title: Ovary H3K4me3

data_processing:
Basecalls performed using CASAVA version 1.8
Reads were aligned to the mm10 genome assembly. H3K4me3 ChIP-Seq reads were mapped to the reference genome using bwa aln (0.7.12).The pipeline described in Khil et al. Genome Res. 2012 was used to align SSDS reads to the genome.
Uniquely mapping fragments unambiguously derived from ssDNA (ssDNA type 1) and having both reads with a mapping quality score ≥ 30 were used for identifying hotspot locations (peak calling). NCIS was used to estimate the background fraction for each library. Peak calling was performed using MACS (v.2.1.0.20150420) with the following parameters : --ratio [output from NCIS]  -g mm --bw 1000 --keep-dup all --slocal 5000. DSB hotspots within regions previously blacklisted were removed and hotspot strength was calculated as described previously(17).
Uniquely mapping reads with a mapping quality score ≥ 30 were used for peak calling. NCIS was used to estimate the background fraction relative to an input DNA library. Peak calling was performed using MACS (v.2.1.0.20150420) with the following parameters : --ratio [output from NCIS]  -g mm --bw 1000 --keep-dup all --slocal 5000. Peak strength was subsequently calculated by subtracting the NCIS normalized input read count from the ChIP-Seq read count.
Genome_build: mm10
For each SSDS experiment, the associated BAM file contains all paired end ssDNA reads identified for that sample. Custom BAM tags describe properties of each read pair used for ssDNA detection:
it : ITR length
uh : microhomology length
os : microhomology offset
mm : # mismatches
For each SSDS experiment, a BED file of ssDNA fragments is also provided. Colum 4 gives the alignment phred score for each read (underscore separated). Column 5 gives the ITR-associated detail listed above (it_uh_os). DSB hotspots are provided in bedgraph format for each DMC1 SSDS sample. Background corrected hotspot strength is listed in the fourth column. DSB hotspots in blacklisted regions were not used for analyses.
For the Testis_H3K4me3_BSSeq sample, the processed data file is the bedgraph generated by bismark. This gives the percent methyated reads for each locus.

source_name: Ovary

extract_protocol:
Embryos were collected at E15.5, ovaries were dissected in cold PBS and stored at -80oC until use. 90 or 250 ovaries were fixed in 1 ml PBS with 1% paraformaldehyde for 3 min, quenched and homogenized with Dounce homogenizer.  Cells were collected by 10 minute centrifugation at 900g using a bucket rotor. The pellet was washed in 1 ml of the following buffers: 1) PBS 2) 0.25% Triton X-100, 10 mM EDTA, 0.5 mM EGTA, 10 mM Tris pH8. Cells were lysed in 0.5 ml of the lysis buffer (1% SDS, 10 mM EDTA, 50 mM TrisCl pH8 with complete protein inhibitor cocktail (Roche) and the chromatin was sheared with Misonix sonicator with the following parameters: efficiency 1, 10 s on, 20 s off, total sonication time 4 min. Chromatin was cleared by 10 min centrifugation at 12,000g at 4C. The supernatant was diluted 2-fold by ChiP buffer (0.01% SDS, 1.1% Triton X-100, 1.2 mM EDTA, 16.7 mM TrisHCl , 167 mM NaCl) and dialyzed against the same buffer for 5 hour at 4oC.     Chromatin was incubated with 6µg of custom made anti-DMC1 antibody and 20µl Dynabead beads  (10002D, Invitrogen) at 4oC overnight followed by washing with 500µl of the following buffers: 1) 0.1% SDS, 1% Triton X-100, 2 mM EDTA, 20 mM TrisHCl, 150 mM NaCl; 2) 0.1% SDS, 1% Triton X-100, 2 mM EDTA, 20 mM TrisCl pH8, 500 mM NaCl; 3) 0.25 M LiCl, 1% Igepal, 1 mM EDTA, 10 mM TrisCl, pH8, 1% Deoxycholic acid. DNA protein complexes were eluted by two consecutive 15 min incubations at 650C using elution buffer (0.1M NaHCO3, 1%SDS, 5mM  DTT). The eluates were combined and crosslinking was reversed at 650C for 5 hours. The samples were deproteinized and cleaned up with MinElute PCR purification kit (QIAGEN). The sequencing library was prepared as previously described (Brick et al. 2012).     For H3K4me3 CHiP-Seq and DMC1 SSDS protocols from testis, testicular samples were obtained frozen and were directly thawed in 1% paraformaldehyde and gently dissociated. Testis sample preparation was performed as described previously (Brick et al., 2012).
DMC1 and H3K4Me3 ChIP were performed as described previously (Smagulova et al., 2011; Khil et al., 2012; Brick et al., 2012) with minor modifications.  Sequencing libraries for anti-H3K4Me3 samples were prepared according to manufacturer's protocol (Illumina) and anti-DMC1 libraries were prepared following the method described in (Khil et al., 2012).

value: 15.5 days post coitum

value: wild-type

value: Anti-H3K4me3 Millipore (#07-473)

value: ChIP-Seq

value: For H3K4me3 ChIP-Seq in oocytes, we isolated SCP3 positive meiotic oocytes from 14 15.5 dpc females using BiTS-CHiP54. Briefly, this method uses FACS to isolate nuclei on the basis of the presence of an intra-nuclear marker (in this case, anti-SCP3 (Santa Cruz:sc-74569)). Kapa Hyper Prep kit (catalog #KR0961) was used to prepare the sequencing library due to the limited starting material relative to experiments in whole testis.


title: cortex_systemic_rep3

status: Public on Sep 20 2018

type: RNA

city: Irapuato

first: Ruairidh

middle: J H

last: Sawers

address: Km9.6 Car. N. Irapuato-Leon

country: Mexico

institute: LANGEBIO-CINVESTAV

description: Gene expression data from arbusculated and systemtic cells in mycorrhizal rice roots

hyb_protocol: The labeled cRNA samples were mixed with 0.1 mg•ml–1 sonicated herring sperm DNA in a hybridization buffer containing 100 mM 2-N-morpholinoethanesulfonic acid (Mes), 1 M NaCl, 20 mM EDTA, and 0.01% Tween 20, denatured at 99°C for 5 min, and equilibrated at 45°C for 5 min before hybridization. The hybridization mix was then transferred to the rice GeneChip microarray cartridge and hybridized at 45°C for 16 h on a rotisserie at 60 rpm. The hybridized arrays were then rinsed and stained in a fluidics station (Affymetrix). The arrays were first rinsed with wash buffer A [6× SSPE (0.9 M NaC/0.06 M NaH2PO4/0.006 M EDTA)/0.01% Tween 20/0.005% antifoam] at 25°C for 10 min, incubated with wash buffer B (100 mM Mes/0.1 M NaCl/0.01% Tween 20) at 50°C for 20 min, stained with streptavidin-phycoerythrin (SAPE) (100 mM Mes/1 M NaCl/0.05% Tween 20 containing 10 mg/ml SAPE and 2 mg/ml BSA) at 25°C for 10 min, washed with wash buffer A at 25°C for 20 min, and stained with biotinylated anti-streptavidin antibody at 25°C for 10 min. After staining, arrays were stained with SAPE at 25°C for 10 min and washed with wash buffer A at 30°C for 30 min.

scan_protocol: The probe arrays were scanned twice, and the intensities were averaged with a GeneArray Scanner (Agilent Technologies, Palo Alto, CA) by using MAS 5.0 (Affymetrix).

data_processing: RMA using R/Bioconductor AFFY::rma (normalize  =  TRUE; background  = TRUE)

label: biotin

molecule: total RNA

organism: Oryza sativa Japonica Group

source_name: systemic cells, mycorrhizal rice root

label_protocol: Total RNA (5 m g) from each sample was reverse-transcribed at 42°C for 1 h by using 100 pmol of an oligo(dT)24 primer containing a 5' T7 RNA polymerase promoter sequence [5'-GGCCAGTGAATTGTAATACGACTCACTATAGGGAGGCGG(dT)24-3'], 50 mM Tris•HCl (pH 8.3), 75 mM KCl, 3 mM MgCl2, 10 mM DTT, 0.5 mM dNTPs, and 200 units of SuperScript II reverse transcriptase (Invitrogen). The second strand of cDNA was synthesized by using 40 units of Escherichia coli DNA polymerase I, 10 units of E. coli DNA ligase, and 2 units of RNase H in a reaction containing 25 mM Tris•HCl (pH 7.5), 100 mM KCl, 5 mM MgCl2, 10 mM (NH4)2SO4, 0.15 mM b -NAD+, 1 mM dNTPs, and 1.2 mM DTT. The reaction proceeded at 16°C for 2 h and was terminated by using EDTA. Double-stranded cDNA products were purified by phenol/chloroform extraction and ethanol precipitation. Biotinylated complementary RNAs (cRNAs) were transcribed in vitro from synthesized cDNA by T7 RNA Polymerase (ENZO BioArray High Yield RNA transcript-labeling kit, Enzo Biochem). cRNAs were purified by using affinity resin (Qiagen RNEasy spin columns) and randomly fragmented by incubating at 94°C for 35 min in a buffer containing 40 mM Tris•acetate (pH 8.1), 100 mM potassium acetate, and 30 mM magnesium acetate to produce molecules of » 35–200 bases.

