Following activation and electroporation, cells were split 1:2 every 48 hours to maintain an approximate density of 1x106 cells/mL and supplemented with respective doses of IL-2.

extract_protocol:
8 and 10 days after editing, respectively, resting and 48-hour restimulated cells were pelleted and resuspended at 1x106 cells per 300 µL of RNA lysis buffer (Zymo, #R1060-1-100). Cells were pipette mixed and vortexed to lyse and frozen at -80 until RNA isolation was performed. RNA was isolated using the Zymo-Quick RNA micro prep kit (#R1051) according to the manufacturer’s protocol with the following modifications: After thawing the samples, each sample was vortexed vigorously to ensure total lysis prior to loading into the extraction columns. The optional kit provided DNAse step was skipped, and instead RNA was eluted from the isolation column after the recommended washes and digested with Turbo-DNAse (Fisher Scientific, AM2238) at 37 C for 20 minutes. Following digestion, RNA was purified using the RNA Clean & Concentrator-5 kit (Zymo, #R1016) according to the manufacturer’s protocol.
The purified RNA was submitted to the UC Davis DNA Technologies and Expression Analysis Core to generate 3′ Tag-seq libraries with unique molecular indices (UMIs). Barcoded sequencing libraries were prepared using the QuantSeq FWD kit (Lexogen) for multiplexed sequencing.

treatment_protocol: Guide loaded Cas9 RNPs were assembled with custom crRNAs (Dharmacon) which were resuspended in IDT duplex buffer (IDT, #11-01-03-01) at 160uM. Dharmacon Edit-R CRISPR-Cas9 Synthetic tracrRNA (Dharmacon, #U-002005-20) also resuspended in Nuclease Free Duplex Buffer at 160uM was combined at a 1:1 molar ratio in a 96 well plate and incubated at 37°C for 30 minutes. Single-stranded donor oligonucleotides (ssODN; sequence: TTAGCTCTGTTTACGTCCCAGCGGGCATGAGAGTAACAAGAGGGTGTGGTAATATTACGGTACCGAGCACTATCGATACAATATGTGTCATACGGACACG, 100uM stock) was added to the complex at a 1:1 molar ratio and incubated at 37°C for 5 minutes. Finally, Cas9 protein (MacroLab, Berkeley, 40 µM stock) was added at a 1:2 molar ratio and incubated at 37°C for 15 minutes. The resulting RNPs were frozen at -80°C until the day of electroporation and were thawed to room temperature prior to use. 48 hours following T cell activation, the cells were pelleted at 100x g for 10 minutes and resuspended in room temperature P3 Primary Cell Nucleofector Buffer (Lonza, catalog no. V4XP-3032) at 1.5E6 cells per 17.8 µL. 1.5E6 cells were transferred to each RNP containing well and mixed gently. 25 µL of the combined RNP cell solution was transferred to a 96-well electroporation cuvette plate (Lonza, #VVPA-1002) and nucleofected with pulse code DS-137. Immediately following electroporation, the cells were gently resuspended in 90 µL warmed media and incubated at 37 C for 15 minutes. After recovery, the cells were cultured in 96 well round-bottom plates at 1x106 cells/mL for the duration of the experiment. To prevent edge effects, the guides were randomly distributed across each plate and the first and last columns and rows of each plate was filled with PBS to prevent evaporation. CRISPR-Cas9 edited cells were restimulated 48 hours prior to collection for stimulation response arrayed assays using ImmunoCult™ Human CD3/CD28/CD2 T Cell Activator (STEMCELL Technologies, #10990)

value: Lymphocyte

value: CD4+ Teff

tag: crispr experiment

value: CRISPR KO RNA

tag: sgrna

tag: group

tag: condition

value: Stimulated

tag: donor


title: ECA701, parental, rep1

value: whole animal

value: wildtype

tag: strain


title: N2, parental, rep1

value: parental


title: TG-treated CRISPR screen - input - biological replicate 1


title: N6215CT96h


title: D3_Teff_Resting_MED24_KO_RNA

value: Resting


title: N6203HS48h


title: D1_Teff_Stimulated_MED30_KO_RNA


title: N6203CT48h


title: N6215HS48h


title: D3_Teff_Resting_MED31_KO_RNA


title: D3_Teff_Resting_AAVS1_3_KO_RNA


title: D3_Teff_Resting_MED14_KO_RNA


title: D1_Teff_Stimulated_MED12_KO_RNA


title: N6215CT48h


title: hiPSC-derived NPC, M3_CTR_36S, IL-6 [IL6_36S_NPC]