growth_protocol: Rice seeds were surface-sterilized, pre-germinated and grown in a growth chamber with a 12h/12h day/night cycle at 28°C/22°C and 60% humidity. Plants were fertilized every second day with half Hoagland solution, adjusted to 25mM KH2PO4. Plants were harvested at 7 weeks.

extract_protocol: Fresh roots of mycorrhizal and non-mycorrhizal rice plants were collected at 7 wpi, cut into 3-5 mm pieces and immediately fixed, embedded and processed as previously described (Gutjahr et al., 2012). Root sections were arranged in parallel on UV-treated poly-L-lysine slides. A laser microbeam system (P.A.L.M. Microlaser Technologies, http://www.zeiss.de/ micro-dissection) was utilized for microdissection of cortical root cells. For cutting, the following parameters were selected using P.A.L.M. Robosoftware 2003 (http://www.zeiss.de/microdissection): auto- LPC focus 50, with energy of 95 and a speed of 100. Approximately 2000 cortical cells were isolated as combined cell units. Isolated specimens were catapulted onto the adhesive surface on the inside of the lid of a 0.5ml reaction tube (Carl Zeiss, http://www.zeiss.com/), collected by centrifugation (11 000g for 30 sec) and immediately subjected to 350μl RLT buffer (Qiagen, http:// www.qiagen.com/), 40 units),1μl RNaseOUT (Invitrogen, http:// www.invitrogen.com/) and 3.5 μl β-mercaptoethanol (Sigma-Aldrich, http://www.sigmaaldrich.com/). The sample was vigorously mixed and incubated at 56°C for 5 min to facilitate cell-wall disruption. At this stage, the lysate was either stored at 80°C or further processed for RNA isolation. Total RNA of each cell population was extracted using an RNeasy micro kit (Qiagen) according to the manufacturer’s instructions.

treatment_protocol: Plants in the treatment group were inoculated at planting with 250 spores of Rhizophagus irregularis.

tag: cultivar

value: Nipponbare

tag: inoculation

value: Rhizophagus irregularis

tag: cell type

value: systemic root cortical cell


title: 171219_CEMBA_mm_P56_P63_4B_CEMBA171213_4B_1_CEMBA171213_4B_2_A10_AD008_indexed

type: SRA

city: La Jolla

first: Joseph

middle: R

last: Ecker

state: CA

address: 10010 North Torrey Pines Road

department: Genomic Analysis Laboratory

country: USA

institute: HHMI-Salk-Institute

library_source: genomic

data_processing:
Sequencing reads mapping, quality filtering and the summary of DNA methylation level for each cytosine was performed as previously described in Luo et al., 2017 with minor modifications. Non-clonal mapped reads were filtered for MAPQ > 10 using samtools view -bq10 option.
Supplementary_files_format_and_content: tab delimited text files of methylcytosine calls; columns in allc files are: column 1 - chromosome; column 2 - position; column 3 - strand; column 4 - class; column 5 - mC reads; column 6 - total reads; column 7 - methylated (Boolean value indicating the result of statistical test for methylated cytosines)

molecule: genomic DNA

organism: Mus musculus

source_name: Mouse primary motor cortex

growth_protocol: Male C57Bl/6J mice were purchased from Jackson laboratories at 8 weeks of age and maintained in our facility for 48h before dissection. Animals were maintained in the Salk animal barrier facility on 12h dark-light cycles with food ad-libitum. Human brain specimens were obtained from University of Miami Brain Endowment Bank. snmC-seq with Shrimp Alkaline Phosphatase treatment (snmC-seq + SAP) was applied to frontal cortex (Brodmann Area 10, BA10) tissue of a deceased 58-year-old Caucasian male with PMI = 23.4. snmC-seq2 was applied to BA10 tissue of a deceased 25-year-old Caucasian male with PMI = 20.8.

extract_protocol: Bisulfite-converted samples were denatured at 95°C for 3 minutes, then placed on ice for 2 minutes. 5µL Random Priming master mix [1µL 10x Blue Buffer (Enzymatics B0110), 0.25µL Klenow Exo- (50U/µL, Enzymatics, P7010-HC-L), 0.5µL dNTP (10mM each, NEB N0447L), 3.25µL water] was added and incubated at 4°C for 5 min, 25°C for 5 min, and 37°C for 60 min followed by 4°C. 1.5µL enzyme mix containing Exonuclease 1 (20U/µL, Enzymatics X8010L) and rSAP (1U/µL, NEB M0371L) was added, then incubated at 37°C for 30 min followed by 4°C. 0.8x SPRI beads were added, mixed and incubated for 5 minutes at room temperature to allow DNA to bind. The beads were washed 3 times with 80% ethanol and eluted in 10µL EB buffer (Qiagen 19086). The plates were denatured again at 95°C for 3 minutes, and 10.5µL Adaptase master mix [2µL G1, 2µL G2, 1.25µL G3, 0.5µL G4 and 0.5µL G5 (Swift Biosciences 33096)] was added and incubated at 37°C for 30 minutes, then 95°C for 2 minutes. 25µL 2x KAPA HiFi HotStart ReadyMix (Kapa, KK2602) and 5µL custom indexing primer mix (6µM P5, 10µM P7) were added. The PCR was programmed as follows: 1) 95°C 2 min, 2) 98°C 30 sec, 3) 98°C 15 sec, 4) 64°C for 30 sec, 5) 72°C for 2 min, 6) 72°C 5 min, 7) 4°C hold. Repeat steps 3-5 for 15 total cycles. PCR reactions were cleaned with 0.8X SPRI beads for three rounds. Library concentration was determined with Qubit™ dsDNA BR Assay Kit (ThermoFisher Q32853). Libraries were sequenced using Illumina Novaseq instrument.

tag: Sex

value: Male

tag: age

tag: genome build

tag: library strategy


title: 171213_CEMBA_mm_P56_P63_3C_MOp_CEMBA171207_3C_1_CEMBA171207_3C_2_A2_AD008_indexed


title: Pool_2082_AD010_indexed

organism: Homo sapiens

source_name: Human frontal cortex (BA10)

value: 58 yr

value: snmC-seq + SAP


title: Pool_2068_AD010_indexed


title: 171213_CEMBA_mm_P56_P63_3C_MOp_CEMBA171207_3C_1_CEMBA171207_3C_2_A2_AD007_indexed


title: Pool_2066_AD002_indexed


title: 44D_tumor vs reference

type: genomic

city: Amsterdam

first: Daoud

last: Sie

address: De Boelelaan 1117

department: Pathology

country: Netherlands

institute: Vrije Universiteit Medical Center

description:
raw data file: US22502676_251495010775_S01_CGH_105_Dec08_1_4.txt
raw data file: US22502676_251495010775_S01_CGH_105_Dec08_1_4.txt

hyb_protocol: samples were applied to microarrays enclosed in Agilent SureHyb-enabled hybridization chambers. After hybridization, slides were washed following manufacturer's instructions

scan_protocol: Images were quantified using Agilent Feature Extraction Software (version 10.7.3.1) according to the specifications of the used platforms: 4x44k, 4x105k, 4x180k. All data was processed with HG18.

data_processing: Agilent Feature Extraction Software (v 10.7.3.1) was used for background subtraction. Sample processed data reflects normalized log2 ratio representing test/reference (ch1/ch2) according to sample information.

source_name: tumor

label_protocol: ENZO labeling for Cy3 and Cy5 following manufacturer's instructions

extract_protocol: 10x10mu FFPE sections were cut for DNA isolation according to protocol by HF van Essen et al.  (2012)

tag: histology

value: adenocarcinoma

source_name: reference

value: normal


title: 44C_tumor vs reference

description:
raw data file: US22502676_251495010775_S01_CGH_105_Dec08_1_3.txt
raw data file: US22502676_251495010775_S01_CGH_105_Dec08_1_3.txt


title: Pool_2047_AD010_indexed


title: 171213_CEMBA_mm_P56_P63_3C_MOp_CEMBA171207_3C_1_CEMBA171207_3C_2_A1_AD010_indexed


title: Pool_2065_AD008_indexed


title: 171213_CEMBA_mm_P56_P63_3C_MOp_CEMBA171207_3C_1_CEMBA171207_3C_2_A2_AD001_indexed


title: Pool_2051_AD002_indexed


title: cortex_mock_rep2

source_name: cortical cells, mock-inoculated rice root

value: mock

value: root cortex


title: Pool_2035_AD002_indexed


title: 171213_CEMBA_mm_P56_P63_3C_MOp_CEMBA171207_3C_1_CEMBA171207_3C_2_A1_AD007_indexed


title: 44A_tumor vs reference

description:
raw data file: US22502676_251495010775_S01_CGH_105_Dec08_1_1.txt
raw data file: US22502676_251495010775_S01_CGH_105_Dec08_1_1.txt


title: cortex_arbusculated_rep1

source_name: arbusculated cells, mycorrhizal rice root

value: arbusculated root cortical cell


title: cortex_systemic_rep1


title: cortex_arbusculated_rep3


title: 171213_CEMBA_mm_P56_P63_3C_MOp_CEMBA171207_3C_1_CEMBA171207_3C_2_A12_AD007_indexed


title: Pool_2043_AD002_indexed


title: 171213_CEMBA_mm_P56_P63_3C_MOp_CEMBA171207_3C_1_CEMBA171207_3C_2_A12_AD010_indexed


title: cortex_arbusculated_rep2


title: Pool_2020_AD010_indexed


title: cortex_mock_rep3


title: cortex_mock_rep1


title: Pool_2026_AD010_indexed


title: 44B_tumor vs reference

description:
raw data file: US22502676_251495010775_S01_CGH_105_Dec08_1_2.txt
raw data file: US22502676_251495010775_S01_CGH_105_Dec08_1_2.txt

value: WT

value: positive


title: 171213_CEMBA_mm_P56_P63_3C_MOp_CEMBA171207_3C_1_CEMBA171207_3C_2_A1_AD012_indexed


title: 171213_CEMBA_mm_P56_P63_3C_MOp_CEMBA171207_3C_1_CEMBA171207_3C_2_A12_AD006_indexed


title: Pool_1992_AD006_indexed


title: H4K16ac_ChIP-seq_aged-HSC

city: Heidelberg

first: Daniel

middle: B.

last: Lipka

address: INF 280

department: Epigenomics and Cancer Risk Factors

country: Germany

institute: German Cancer Research Center

data_processing:
Raw reads were trimmed using the trimLinDA script (available at http://www-igbmc.u-strasbg.fr/Gronemeyer)(Shankaranarayanan et al., 2012)
Trimmed reads were mapped to the mouse reference genome (mm10) using the BWA aligner(Li and Durbin, 2009), and subsequently duplicates and low-quality reads (MAQ score <20) were removed using SAMtools(Li et al., 2009).
For peak calling, generation of bedgraphs, genome and gene annotation, we used the Homer software package; the .bed file conforms to the UCSC browser format, but still contains a header line that explains explicitly the content of each column. To view the data in the UCSC genome browser one would have to delete this header line. (http://biowhat.ucsd.edu/homer/)(Heinz et al., 2010). The histone style parameters and the appropriate input sample were used to detect H4K16ac peaks.
Genome_build: mm10
Supplementary_files_format_and_content: BED files and BEDGRAPH files were generated using the Homer software package; the .bed file conforms to the UCSC browser format, but still contains a header line that explains explicitly the content of each column. To view the data in the UCSC genome browser one would have to delete this header line.

extract_protocol:
Cells were fixed in 1% formalin for 10 min at room temperature and fixation was stopped by adding glycine to a final concentration of 125 mM. After three washes with PBS, cell pellets were snap-frozen and stored at -80°C. For sonication, cell pellets were resuspended in lysis buffer (1% SDS, 10 mM EDTA, 50 mM Tris, pH 8.1) supplemented with protease inhibitor cocktail (Roche) and processed in a Covaris S2 instrument using empirically established conditions that resulted in chromatin fragments between 150 and 300 bp in length (Duty Cycle 5%, Intensity 4, Cycles/Burst 200; 40 cycles: 30 sec on, 20 sec off). Sonicated chromatin was cleared by centrifugation for 10 min at 13000 rpm. Ten percent of cleared chromatin was taken as input control. The H4K16ac antibody (07-329, Millipore) was coupled to protein A/G magnetic beads and washed 2x with ChIP dilution buffer (0.01% SDS, 1.1% Triton X-100, 1.2 mM EDTA, 16.7 mM Tris-HCl, pH 8.1, 167 mM NaCl). Sonicated chromatin was first diluted 10-fold with ChIP dilution buffer, then the antibody-bead conjugates were added and incubated at 4°C over night. Beads were first washed 2x with Low Salt Immune Complex Wash Buffer (0.1% SDS, 1% Triton X-100, 2mM EDTA, 20mM Tris-HCl, pH 8.1, 150mM NaCl) and then washed 2x with LiCl Immune Complex Wash Buffer (0.25M LiCl, 1% IGEPAL CA630, 1% deoxycholic acid (sodium salt), 1mM EDTA, 10mM Tris, pH 8.1). Then precipitated chromatin was eluted and decrosslinked (2% SDS, 0.1 M NaHCO3, 0.25 M NaCl) for 4 h at 65°C and afterwards treated with proteinase K for 1 h at 45°C. DNA was purified using SPRI beads (Agencourt AMPure XP, Beckman Coulter) at a ratio of 1.8/1 (beads/DNA) and eluted in 15 µl ddH2O. Linear DNA amplification (LinDA) of ChIPed DNA was performed as described (Shankaranarayanan et al., 2011). Briefly, ChIPed DNA was dephosphorylated for 10 min at 37°C using 1 U TSAP (M9910, Promega) in NEB 4 buffer (B7004, NEB) followed by heat inactivation for 15 min at 75°C. DNA was then T-tailed for 20 min at 37°C in the presence of 5 µM T-mix (92 % dTTP, 8 % ddCTP) and 0.25 mM CoCl2 using 20 U terminal transferase (M0252, NEB) followed by heat inactivation for 10 min at 70°C. Next, 5 pmol T7-BpmI-oligo(A)14 primer (AATTAATACGACTCACTATAGGGCTGGAGAAAAAAAAAAAAAA) was annealed at 37°C for 5 min and then double strand synthesis was performed for 55 min at 37°C using 10 U Klenow fragment (M0210, NEB), 0.2 mM dNTPs. After heat inactivation, DNA was purified using SPRI beads at a ratio of 1.8/1 (beads/DNA) and eluted in 12.5 µl ddH2O. In vitro transcription was performed over night using the RNAMaxx high yield kit (200339, Stratagene) and resulting RNA was purified using SPRI beads (Agencourt AMPure XP, Beckman Coulter; beads/DNA ratio 1.8/1) followed by a reverse transcription (1h, 42°C; 10 min, 75°C) using the T7-BpmI-oligo(A)14 primer and the Superscript III reverse transcription kit (18080044, Invitrogen). Second strand synthesis was performed using a custom reaction (20 µl first strand cDNA, 10 µl 10x thermopol buffer, 1 ml 100x BSA, 3 ml 10 mM dNTPs, 0.5 µl RnaseH, 1 µl Taq polymerase, 0.1 µl Pfu polymerase, 64.4 ml H2O) which was incubated in a thermocycler under the following conditions: 37°C 5 min, 65°C 1 min, 72°C 30 min) followed by SPRI purification (beads/DNA ratio 1.8/1). Resulting double-stranded DNA was then digested with BpmI for 2 h at 37°C, purified again with SPRI beads (beads/DNA ratio 1.5/1) and eluted in 13 µl ddH2O.
Illumina sequencing libraries were prepared using the NEBNext Ultra DNA Library Prep Kit for Illumina (E7370, NEB) and the NEBNext Multiplex-Oligos for Illumina (E7335, NEB) following the manufacturer’s protocol. Since for all samples, input amount of DNA was below 100 ng, the adaptors were diluted 1:10 in sterile water before adaptor ligation. Cleanup of adaptor-ligated DNA was performed without size selection using a 1.0/1 ratio of SPRI beads/DNA. For the PCR amplification of libraries, 0.5 µl of 100x Syber Green was added to each reaction and the PCR amplification was performed on a LightCycler 480 (Roche) using the following cycling conditions: initial denaturation (98°C, 30 sec) followed by 9-12 cycles (98°C, 10 sec; 65°C, 30 sec; 72°C, 30 sec). The PCR reaction was stopped, when the amplification curves had reached a fluorescence intensity of about 10. The PCR reactions were purified using SPRI beads at a ratio of 1.0/1 (beads/DNA). Sequencing libraries were multiplexed and subjected to 50 bp single-end sequencing on an Illumina HiSeq 2000 instrument (Illumina Inc., San Diego).

treatment_protocol: Sorted young, aged, aged±CASIN 5µM HSCs where incubated for 16 hrs in IMDM+10%FBS at 37°C, 5%CO2, 3%O2.