city: London

first: Amalie

middle: Charlotte Maria

last: Couch

address: 5 Cutcombe Road

department: Basic and Clinical Neuroscience

country: United Kingdom

institute: King's College London

data_processing:
Paired-end fastq files were aligned to the human reference genome (GRCh38) with STAR (Dobin et al., 2013).
Read count extraction was prepared and filtered for counts ≥ 1 using featureCounts (Liao et al., 2014) from the Rsubread (Yang Liao et al., 2019) package, version 2.4.3.
Assembly: GRCh38 - hg38
Supplementary files format and content: tab-deliminated text file includes raw counts for each Sample, plus annotation values for entrezgene IDs, ensembl IDs, HGNC symbols, chromosome location, start and end position, strand, band and gene biotype.

growth_protocol: hiPSC-derived MGLs were differentated to 14 days following the protocol published by Haenseler et al. (2017), and hiPSC-derived NPCs following the dual SMAD inhibition protocol published by Shi et al., (2012). Cells were independently differentiated until day 16 (NPC) and 12 (MGL) of their differentiation protocols, were they were assembled into a transwell system prior to IL-6 exposure.

extract_protocol:
RNA was extracted using Trizol Reagent (Invitrogen; 15596026) following the manufacturers protocol.
Libraries were prepared using a polyA selection method using the NEBNext Ultra II RNA Library Prep Kit for Illumina following manufacturer’s instructions (NEB, Ipswich, MA, USA).

treatment_protocol: Cells were treated for 24h with 100ng/ml IL-6 (Gibco; PHC0066) or 100pM Acetic Acid as per manufacturers reconsitution instruction.

value: hiPSC-derived neural progenitor cell

value: Control

value: 100ng/ml IL-6


title: hiPSC-derived NPC, 127_CTM_01, Vehicle [Veh_127_NPC]

value: 100pM Acetic Acid Vehicle


title: hiPSC-derived NPC, 014_CTM_02, IL-6 [IL6_014_NPC]


title: hiPSC-derived NPC, 069_CTF_01, Vehicle [Veh_069_NPC]


title: hiPSC-derived NPC, 069_CTF_01, IL-6 [IL6_069_NPC]


title: hiPSC-derived NPC, 014_CTM_02, Vehicle [Veh_014_NPC]


title: hiPSC-derived NPC, 127_CTM_01, IL-6 [IL6_127_NPC]


title: hiPSC-derived MGL, 069_CTF_01, Vehicle [Veh_069_MGL]

value: hiPSC-derived microglia-like cell


title: D3_Teff_Resting_MED11_KO_RNA


title: hiPSC-derived MGL, 069_CTF_01, IL-6 [IL6_069_MGL]


title: N6203HS24h


title: hiPSC-derived MGL, 014_CTM_02, IL-6 [IL6_014_MGL]


title: hiPSC-derived MGL, 127_CTM_01, IL-6 [IL6_127_MGL]


title: D3_Teff_Resting_MED30_KO_RNA


title: N6203CT24h


title: D3_Teff_Resting_MED12_KO_RNA


title: hiPSC-derived MGL, 127_CTM_01, Vehicle [Veh_127_MGL]


title: hiPSC-derived NPC, M3_CTR_36S, Vehicle [Veh_36S_NPC]


title: N6215CT24h


title: N6215HS24h


title: D3_Teff_Resting_AAVS1_1_KO_RNA


title: SUM149CR, DMSO, day7

city: Boston

first: Kornelia

last: Polyak

state: MA

address: 450 Brookline Ave

department: Medical Oncology

institute: Dana-Farber Cancer Institute

description: 10X Genomics: 3' v3.1

data_processing:
Raw bcl2 files were converted to fastq using cellranger mkfastq (Cell Ranger v5.0.1). Count information was obtained using cellranger count and aligned to the GRCh38 reference transcriptome provided by 10X: refdata-gex-GRCh38-2020-A.tar.gz. The filtered h5 files output from cellranger count are provided as the processed files.
Assembly: GRCh38
Supplementary files format and content: h5 files

growth_protocol: SUM149 and SUM149CR were cultured in DMEM/F12 with 5% FBS, 5 µg/mL Insulin, 1 µg/mL Hydrocortisone, 100 U/mL penicillin and 100 µg/mL streptomycin. All cells were maintained in a humidified atmosphere with 5% CO2 at 37C.

extract_protocol:
Cells were trypsinized and counted. 1 million cells were pelleted, washed 1 time in ice cold 0.04% BSA in PBS, then resuspended in 1 mL of ice cold 0.04% BSA in PBS.
Library was constructed according to the manufacturer's (single cell 3' v3.1)

treatment_protocol: Cells were seeded in 10 cm dishes in media containing 0.01% DMSO or 10 µM C70 and treated for 7 days.

value: Human breast cancer cell line

value: DMSO


title: SUM149, C70, day7

description: 10X Genomics: 3' v3.1


title: D2_Teff_Resting_MED14_KO_RNA


title: NHP3, control

first: Yantao

last: Duan

address: 270 Dong'an Road

institute: Fudan University Shanghai Cancer Center

data_processing:
Raw data (raw reads) were processed using Trimmomatic. The reads containing ploy-N and the low quality reads were removed to obtain the clean reads. Then the clean reads were mapped to reference genome using hisat2.
Assembly: The reads were reassembled using StringTie.
Supplementary files format and content: counts_anno.txt file includes raw counts for each sample.
Supplementary files format and content: FPKM_anno.txt file includes FPKM values for each sample.

source_name: MGC 803

growth_protocol: MGC803 cells were cultured in Dulbecco’s Modified Eagle Medium (DMEM) with 10% FBS at 37°C in a humidified atmosphere of 5% CO2

extract_protocol:
Total RNA was extracted using the mirVana miRNA Isolation Kit (Ambion) following the manufacturer’s protocol. Total RNA (4μg) was used for the construction of sequencing libraries.
The libraries were constructed using TruSeq Stranded mRNA LTSample Prep Kit (Illumina, San Diego, CA, USA) according to the manufacturer’s instructions.

value: MGC 803

value: Gastric cancer

value: WT

value: control


title: SUM149CR, C70, day7

description: 10X Genomics: 3' v3.1


title: NHP1, control


title: BCH_002_hG1E

first: Alexis

last: Caulier

state: Massachusetts

address: 1 Blackfan street

department: Oncology/Hematology

institute: Boston Children's Hospital

description: demultiplexed with BCL Convert

data_processing:
Files were demultiplexed with BCL convert then aligned with Cellranger
Assembly: GRCh38
Supplementary files format and content: h5 matrix

source_name: erythroid cells

growth_protocol: Erythroid media

extract_protocol: 10x Genomics 3' v3.1

value: CD34+ cells from DBA patients

value: erythroid cells

value: transgene


title: D2_Teff_Resting_AAVS1_3_KO_RNA

value: Lymphocyte

value: CD4+ Teff

value: CRISPR KO RNA


title: BCH_008_GFP

description: demultiplexed with BCL Convert


title: NHP2, control


title: D2_Teff_Resting_MED31_KO_RNA

value: Resting


title: hiPSC-derived MGL, 014_CTM_02, Vehicle [Veh_014_MGL]

value: Control


title: SUM149, ZBTB7A_KO, C70, day7, rep2

data_processing:
All RNA-seq data was analyzed using the VIPER pipeline (Cornwell et al., 2018). Briefly, the STAR aligner (Dobin et al., 2013) was used to align fastq files to the human reference GRCh37/hg19 genome and produce raw gene counts. Cufflinks (Trapnell et al., 2010) was used to create normalized gene counts in terms of FPKM.
Assembly: hg19
Supplementary files format and content: CSV (comma-separated values) matrix files that include raw counts for each sample.
Supplementary files format and content: CSV (comma-separated values) matrix files that include FPKM normalized counts for each sample.

growth_protocol: SUM149 and SUM149CR were cultured in DMEM/F12 with 5% FBS, 5 µg/mL Insulin, 1 µg/mL Hydrocortisone, and 100 µg/mL Penicillin-Streptomycin. MDAMB436 was cultured in McCoy's with 10% FBS, 10 µg/mL Insulin, and 100 µg/mL Penicillin-Streptomycin. MCF7 was cultured in DMEM/F12 with 10% FBS, 10 µg/mL Insulin, and 100 µg/mL Penicillin-Streptomycin. T47D was cultured in RPMI with 10% FBS, 10 µg/mL Insulin, and 100 µg/mL Penicillin-Streptomycin. All cells were maintained in a humidified atmosphere with 5% CO2 at 37C.