tag: strain

value: hematopoietic stem cells

tag: cell status

value: aged

tag: chip antibody

value: H4K16ac (Millipore, 07-329)


title: H4K16ac_ChIP-seq_rejuvenated-HSC

value: rejuvenated


title: input_DNA


title: 171213_CEMBA_mm_P56_P63_3C_MOp_CEMBA171207_3C_1_CEMBA171207_3C_2_A12_AD004_indexed


title: 171213_CEMBA_mm_P56_P63_3C_MOp_CEMBA171207_3C_1_CEMBA171207_3C_2_A11_AD010_indexed


title: Testis SSDS Dnmt3Lko

city: Bethesda

first: Kevin

last: Brick

state: MD

address: 5/205 Memorial Drive

department: GBB

institute: NIDDK

source_name: Testis

value: 30 days post partum

tag: genotype

tag: antibody

value: Anti-DMC1 Santa Cruz (C-20, sc 8973)

tag: sequencing technique

value: ChIP-Single Stranded DNA Sequencing (SSDS)


title: Pool_2015_AD006_indexed

value: Male


title: H4K16ac_ChIP-seq_young-HSC

value: young


title: Testis SSDS Prdm9ko

description: Data from GSM869790

value: Adult


title: 171213_CEMBA_mm_P56_P63_3C_MOp_CEMBA171207_3C_1_CEMBA171207_3C_2_A11_AD012_indexed


title: Pool_1985_AD006_indexed

value: 58 yr

value: snmC-seq + SAP


title: Ovary Input SSDS

data_processing:
Library strategy: DNA-seq
Basecalls performed using CASAVA version 1.8
Reads were aligned to the mm10 genome assembly. H3K4me3 ChIP-Seq reads were mapped to the reference genome using bwa aln (0.7.12).The pipeline described in Khil et al. Genome Res. 2012 was used to align SSDS reads to the genome.
Uniquely mapping fragments unambiguously derived from ssDNA (ssDNA type 1) and having both reads with a mapping quality score ≥ 30 were used for identifying hotspot locations (peak calling). NCIS was used to estimate the background fraction for each library. Peak calling was performed using MACS (v.2.1.0.20150420) with the following parameters : --ratio [output from NCIS]  -g mm --bw 1000 --keep-dup all --slocal 5000. DSB hotspots within regions previously blacklisted were removed and hotspot strength was calculated as described previously(17).
Uniquely mapping reads with a mapping quality score ≥ 30 were used for peak calling. NCIS was used to estimate the background fraction relative to an input DNA library. Peak calling was performed using MACS (v.2.1.0.20150420) with the following parameters : --ratio [output from NCIS]  -g mm --bw 1000 --keep-dup all --slocal 5000. Peak strength was subsequently calculated by subtracting the NCIS normalized input read count from the ChIP-Seq read count.
Genome_build: mm10
For each SSDS experiment, the associated BAM file contains all paired end ssDNA reads identified for that sample. Custom BAM tags describe properties of each read pair used for ssDNA detection:
it : ITR length
uh : microhomology length
os : microhomology offset
mm : # mismatches
For each SSDS experiment, a BED file of ssDNA fragments is also provided. Colum 4 gives the alignment phred score for each read (underscore separated). Column 5 gives the ITR-associated detail listed above (it_uh_os). DSB hotspots are provided in bedgraph format for each DMC1 SSDS sample. Background corrected hotspot strength is listed in the fourth column. DSB hotspots in blacklisted regions were not used for analyses.
For the Testis_H3K4me3_BSSeq sample, the processed data file is the bedgraph generated by bismark. This gives the percent methyated reads for each locus.

value: 15.5 days post coitum

value: wild-type

value: Single Stranded DNA Sequencing (SSDS)


title: eye_diabets_rep3

city: Beijing

first: Qian

last: Zhang

address: No. 1 Shuaifuyuan, Dongcheng District

country: China

institute: Peking Union Medical College Hospital

hyb_protocol: The sample preparation and hybridization of each cDNAs on Affymetrix GeneChip® Mouse Gene 2.0 ST arrays based on manufacturer's instruction.

scan_protocol: Arrays were scanned using the GeneChip® Scanner 3000 (Affymetrix).

data_processing: All arrays were uploaded into GeneSpring v12.5 (Agilent Technologies, USA) for data normalization, quality control and first-pass analysis. Differentially expressed genes were then identified through fold change as well as P value calculated with t-test. The threshold set for differentially regulated genes was a fold change>1.5 and a P value<0.05.

organism: Rattus norvegicus

source_name: diabetic rats, 17week

label_protocol: biotin

extract_protocol: Total tissue RNA was extracted using total RNA Isolation Kit (mirVanaTM, ABI).

tag: disease status

value: diabetic

tag: tissue

value: retina

value: 17 weeks


title: 171213_CEMBA_mm_P56_P63_3C_MOp_CEMBA171207_3C_1_CEMBA171207_3C_2_A11_AD004_indexed


title: Pool_1872_AD010_indexed


title: Testis Input 30dpp Dnmt3l-/-

value: DNA-Seq


title: Testis H3K4me3 30dpp Dnmt3l-/-

value: Anti-H3K4me3 Millipore (#07-473)

value: ChIP-Seq


title: 171213_CEMBA_mm_P56_P63_3C_MOp_CEMBA171207_3C_1_CEMBA171207_3C_2_A11_AD007_indexed


title: 171213_CEMBA_mm_P56_P63_3C_MOp_CEMBA171207_3C_1_CEMBA171207_3C_2_A11_AD002_indexed


title: Pool_1872_AD002_indexed


title: 171213_CEMBA_mm_P56_P63_3C_MOp_CEMBA171207_3C_1_CEMBA171207_3C_2_A11_AD001_indexed


title: Pool_1859_AD006_indexed


title: Testis H3K4me3 30dpp wt

value: 30 days post partum


title: 14A_tumor vs reference

description:
MAL_251901510430_S01_CGH_105_Dec08_1_1.txt
MAL_251901510430_S01_CGH_105_Dec08_1_1.txt

value: small cell (lung) carcinoma

value: normal


title: Pool_1842_AD006_indexed

value: Male


title: Ovary SSDS Sample 2

value: Anti-DMC1 GP1 custom

value: ChIP-Single Stranded DNA Sequencing (SSDS)


title: Pool_1856_AD006_indexed

value: 58 yr


title: 171213_CEMBA_mm_P56_P63_3C_MOp_CEMBA171207_3C_1_CEMBA171207_3C_2_A10_AD012_indexed


title: 14B_tumor vs reference

description:
MAL_251901510430_S01_CGH_105_Dec08_1_2.txt
MAL_251901510430_S01_CGH_105_Dec08_1_1.txt


title: Pool_1822_AD010_indexed

value: snmC-seq + SAP


title: 171213_CEMBA_mm_P56_P63_3C_MOp_CEMBA171207_3C_1_CEMBA171207_3C_2_A10_AD010_indexed


title: Pool_1819_AD006_indexed


title: eye_diabets_rep2


title: Danshen-treated_rep2

source_name: Danshen-treated rats, 17week

tag: treatment

value: Danshen


title: Danshen-treated_rep3


title: 171213_CEMBA_mm_P56_P63_3C_MOp_CEMBA171207_3C_1_CEMBA171207_3C_2_A10_AD008_indexed


title: Danshen-treated_rep1


title: eye_diabets_rep1


title: 73A_tumor vs reference

description:
raw data file: US22502676_251901510369_S01_CGH_105_Dec08_1_1.txt
raw data file: US22502676_251901510369_S01_CGH_105_Dec08_1_2.txt

value: squalors cell carcinoma


title: Pool_1818_AD002_indexed


title: Testis Input SSDS

description: Data from GSM1954850, GSM1954851, GSM1954852, GSM1954853, GSM1954854

value: Various

value: Adult

value: wild-type

value: Single Stranded DNA Sequencing (SSDS)


title: 171213_CEMBA_mm_P56_P63_3C_MOp_CEMBA171207_3C_1_CEMBA171207_3C_2_A10_AD007_indexed


title: 171213_CEMBA_mm_P56_P63_3C_MOp_CEMBA171207_3C_1_CEMBA171207_3C_2_A10_AD006_indexed


title: 43B_tumor vs reference

description:
raw data file: MAF_251901511426_S01_CGH-v4_95_Feb07_1_2.txt
raw data file: MAF_251901511426_S01_CGH-v4_95_Feb07_1_2.txt


title: Pool_1795_AD006_indexed


title: 43A_tumor vs reference

description:
raw data file: MAF_251901511426_S01_CGH-v4_95_Feb07_1_1.txt
raw data file: MAF_251901511426_S01_CGH-v4_95_Feb07_1_2.txt


title: Pool_1788_AD010_indexed


title: 171213_CEMBA_mm_P56_P63_3C_MOp_CEMBA171207_3C_1_CEMBA171207_3C_2_A10_AD004_indexed


title: 13B_tumor vs reference

description:
raw data file: MAF_251901511424_S01_CGH_105_Dec08_1_2.txt
raw data file: MAF_251901511424_S01_CGH_105_Dec08_1_1.txt

value: adenocarcinoma


title: Ovary SSDS Sample 1

value: 15.5 days post coitum


title: Testis SSDS Sample 6

value: Anti-DMC1 Santa Cruz (C-20, sc 8973)