extract_protocol:
Total RNA was extracted using the Qiagen RNeasy Mini Kit (Qiagen, #74106) following the manufacturer's protocol.
RNA-seq libraries were prepared using Illumina TruSeq Stranded mRNA sample preparation kits from 500 ng of purified total RNA according to the manufacturer’s protocol. The finished dsDNA libraries were quantified by Qubit fluorometer, Agilent TapeStation 2200, and RT-qPCR using the Kapa Biosystems library quantification kit according to the manufacturer’s protocols.

treatment_protocol: Cells were seeded in 10 cm dishes in media containing 0.01% DMSO or 10 µM C70. Cells were treated for 2 or 7 days before RNA collection.

tag: time

value: 7 days


title: BCH_002_GFP

description: demultiplexed with BCL Convert


title: SUM149, ZBTB7A_KO, DMSO, day7, rep2


title: hiPSC-derived MGL, M3_CTR_36S, IL-6 [IL6_36S_MGL]

value: 100ng/ml IL-6


title: HP3, HP-treated

value: Helicobacter pylori treatment


title: D2_Teff_Resting_MED11_KO_RNA


title: D2_Teff_Resting_MED24_KO_RNA


title: HP2, HP-treated


title: SUM149, ZBTB7A_KO, DMSO, day7, rep1

value: Human breast cancer cell line


title: hiPSC-derived MGL, M3_CTR_36S, Vehicle [Veh_36S_MGL]

value: hiPSC-derived microglia-like cell

value: 100pM Acetic Acid Vehicle


title: HP1, HP-treated

value: MGC 803

value: Gastric cancer


title: SUM149, DMSO, day7

description: 10X Genomics: 3' v3.1

value: DMSO


title: D2_Teff_Resting_MED30_KO_RNA


title: D2_Teff_Resting_MED12_KO_RNA


title: BCH_008_hG1E

description: demultiplexed with BCL Convert

value: CD34+ cells from DBA patients


title: SUM149, ZBTB7A_KO, C70, day7, rep1


title: ACE2-OE3

data_processing:
Sequence reads were trimmed using Cutadapt and bases with average quality scores below 20 were filtered
Sequencing reads were mapped by using HISAT2 with BWT algorithm
Assembly: mm9
Supplementary files format and content: tab-delimited text files include FPKM values for each Sample

growth_protocol: All cells were cultured in Dulbecco’s modified Eagle’s medium (DMEM, Gibco) with 10% FBS (Thermo Fisher Scientific, MA, USA) containing 10% fetal bovine serum (FBS) (Gibco, USA) and 1% penicillin/streptomycin (Gibco) at 5% CO2 at 37°C.

extract_protocol:
RNA was extracted by RNAiso Plus (9109, Takara) following the manufacturer’s instructions, 1.5 ug of total RNA was used to construct sequencing libraries
RNA libraries for RNA-seq were prepared by using Agilent 2100 Bioanalyzer

treatment_protocol: HCC cells with ACE2 overexpression or control (5 × 105 /well) were seeded in the 6-well plate.  For Ang II treatment, 20 nM Ang II (HY-13948, MedChemExpress) was used to treat HCC cells for 24 h before experiments.