title: Testis SSDS Sample 9

description: Data from GSM869782


title: 73B_tumor vs reference

description:
raw data file: US22502676_251901510369_S01_CGH_105_Dec08_1_2.txt
raw data file: US22502676_251901510369_S01_CGH_105_Dec08_1_2.txt


title: Pool_1775_AD010_indexed


title: Testis SSDS Sample 5


title: Pool_1787_AD006_indexed


title: 171213_CEMBA_mm_P56_P63_3C_MOp_CEMBA171207_3C_1_CEMBA171207_3C_2_A10_AD001_indexed


title: Testis SSDS Sample 4


title: 72B_tumor vs reference

description:
raw data file: MAF_251901510151_S01_CGH_105_Dec08_1_2.txt
raw data file: MAF_251901510151_S01_CGH_105_Dec08_1_1.txt


title: 42B_tumor vs reference

description:
raw data file: MAF_251901510150_S01_CGH_105_Dec08_1_2.txt
raw data file: MAF_251901510150_S01_CGH_105_Dec08_1_1.txt


title: Testis SSDS Sample 3


title: Pool_1758_AD010_indexed


title: Pool_1769_AD006_indexed


title: 171213_CEMBA_mm_P56_P63_3C_MOp_CEMBA171206_3C_1_CEMBA171206_3C_3_A1_AD012_indexed


title: Testis SSDS Sample 8


title: 171213_CEMBA_mm_P56_P63_3C_MOp_CEMBA171206_3C_1_CEMBA171206_3C_3_A1_AD010_indexed


title: Testis SSDS Sample 2


title: Testis SSDS Sample 7


title: 171213_CEMBA_mm_P56_P63_3C_MOp_CEMBA171206_3C_1_CEMBA171206_3C_3_A1_AD008_indexed


title: 72A_tumor vs reference

description:
raw data file: MAF_251901510151_S01_CGH_105_Dec08_1_1.txt
raw data file: MAF_251901510151_S01_CGH_105_Dec08_1_1.txt


title: 171213_CEMBA_mm_P56_P63_3C_MOp_CEMBA171206_3C_1_CEMBA171206_3C_3_A1_AD007_indexed


title: Pool_1751_AD006_indexed


title: Pool_1750_AD006_indexed


title: 13A_tumor vs reference

description:
raw data file: MAF_251901511424_S01_CGH_105_Dec08_1_1.txt
raw data file: MAF_251901511424_S01_CGH_105_Dec08_1_1.txt


title: 171213_CEMBA_mm_P56_P63_3C_MOp_CEMBA171207_3C_1_CEMBA171207_3C_2_A10_AD002_indexed


title: Testis SSDS 200k cells


title: 171213_CEMBA_mm_P56_P63_3C_MOp_CEMBA171206_3C_1_CEMBA171206_3C_3_A1_AD006_indexed


title: 42A_tumor vs reference

description:
raw data file: MAF_251901510150_S01_CGH_105_Dec08_1_1.txt
raw data file: MAF_251901510150_S01_CGH_105_Dec08_1_1.txt


title: Testis SSDS Sample 1

description: Data from GSM869781


title: 12B_tumor vs reference

description:
raw data file: MAF_251901510153_S01_CGH_105_Dec08_1_1.txt
raw data file: MAF_251901510153_S01_CGH_105_Dec08_1_1.txt

value: large cell carcinoma


title: Testis SSDS Hop2ko

data_processing:
Library strategy: Single Stranded DNA Sequencing (SSDS)
Basecalls performed using CASAVA version 1.8
Reads were aligned to the mm10 genome assembly. H3K4me3 ChIP-Seq reads were mapped to the reference genome using bwa aln (0.7.12).The pipeline described in Khil et al. Genome Res. 2012 was used to align SSDS reads to the genome.
Reads from Testis_H3K4me3_BSSeq were aligned to the mm10 reference using bismark 0.19.0.
Uniquely mapping fragments unambiguously derived from ssDNA (ssDNA type 1) and having both reads with a mapping quality score ≥ 30 were used for identifying hotspot locations (peak calling). NCIS was used to estimate the background fraction for each library. Peak calling was performed using MACS (v.2.1.0.20150420) with the following parameters : --ratio [output from NCIS]  -g mm --bw 1000 --keep-dup all --slocal 5000. DSB hotspots within regions previously blacklisted were removed and hotspot strength was calculated as described previously(17).
Uniquely mapping reads with a mapping quality score ≥ 30 were used for peak calling. NCIS was used to estimate the background fraction relative to an input DNA library. Peak calling was performed using MACS (v.2.1.0.20150420) with the following parameters : --ratio [output from NCIS]  -g mm --bw 1000 --keep-dup all --slocal 5000. Peak strength was subsequently calculated by subtracting the NCIS normalized input read count from the ChIP-Seq read count.
Genome_build: mm10
For each SSDS experiment, the associated BAM file contains all paired end ssDNA reads identified for that sample. Custom BAM tags describe properties of each read pair used for ssDNA detection:
it : ITR length
uh : microhomology length
os : microhomology offset
mm : # mismatches
For each SSDS experiment, a BED file of ssDNA fragments is also provided. Colum 4 gives the alignment phred score for each read (underscore separated). Column 5 gives the ITR-associated detail listed above (it_uh_os). DSB hotspots are provided in bedgraph format for each DMC1 SSDS sample. Background corrected hotspot strength is listed in the fourth column. DSB hotspots in blacklisted regions were not used for analyses.
For the Testis_H3K4me3_BSSeq sample, the processed data file is the bedgraph generated by bismark. This gives the percent methyated reads for each locus.

extract_protocol:
For H3K4me3 CHiP-Seq and DMC1 SSDS protocols from testis, testicular samples were obtained frozen and were directly thawed in 1% paraformaldehyde and gently dissociated. Testis sample preparation was performed as described previously (Brick et al., 2012).
DMC1 and H3K4Me3 ChIP were performed as described previously (Smagulova et al., 2011; Khil et al., 2012; Brick et al., 2012) with minor modifications.  Sequencing libraries for anti-H3K4Me3 samples were prepared according to manufacturer's protocol (Illumina) and anti-DMC1 libraries were prepared following the method described in (Khil et al., 2012).
For H3K4me3 CHiP-Seq and DMC1 SSDS protocols from testis, testicular samples were obtained frozen and were directly thawed in 1% paraformaldehyde and gently dissociated. Testis sample preparation was performed as described previously (Brick et al., 2012).
DMC1 and H3K4Me3 ChIP were performed as described previously (Smagulova et al., 2011; Khil et al., 2012; Brick et al., 2012) with minor modifications.  Sequencing libraries for anti-H3K4Me3 samples were prepared according to manufacturer's protocol (Illumina) and anti-DMC1 libraries were prepared following the method described in (Khil et al., 2012).


title: N+PD30_3

city: Göttingen

first: Anusha

last: Sriraman

address: Justus-von-liebig Weg 11

department: Department of Molecular Oncology

institute: Georg-August University of Göttingen

library_source: transcriptomic

data_processing:
The FASTQ  files were mapped to the human genome (assembly hg19) using TopHat Gapped-read mapper for RNA-seq data with very-sensitive BOWTIE2 settings on Galaxy Platform (Version 0.9). The read counting was performed via HTSeq (version 0.6.0) with following parameters: -f bam -r pos -s reverse -a 10 -t exon -m union. The count files were subsequently subjected for differential analysis using the DESeq2 Package on R (Bioconductor version 3.2.2). The Wald Test was utilized for a comparisons. The resulting tables for log2fold gene expression changes wrt basemeans are provided for Nutlin_vs_DMSO as CSV files. With the selection of the upregulated and downregulated genes, z-score for all conditions for these genes were calculated and heatmaps were generated. GSEA analysis for NUT_PD_30_vs_NUT and PD30_vs_DMSO was carried out using the basemeans provided here as CSV file.
hg19
The provided mRNA-Seq analysis files in .csv format represent the differential gene expression analysis results obtained by DESeq2 and basemeans.
Genome_build: hg19

source_name: human osteosarcoma cell line SJSA-1

growth_protocol: Cells were cultured in DMEM (home-made) supplemented with 10 % fetal bovine serum (FBS) and antibiotics

extract_protocol:
Total RNA was extracted from cells using TRIzol® (Invitrogen/Life Technologies).
For RNA-sequencing, the quality of total RNA was determined using the bioanalyzer 2100 from Agilent. All samples analyzed exhibited a RIN>8. Library preparation was conducted using the TruSeq RNA LT SamplePrep Kit (Illumina) starting from 1000 ng of total RNA. Barcodes used for sample preparation were used according to the indications given by the protocol. Accurate library quantitation of cDNA libraries was performed with the QuantiFluor™dsDNA System (Promega). The size range of final cDNA libraries was determined applying the DNA 1000 chip on the Bioanalyzer 2100 from Agilent (290-310 bp). cDNA libraries were amplified and sequenced via cBot and HiSeq 2000 from Illumina (SR, 1×50 bp, 6 Gb/sample ca. 30 million reads per sample). Sequence images were transformed with Illumina software BaseCaller to bcl files, which were demultiplexed to fastq files with CASAVA (version 1.8.2).

treatment_protocol: Cells were treated with 20µM Nutlin for 6 hours and PD0332991 10µM for 24h/30h and in combination

tag: cell line

value: human osteosarcoma cell line


title: Pool_1747_AD006_indexed


title: Pool_1732_AD010_indexed


title: 171213_CEMBA_mm_P56_P63_3C_MOp_CEMBA171206_3C_1_CEMBA171206_3C_3_A1_AD004_indexed


title: N+PD30_2


title: N+PD30_1


title: Pool_1729_AD010_indexed


title: Pool_1724_AD010_indexed


title: 71B_tumor vs reference

description:
MAL_251901510131_S01_CGH_105_Dec08_1_2.txt
MAL_251901510131_S01_CGH_105_Dec08_1_2.txt

value: normal


title: 171213_CEMBA_mm_P56_P63_3C_MOp_CEMBA171206_3C_1_CEMBA171206_3C_3_A1_AD001_indexed

value: Male


title: 71A_tumor vs reference

description:
raw data file: US22502676_251901510129_S01_CGH_105_Dec08_1_1.txt
MAL_251901510131_S01_CGH_105_Dec08_1_2.txt

value: WT


title: 171213_CEMBA_mm_P56_P63_3C_MOp_CEMBA171206_3C_1_CEMBA171206_3C_3_A12_AD006_indexed


title: N+PD24_3


title: 171213_CEMBA_mm_P56_P63_3C_MOp_CEMBA171206_3C_1_CEMBA171206_3C_3_A11_AD012_indexed


title: Pool_1718_AD006_indexed


title: 41A_tumor vs reference

description:
raw data file: US22502676_251901510377_S01_CGH_105_Dec08_1_1.txt
raw data file: US22502676_251901510377_S01_CGH_105_Dec08_1_1.txt


title: N+PD24_1


title: 11A_tumor vs reference

description:
raw data file: US22502676_251901512332_S01_Cropped_CGH_105_Dec08_1_1.txt
raw data file: US22502676_251901512332_S01_Cropped_CGH_105_Dec08_1_1.txt


title: 171213_CEMBA_mm_P56_P63_3C_MOp_CEMBA171206_3C_1_CEMBA171206_3C_3_A1_AD002_indexed


title: 171213_CEMBA_mm_P56_P63_3C_MOp_CEMBA171206_3C_1_CEMBA171206_3C_3_A11_AD010_indexed


title: Pool_1710_AD008_indexed


title: 171213_CEMBA_mm_P56_P63_3C_MOp_CEMBA171206_3C_1_CEMBA171206_3C_3_A11_AD008_indexed


title: 41B_tumor vs reference

description:
raw data file: US22502676_251901510377_S01_CGH_105_Dec08_1_2.txt
raw data file: US22502676_251901510377_S01_CGH_105_Dec08_1_2.txt


title: PD30_3


title: Pool_1699_AD008_indexed


title: 11B_tumor vs reference

description:
raw data file: US22502676_251901512332_S01_Cropped_CGH_105_Dec08_1_2.txt
raw data file: US22502676_251901512332_S01_Cropped_CGH_105_Dec08_1_1.txt


title: 12A_tumor vs reference

description:
raw data file: MAF_251901510153_S01_CGH_105_Dec08_1_2.txt
raw data file: MAF_251901510153_S01_CGH_105_Dec08_1_1.txt


title: Pool_1696_AD002_indexed


title: PD30_1


title: 171213_CEMBA_mm_P56_P63_3C_MOp_CEMBA171206_3C_1_CEMBA171206_3C_3_A11_AD007_indexed


title: 71C_tumor vs reference

description:
MAL_251901510131_S01_CGH_105_Dec08_1_1.txt
MAL_251901510131_S01_CGH_105_Dec08_1_2.txt


title: 171213_CEMBA_mm_P56_P63_3C_MOp_CEMBA171206_3C_1_CEMBA171206_3C_3_A11_AD006_indexed


title: PD30_2


title: Pool_1693_AD008_indexed


title: 171213_CEMBA_mm_P56_P63_3C_MOp_CEMBA171206_3C_1_CEMBA171206_3C_3_A11_AD004_indexed


title: Pool_1686_AD006_indexed


title: PD24_3


title: PD24_2


title: N+PD24_2


title: 171213_CEMBA_mm_P56_P63_3C_MOp_CEMBA171206_3C_1_CEMBA171206_3C_3_A11_AD002_indexed


title: 68B_tumor vs reference

description:
raw data file: US22502676_251901512356_S01_CGH_105_Dec08_1_2.txt
raw data file: US22502676_251901512356_S01_CGH_105_Dec08_1_2.txt


title: 68A_tumor vs reference

description:
raw data file: US22502676_251901512356_S01_CGH_105_Dec08_1_1.txt
raw data file: US22502676_251901512356_S01_CGH_105_Dec08_1_1.txt


title: Pool_1693_AD010_indexed


title: Nutlin_3


title: PD24_1


title: 171213_CEMBA_mm_P56_P63_3C_MOp_CEMBA171206_3C_1_CEMBA171206_3C_3_A10_AD012_indexed


title: 171213_CEMBA_mm_P56_P63_3C_MOp_CEMBA171206_3C_1_CEMBA171206_3C_3_A11_AD001_indexed


title: Pool_1681_AD010_indexed


title: 2A_tumor vs reference

description:
raw data file: US22502676_251901512359_S01_CGH_105_Dec08_1_2.txt
raw data file: US22502676_251901512359_S01_CGH_105_Dec08_1_2.txt


title: 171213_CEMBA_mm_P56_P63_3C_MOp_CEMBA171206_3C_1_CEMBA171206_3C_3_A10_AD010_indexed


title: 171213_CEMBA_mm_P56_P63_3C_MOp_CEMBA171206_3C_1_CEMBA171206_3C_3_A10_AD008_indexed


title: Nutlin_1


title: 2B_tumor vs reference

description:
raw data file: US22502676_251901512359_S01_CGH_105_Dec08_1_1.txt
raw data file: US22502676_251901512359_S01_CGH_105_Dec08_1_1.txt


title: 171213_CEMBA_mm_P56_P63_3C_MOp_CEMBA171206_3C_1_CEMBA171206_3C_3_A10_AD007_indexed


title: 171213_CEMBA_mm_P56_P63_3C_MOp_CEMBA171206_3C_1_CEMBA171206_3C_3_A10_AD004_indexed


title: 69A_tumor vs reference

description:
raw data file: US22502676_251901512355_S01_CGH_105_Dec08_1_2.txt
raw data file: US22502676_251901512355_S01_CGH_105_Dec08_1_1.txt


title: K562_Input_AR17

city: Chicago

first: Rohan

middle: Nishant

last: Shah

state: IL

address: CLSC 856, 920 E. 58th Street

institute: University of Chicago

description:
Sample 20
Input chromatin used for ICeChIP-seq in Samples 17-19.

data_processing:
To align reads, a reference genome was first created, consisting of the human genome (GRCh38/hg38) appended by the sequences of each of the nucleosome standard barcodes. Reads were then mapped to this reference genome using Bowtie2 using the sensitive preset and end-to-end alignment options.
Using SAMTools, any reads which were not paired, not mapped in a proper pair, or mapped with a map quality below 20 were then discarded to prevent low-quality mapping or sequencing reads from impacting downstream analyses. Reads were then flattened to create a single mapping from each matched pair of reads, and any mappings with lengths greater than 200bp were also discarded to ensure only mononucleosomes were being analyzed.
Bedgraphs of genome coverage were then generated using BEDTools, and IP and input genome coverage bedgraphs were merged using BEDTools7. The sum of reads across ladder members for each nucleosomal standard was computed for each sample and HMD bedgraphs were then generated from the merged bedgraphs using awk to apply the following formula: HMD (%) = 100% * (IPlocus/INlocus) / (IPbarcode/INbarcode). Error and 95% confidence intervals were computed with Poisson statistics and error propagation from the merged bedgraphs using awk to apply the following formula: 95CI Error (%) = 1.96 * HMD (%) * sqrt(1/IPlocus + 1/INlocus). Bigwig files were generated for visualization using the bedGraphToBigWig tool.
To compute average HMD across a series of intervals, a “double mapping” procedure was used. First, the HMD bedgraph was mapped onto 1bp windows made for each interval using BEDTools. Then, the mapped windows were mapped onto the original intervals using BEDTools. This procedure ensured that the degree of overlap of the interval with each value of the HMD bedgraph was accounted for in the mapping procedure.
Peak calling was conducted using Macs2 using the bdgpeakcall, with the input being the HMD bedgraphs computed for each sample. Peaks within 200bp of each other were then merged using BEDTools. The average HMD and average 95% CI Error of each called peak was computed. At this point, those peaks with greater average HMD than average 95% CI Error were selected as “high-confidence” peaks. All subsequent peak analyses were conducted with these “high-confidence” peaks.
Correction was conducted using the antibodies AB 8895, AB 7766, AB 12209, and AB 8580. Correction was done using our previously-described method against H3K4me1, H3K4me2, H3K4me3, and H4K20me3 off-target binding. Briefly, measured HMD by each antibody can be described by a vector M, and the measured specificities by each antibody can be described by a square matrix S. Then, we can state, if other off-specific binding is negligible, that the correct HMDs for H3K4me1, H3K4me2, H3K4me3, and H4K20me3 can be expressed by the vector C such that M=CS. As such, the vector C can be computed as CSSi = C = MSi. The elements of Si were then used to compute the HMD and Error of the corrected profiles using awk to linearly combine the AB 8895, AB 7766, AB 12209, and AB 8580 profiles.
Genome_build: hg38_bar_Ca1-17 (GRCh38/hg38 appended by synthetic DNA barcodes used with semisynthetic nucleosome standards)
Supplementary_files_format_and_content: Bedgraph files all have the form <chromosome> <start> <end> <value>, where the fourth field represents the value of the measurement. Bigwig files are bedgraph files in binary format, appropriate for viewing on a genome browser. Bed files all have the form <chromosome> <start> <end> and represent a list of intervals. Fasta file (with extension .fa) represents the reference genome used in this study, with the human genome (GRCh38/hg38) appended by the DNA barcode sequences used for the semisynthetic nucleosome standards.