value: human heptocellular carcinoma

value: ACE3 overexpression


title: SUM149, ROSA26_KO, C70, day7, rep1


title: SUM149, KDM5B_KO, DMSO, day7, rep2


title: SUM149, ROSA26_KO, DMSO, day7, rep1


title: SUM149, KDM5B_KO, DMSO, day7, rep1


title: D2_Teff_Resting_AAVS1_1_KO_RNA

value: Lymphocyte


title: ACE2-OE2

value: ACE4 overexpression


title: ACE2-OE1

value: ACE2 overexpression


title: SUM149, KDM5B_KO, C70, day7, rep2


title: SUM149, ROSA26_KO, C70, day7, rep2


title: D3_Teff_Stimulated_MED14_KO_RNA

value: CD4+ Teff

value: CRISPR KO RNA

value: Stimulated


title: D3_Teff_Stimulated_MED24_KO_RNA


title: EV3

value: WT

value: control


title: D3_Teff_Stimulated_MED31_KO_RNA


title: SUM149, ROSA26_KO, DMSO, day7, rep2

value: Human breast cancer cell line

value: 7 days


title: SUM149, KDM5A_KO, C70, day7, rep2


title: SUM149, KDM5B_KO, C70, day7, rep1


title: SUM149, KDM5A_KO, DMSO, day7, rep2


title: SUM149, KDM5A_KO, DMSO, day7, rep1


title: EV2


title: SUM149, KDM5A_KO, C70, day7, rep1


title: D3_Teff_Stimulated_AAVS1_3_KO_RNA


title: EV1


title: SUM149, DMSO, day7, rep3


title: D3_Teff_Stimulated_MED11_KO_RNA


title: Input

city: NEW ORLEANS

first: Suzana

middle: D

last: Savkovic

state: LA

address: 1430 Tulane Ave JBJ 315

institute: Tulane University

description:
20220503-20210809- myc-IgG_peaks.annotated- with pvalue and qvalue
myc_peaks.annotated.xlsx

data_processing:
Quality Control.  All read files processed using FastQC (v0.11.7).  All samples contained >50 million reads except for Input (7Savkovic_SimpleChipseq_S7_L001_R1/R2_001.fastq.gz)
Read Alignment.  Read pairs were aligned to hg38 using BWA (v.0.7.15) with default parameters and converted to BAM using Samtools (v.1.5). Aligned reads were deduplicated using MarkDuplicates (Picard, v2.19.0).
Alignment Quality Standards.  ChIP (myc) and control (IgG) samples contained three replicates.  No replicates for Input.  All samples were within or surpassed acceptable levels for NRF (non-redundant fraction=number of distinctly mapping reads after removing duplicates/total reads), and PBC (PCR Bottlenecking Coefficient).
Peak Calling.  Peaks were called using MACS2 (v2.1.2) using either IgG or Input as control and the MACS2 callpeak function with the following parameters: -g hs (for human genome), -B (save fragment pileups into a bedGraph file), --SPMR (save signal per million reads for pileups).  Peak Annotation.  Peaks were annotated (associated with nearby genes) using annotatePeaks.pl (HOMER v.4.11.1) with hg38 and default parameters.
Motif Discovery.  Motifs (de novo and known) were discovered using findMotifsGenome.pl (HOMER v.4.11.1).
Co-occurring Motif Identification.  After filtering known Motifs by p-value (< 1.00e-2) and percent of target sequences with motif (>5%), instances of motifs near called peaks was found using annotatePeaks.pl (HOMER v.4.11.1) and the -m (motifs) parameter.
Peak Density Heatmap.  Generated using deeptools v. 3.5.0 with hg38 reference gene transfer format (.gtf) file and myc pileup file (bigWig) as input.
Assembly: hg38
Supplementary files format and content: Images and excel files of peaks associated with Myc binding and This file shows filtered known motifs associated with called peaks.
Library strategy: CUT&RUN

extract_protocol:
Cells were bound to Concanavalin A-coated magnetic beads, then permeabilized with digitonin. Myc-primary antibody and pAG-MNAse were incubated with cells, and calcium choloride is added to activate the pAG-MNase to initiate DNA digestion. Cleaved DNA was isolated and purified by addng supernatent to spin column
DNA Libraries were prepared using SimpleChIP® ChIP-seq DNA Library Prep Kit for Illumina® #56795 (Cell Signaling Technology).

treatment_protocol: 6hrs oleic acid in Serum Free Media

value: Colon Cancer Cell

value: Input


title: SUM149, DMSO, day7, rep1


title: SUM149, DMSO, day2, rep3

value: 2 days


title: D3_Teff_Stimulated_MED12_KO_RNA


title: D3_Teff_Stimulated_MED30_KO_RNA


title: SUM149, DMSO, day2, rep2

value: DMSO


title: myc-2

description:
20220503-20210809- myc-IgG_peaks.annotated- with pvalue and qvalue
myc_coOccurMotifs