source_name: Cell Culture

growth_protocol: K562 cells were grown in DMEM (Invitrogen) containing 10% (v/v) HyClone FBS Characterized U.S. and 1x Penicillin/Streptomycin. Cells were grown in supplemented medium in suspension in 25 cm-squared vented flasks. The cells were seeded at 100,000 to 200,000 cells per mL and were grown until they reached a density of 1 million cells per mL, at which point they were passaged 1:10. Cells used in ChIP experiments were harvested at a concentration of approximately 1 million cells per mL.

extract_protocol:
K562 cell pellets were washed twice with 5 mL of PBS, then washed twice with 5 mL of Buffer N (15 mM Tris-HCl pH 7.5, 15 mM NaCl, 60 mM KCl, 8.5% w/v Sucrose, 5 mM MgCl2, 1 mM CaCl2, 1 mM DTT, 200 uM PMSF, 50 ug/mL BSA, 1x Roche Protease Inhibitor Cocktail, Filter Sterilized), with each wash consisting of complete resuspension of the pellet, centrifugation at 500 g for 5 minutes at 4°C, and removal of supernatant. The washed pellet was then resuspended in at least 2 packed cell volumes (PCV) of Buffer N and mixed with 1 volume of Lysis Buffer (Buffer N supplemented with 0.6% NP-40 Substitute) and incubated on ice for 10 minutes to lyse cells. The crude nuclei were spun down at 500 g for 5 minutes at 4°C before being resuspended in at least 6 packed nuclear volumes (PNV) of Buffer N and applied to the top of 7.5 mL of Sucrose Cushion N (15 mM Tris-HCl pH 7.5, 15 mM NaCl, 60 mM KCl, 30% w/v Sucrose, 5 mM MgCl2, 1 mM CaCl2, 1 mM DTT, 200 uM PMSF, 50 ug/mL BSA, 1x Roche Protease Inhibitor Cocktail, Filter Sterilized) in a 15 mL centrifuge tube, then spun down at 500 g for 12 minutes at 4°C in a swinging-bucket rotor. The supernatant was discarded after centrifugation, and the pellet was resuspended in approximately 2 PNV of Buffer N. The nuclei were then quantified by diluting 2 uL of nuclei suspension into 48 uL of 2 M NaCl, water-bath sonicating to solubilize DNA, and spectroscopically measuring DNA concentration by Nanodrop measurement, where 1 A280nm¬ = 50 ng/uL chromatin in the measurement sample. After accounting for the 25-fold dilution of the measurement sample, the concentration of the nuclei was adjusted down to 1ug/uL of chromatin. Nuclei were aliquoted into 100 uL aliquots, flash frozen, and stored at -80°C until use. When nuclei aliquots were used, they were first thawed and spiked approximately 1 uL of each barcoded nucleosome standard per 50 ug of chromatin to be used. The nuclei suspension with spiked calibrants was then mixed with a pipette and transferred to a new tube, then warmed to 37°C for 2 minutes. After warming, 1 unit of micrococcal nuclease (MNase) per 4.375 ug of chromatin was added to the nuclei suspension and incubated at 37°C while shaking at 900 rpm for 12 minutes. After incubation, the digestion was stopped by adding 1/9 volume of 10x MNase Stop Buffer (100 mM EDTA, 100 mM EGTA, Filter Sterilized) while slowly vortexing, and nuclei were lysed by adding 5 M NaCl to a final concentration of 600 mM NaCl while slowly vortexing. 66 mg of HAP resin (BioRad, CHTTM Ceramic Hydroxyapatite, Type I, 20 um) per 100 ug of chromatin digested was rehydrated with 200 uL of HAP Buffer 1 (5 mM Sodium Phosphate pH 7.2, 600 mM NaCl, 1 mM EDTA, 200 uM PMSF, Filter Sterilized) per 100 ug of chromatin digested. The lysed nuclei were then spun down at 18,000 g for 1 minute to pellet insoluble nuclear debris, and the soluble fraction was added to the rehydrated HAP resin and incubated for 10 minutes on a rotator at 4°C. After incubation, the HAP resin slurry was added to a centrifugal filter unit (Millipore Ultrafree MC–HV Centrifugal Filter 0.45 um) and spun at 1000 g for 30 seconds at 4°C. The HAP resin left on the filter unit was then washed 4 times with 200 uL HAP Buffer 1 and 4 times with 200 uL HAP Buffer 2 (5 mM Sodium Phosphate pH 7.2, 100 mM NaCl, 1 mM EDTA, 200 uM PMSF, Filter Sterilized) by spinning at 1000 g for 30 seconds at 4°C. Into a clean tube, the HAP resin was eluted with three elutions of 100 uL HAP Elution Buffer (500 mM Sodium Phosphate pH 7.2, 100 mM NaCl, 1 mM EDTA, 200 uM PMSF, Filter Sterilized). The DNA content of the elution was then quantified spectroscopically with a Nanodrop, and the chromatin concentration was adjusted to 20 ng/uL with ChIP Buffer 1 (25 mM Tris pH 7.5, 5 mM MgCl2, 100 mM KCl, 10% v/v glycerol, 0.1% v/v NP-40 Substitute, Filter Sterilized) with 100 ug/mL of BSA. With the exception of the Koide Lab 304M3B antibody, every ICeChIP was conducted with 12.5 uL of Protein A Dynabeads (Invitrogen) and 3 ug of antibody. The Protein A Dynabeads for each ICeChIP were washed with 50 uL of ChIP Buffer 1 by use of a magnetic rack, then resuspended in 50 uL of ChIP Buffer 1. In a separate set of tubes, 3 ug of each antibody was diluted to 100 uL with ChIP Buffer 1. The antibody suspension and the Protein A Dynabeads suspension were combined and incubated on a rotator at 4°C for at least 1 hour, then washed with 200 uL of ChIP Buffer 1 by use of a magnetic rack and resuspended in 50 uL of ChIP Buffer 1. After antibody washing, 150 uL of the diluted chromatin elution was added to each antibody-bead conjugate and incubated for 15 minutes on a rotator at 4°C. After this incubation, the beads were washed twice with ChIP Buffer 2 (25 mM Tris pH 7.5, 5 mM MgCl2, 300 mM KCl, 10% v/v glycerol, 0.1% v/v NP-40 Substitute, Filter Sterilized) and once with ChIP Buffer 3 (10 mM Tris pH 7.5, 250 mM LiCl, 1 mM EDTA, 0.5% Sodium Deoxycholate, 0.5% v/v NP-40 Substitute, Filter Sterilized), with a wash consisting of removal of the existing supernatant by use of a magnetic rack, resuspension into 150 uL of buffer, transfer to a new siliconized tube, and incubation on the rotator for 10 minutes at 4°C. After these washes, the supernatant was removed, and the beads were resuspended in ChIP Buffer 1, transferred to a new siliconized tube, then rinsed once with 200 uL of TE before being resuspended in 50 uL of ChIP Elution Buffer and incubated at 55°C for 5 minutes. After incubation, the supernatant was transferred to a new set of siliconized tubes, and the beads were discarded. To each supernatant was then added 2 uL of 5 M NaCl, 1 uL of 500 mM EDTA, and 1 uL of 10 mg/mL Proteinase K. 15 uL of Input DNA was also diluted to 50 uL with 35 uL of ChIP Elution Buffer (50 mM Tris pH 7.5, 1 mM EDTA, 1% w/v SDS, Filter Sterilized) and was supplemented with 2 uL of 5 M NaCl, 1 uL of 500 mM EDTA, and 1 uL of 10 mg/mL Proteinase K. The IP elutions and diluted input were then incubated at 55°C for 2 hours for a Proteinase K digestion. After digestion, the DNA was purified by adding 1.5 volumes of Serapure HD (1:50 dilution of Sera-Mag SpeedBeads [Fisher], 20% PEG-8000, 2.5 M NaCl, 10 mM Tris pH 7.5, 1 mM EDTA, 0.05% Tween-20, Filter Sterilized prior to addition of SpeedBeads), incubating at room temperature for 15 minutes, then collecting the beads on a magnetic rack, washing twice with 150 uL of 70% ethanol, and eluting into 50 uL of water, which was then recovered and stored at -20°C.
DNA purified with Serapure was quantified using Quant-iT TM PicoGreen (Thermo Fisher) per manufacturer instructions. Up to 10 ng of each DNA sample for which libraries were to be prepared were then carried forward. Libraries were then generated from the NEBNext Ultra II DNA Library Prep kit (New England Biolabs) per manufacturer instructions. The DNA content of the library was then quantified and pooled for Illumina sequencing.