tag: chip antibody

value: CMYC-101AP, Invitrogen

value: 6h Oleic Acid myc test


title: Rab8a-OE_DC

city: Mohali

first: Sharvan

last: Sehrawat

state: Punjab

address: SAS Nagar

department: Biological Sciences

country: India

institute: IISER Mohali

data_processing:
The sequenced raw data of the samples were processed to obtain high quality clean reads using Trimmomatic v0.39 to remove adapter sequences, ambiguous reads (reads with unknown nucleotides “N” larger than 5%), and low-quality sequences (reads with more than 10% quality threshold (QV) < 25 phred score). A minimum length of 100 nt (nucleotide) after trimming was applied. After removing the adapter and low-quality sequences from the raw data, high quality reads were obtained. These high quality (QV>25), paired-end reads were used for reference based read mapping.
Assembly: mm39
Supplementary files format and content: csv file containing raw counts and FPKM values

source_name: CD11b+CD11c+ Dendritic cells

growth_protocol: Bone marrow cells were cultured in a CO2 incubator with 5% CO2 and at 37 degrees celcius temperature. The Bone marrow cells of each group were differentiated into BMDCs by culturing them in the presence of 5ng of GMCSF and IL4. The media was replenished with the cytokines at every alternate day.

extract_protocol:
RNA was extracted using Trizol method.
The paired end sequencing libraries were prepared from the QC passed RNA samples using NEBNext® UltraTM II Directional RNA Library Prep Kit for Illumina (NEB) as per manufacturer’s instruction.

treatment_protocol: The bone marrow cells of aged animals were transduced with lentiviruses either harboring the construct for Rab8a overexperession (Rab8a-OE) or empty vector (Aged). The young cells underwent the same tretment as the aged cells.

value: CD11b+CD11c+ Dendritic cells

value: Rab8a overexpressors


title: Aged_DC

value: Wild-Type


title: myc-3

tag: treatment


title: D2_Teff_Stimulated_MED24_KO_RNA

value: Lymphocyte


title: SUM149, DMSO, day2, rep1


title: DL238, parental, rep1

value: whole animal

value: wildtype

value: Hermaphrodite

tag: developmental stage

value: day 1 adult

value: parental


title: myc-1


title: IgG 3

description:
20220503-20210809- myc-IgG_peaks.annotated- with pvalue and qvalue
myc_peaks.narrowPeak

value: 6h Oleic Acid Control


title: IgG 2


title: GI.1/GII.4 rep3

city: Tianjin

first: Weiqian

last: Dai

address: Xinxing Sub-district, Heping District

institute: Tianjin Medical University

data_processing:
Illumina Casava 1.8 software used for basecalling
Raw data (raw reads) of fastp format were firstly processed through in-house perl scripts..In this step, clean data (clean reads) were obtained by removing reads containing adapter, reads containing ploy-N and low quality reads from raw data. At the same time, Q20, Q30 and GC content the clean data were calculated. All the downstream analyses were based on the clean data with high quality.
Reference genome and gene model annotation files were downloaded from genome website directly. Index of the reference genome was built using Hisat2 v2.0.5 and paired-end clean reads were aligned to the reference genome using Hisat2 v2.0.5.
featureCounts v1.5.0-p3 was used to count the reads numbers mapped to each gene.
Differential expression analysis of two conditions/groups (four biological replicates per condition) was performed using the DESeq2 R package (1.20.0).
Assembly: mm10
Supplementary files format and content: tab-delimited text files include FPKM values and gene count for each Sample
Supplementary files format and content: tab-delimited text files include FPKM values and gene count for each Sample

source_name: draining lymph nodes

extract_protocol:
Total RNA was isolated using TRIzol (Invitrogen) according to the manufacturer’s instructions.
RNA libraries for RNA-seq were prepared using NEBNext® Ultra™ RNA Library Prep Kit for Illumina® following manufacturer's protocols.

value: draining lymph nodes

value: BABL/c

value: 5μg GI.1/GII.4 VLP


title: D2_Teff_Stimulated_MED14_KO_RNA

value: CD4+ Teff

value: CRISPR KO RNA

value: Stimulated


title: GI.1/GII.4 rep4


title: Young_DC


title: SUM149, C70, day7, rep2

value: Human breast cancer cell line

value: WT

value: 7 days


title: SUM149, C70, day7, rep1


title: D2_Teff_Stimulated_MED31_KO_RNA


title: SUM149, DMSO, day7, rep2

value: DMSO


title: D3_Teff_Stimulated_AAVS1_1_KO_RNA

value: Lymphocyte


title: GI.1/GII.4 rep1


title: SUM149, C70, day2, rep3

value: 2 days


title: D2_Teff_Stimulated_MED11_KO_RNA


title: Alum + GI.1/GII.4 rep4

value: 250μg Alum+ 5μg GI.1/GII.4 VLP injected


title: D2_Teff_Stimulated_MED30_KO_RNA


title: SUM149, C70, day7, rep3


title: MDA-MB-231 triple-negative breast cancer cell CDK4-Wild-Type- Biological replicate 4

city: Lausanne

first: Dorian

middle: Vincent

last: ZIEGLER

state: Vaud

address: Quartier UNIL-Sorge - Batiment Gnopode

department: Center for Integrative Genomics (CIG)

country: Switzerland

institute: University of Lausanne (UNIL)

data_processing:
Reads were mapped to the mouse genome GRCh38 (Ensembl version 102) using STAR (v. 2.7.0f; options: --outFilterType BySJout –outFilterMultimapNmax 20 –outMultimapperOrder Random –alignSJoverhangMin 8 –alignSJDBoverhangMin 1 –outFilterMismatchNmax 999 –alignIntronMin 20 –alignIntronMax 1000000 –alignMatesGapMax 1000000). Read counts in gene loci were evaluated with HTSeq-count (v. 0.13.5).
Assembly: Mouse genome GRCh38
Supplementary files format and content: tab-delimited text files include raw counts for each sample

growth_protocol: TNBC MDA-MB-231 (ATCC)mwere cultured in Roswell Park Memorial Institute (RPMI)-1640 medium with GlutaMAX (GibcoTM, Ref: 61870036) supplemented with 10% heat-inactivated FBS (HyClone, Cat. No: SV30160, Lot: RB35956), 10 mM HEPES (GibcoTM, Ref: 15630122) and 1 mM pyruvate (GibcoTM, Ref: 11360070)

extract_protocol:
RNA was isolated by the Trizol-Chloroforme method extraction and subsequent isopropanol-based precipitation, following by two rinsing with EtOH 75%
RNA libraries for RNA seq were prepared using Truseq Stranded RNA Library Prep Kit following manufacturer's protocols

value: Triple-negative breast cancer cells


title: D2_Teff_Stimulated_MED12_KO_RNA


title: MDA-MB-231 triple-negative breast cancer cell CDK4-Wild-Type - Biological replicate 3


title: GI.1/GII.4 rep2


title: MDA-MB-231 triple-negative breast cancer cell CDK4-Wild-Type - Biological replicate 2


title: SUM149CR, DMSO, day7, rep2


title: Alum + GI.1/GII.4 rep2


title: SUM149CR, DMSO, day7, rep3


title: D2_Teff_Stimulated_AAVS1_1_KO_RNA


title: D2_Teff_Stimulated_AAVS1_3_KO_RNA


title: Alum + GI.1/GII.4 rep1


title: IgG 1

value: Colon Cancer Cell

value: CMYC-101AP, Invitrogen


title: D1_Teff_Stimulated_MED24_KO_RNA


title: SUM149CR, DMSO, day7, rep1


title: High LNP + GI.1/GII.4 rep4

value: 1000μg LNPs+ 5μg GI.1/GII.4 VLP injected


title: Alum + GI.1/GII.4 rep3

value: draining lymph nodes


title: SUM149, C70, day2, rep2


title: MDA-MB-231 triple-negative breast cancer cell CDK4-Knockout - Biological replicate 4


title: D1_Teff_Stimulated_MED14_KO_RNA


title: SUM149CR, DMSO, day2, rep3


title: MDA-MB-231 triple-negative breast cancer cell CDK4-Knockout - Biological replicate 3


title: MDA-MB-231 triple-negative breast cancer cell CDK4-Knockout - Biological replicate 2


title: SUM149CR, DMSO, day2, rep2


title: MDA-MB-231 triple-negative breast cancer cell CDK4-Wild-Type - Biological replicate 1


title: High LNP + GI.1/GII.4 rep2

value: BABL/c


title: High LNP + GI.1/GII.4 rep3


title: D1_Teff_Stimulated_MED31_KO_RNA


title: D1_Teff_Stimulated_AAVS1_3_KO_RNA


title: MDA-MB-231 triple-negative breast cancer cell CDK4-Knockout - Biological replicate 1


title: High LNP + GI.1/GII.4 rep1