tag: amplification cycles


title: 69B_tumor vs reference

description:
raw data file: US22502676_251901512355_S01_CGH_105_Dec08_1_1.txt
raw data file: US22502676_251901512355_S01_CGH_105_Dec08_1_1.txt


title: DMSO_2

tag: cell type


title: K562_309M3B_H3K9me3_ICeChIP

description:
Sample 19
ICeChIP-seq conducted on K562 cells against H3K9me3 using the antibody  Koide Lab 309M3B recombinant anti-H3K9me3.


title: K562_EPG-A-4033-050_H3K4me3_ICeChIP

description:
Sample 18
ICeChIP-seq conducted on K562 cells against H3K4me3 using the antibody  EpiGentek A-4033-050, Lot 606361, anti-H3K4me3.


title: 171213_CEMBA_mm_P56_P63_3C_MOp_CEMBA171206_3C_1_CEMBA171206_3C_3_A10_AD002_indexed

tag: Sex


title: DMSO_1


title: Rectum tissue isolated from African Green Monkey at Mock in Animal ID AGM9

city: Minneapolis

first: Michael

last: Gale

state: Minnesota

address: 689 23rd Ave SE

department: Microbiology and Immunology

institute: University of Minnesota

description: RNA isolated from Rectum from African Green Monkey

data_processing:
Quality control of the primary sequencing data was performed using FastQC (version 0.11.3) and included adapter trimming using cutadapt (version 1.8.3).
Reads were aligned using STAR (version 2.4.2)
Ensembl build Mmul_1 using illumina's igenomes resource
We mapped reads to the genome Macaca mulatta genome (build Mmul_1) with STAR. After mapping, we assigned aligned read counts from BAM files to exons and genes using the python package HT-Seq (version 0.6.1). HT-Seq provided the most accurate way of aligning read counts to overlapping exons. Reads that mapped to multiple positions were removed.
Genome_build: Macaca mulatta Ensembl (build Mmul_1) from illumina's igenomes resource: https://support.illumina.com/sequencing/sequencing_software/igenome.html
Genome_build: Chlorocebus_sabeus 1.0 from https://www.ncbi.nlm.nih.gov/assembly/GCA_000409795.1/
Supplementary_files_format_and_content: A raw gene count matrix of files generated with ht-seq is attached

organism: Chlorocebus aethiops

growth_protocol: please refer to https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4097788/

extract_protocol:
mRNA was isolated using the QIAGEN miRNeasy Kit.
Whole transcriptome libraries were constructed using the TruSeq Stranded Total RNA with Ribo-Zero Gold (Illumina, San Diego, CA) according to the manufacturer’s guide.
Libraries were quality controlled and quantitated using the BioAnalzyer 2100 system and qPCR (Kapa Biosystems, Woburn, MA).

value: Rectum

tag: timepoint

value: Mock


title: Dropseq-H6

city: Boston

first: Chen

last: Weng

state: OH

address: 415 Main St, Cambridge

department: Genetics

institute: Broad Institute and Whitehead Institute

description: Droplet based single cell RNAseq(Dropseq)

data_processing:
We performed raw reads processing following the instructions described in the original Drop-Seq publication (Macosko et al., 2015). The sequenced Drop-Seq libraries yield 50-base paired-end reads (PE50). However, since only the first 20bp of read 1 is informative (base 1-12 cell barcode, base 13-20 UMI), we trimmed base 21-50 of read 1 before further analysis. We first remove all data with the quality score of read 1 (base 1-20) lower than 10. Read 2 was trimmed at 3’ end to remove ployA tails of at least 6 bases, and trimmed at 5’ if template switching oligo (TSO) adapter sequence appears. Clean reads were then aligned to hg18 or mm9 using STAR with default settings. We only keep uniquely mapped reads on gene exons. We next filtered out PCR duplicates with the same coordinates, cell barcode, and UMI. We then grouped the reads by cell barcode, and generated the digital UMI-count matrix after counting transcripts for every gene with different cell barcode.
Genome_build: hg18
Supplementary_files_format_and_content: DGE(Digital gene expression) table was generated.   Row stands for genes; Column stands for single cells.  Number is the transcripts number(UMI) of any given gene in any given cell

molecule: polyA RNA

source_name: Non-diabetic donor 6

growth_protocol: To dissociate islets into single cells, cells were washed once in HBSS (Sigma-aldrich, #6648) and incubated in Accutase (Innovative Cell Technologies, #AT104) at 37°C for 20-25 min. The islets were broken up gently with a 5 ml pipette every 5 min. When >95% of the islets were digested into single cells, PIM(S) medium were added to neutralize the Accutase. Single cells were washed again with HBSS and resuspended in HBSS at the concentration of 2 x 105/mL for Drop-Seq analysis.

extract_protocol:
We performed Drop-Seq following the protocol as previously described (Macosko et al., 2015). Briefly, three pump-controlled syringes with cell suspension (200,000 cells/mL), barcoded beads in lysis buffer (120,000 beads/mL), and droplet generation oil were connected to a microfluidic device under microscope supervision. During droplets generation, we set the cell and bead flow speed at 4,000uL/hr, and the oil speed at 15,000uL/hr. The droplets are collected into 50mL falcon tubes (usually less than 5mL). Under this setting, most droplets have at most one beads or one cell.
Following droplet breakage,we performed 1st strand cDNA synthesis on beads followed by SMART-PCR (Macosko et al., 2015). Finally, the resulting full-length cDNA library are prepared for sequencing exactly following the published Drop-Seq protocol (Macosko et al., 2015).

value: Cadaveric pancreatic islet

tag: donor age

tag: gender

value: M

tag: bmi


title: Dropseq-T2D3

source_name: Diabetic donor 3


title: K562_DIA-C15410003_H3K4me3_ICeChIP

description:
Sample 17
ICeChIP-seq conducted on K562 cells against H3K4me3 using the antibody  Diagenode C15410003, Lot A1052D, anti-H3K4me3.


title: K562_Input_AR16

description:
Sample 16
Input chromatin used for ICeChIP-seq conducted in Samples 1-15.

source_name: Cell culture


title: Rectum tissue isolated from African Green Monkey at Mock in Animal ID AGM11

treatment_protocol: please refer to https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4097788/


title: Rectum tissue isolated from African Green Monkey at Mock in Animal ID AGM29


title: Nutlin_2


title: Rectum tissue isolated from African Green Monkey at Mock in Animal ID AGM10


title: DMSO_3


title: K562_EMD-07-473_H3K4me3_ICeChIP

description:
Sample 14
ICeChIP-seq conducted on K562 cells against H3K4me3 using the antibody  EMD Millipore 07-473, Lot 2839113, anti-H3K4me3.


title: K562_TF-PA5-40086_H3K4me3_ICeChIP

description:
Sample 15
ICeChIP-seq conducted on K562 cells against H3K4me3 using the antibody  Thermo Fisher PA5-40086, Lot SC2349865, anti-H3K4me3.


title: K562_ABC-46698_H3K4me3_ICeChIP

description:
Sample 12
ICeChIP-seq conducted on K562 cells against H3K4me3 using the antibody  Abclonal A2357, Vial 46698, anti-H3K4me3.


title: Rectum tissue isolated from African Green Monkey at Day 9 in Animal ID AGM28

value: Day 9


title: K562_EMD-05-745R_H3K4me3_ICeChIP

description:
Sample 13
ICeChIP-seq conducted on K562 cells against H3K4me3 using the antibody  EMD Millipore 05-745R, Lot 2813867, anti-H3K4me3.


title: Dropseq-H5

source_name: Non-diabetic donor 5


title: Dropseq-H3

source_name: Non-diabetic donor 3

value: F


title: Dropseq-T2D2

source_name: Diabetic donor 2


title: 171213_CEMBA_mm_P56_P63_3C_MOp_CEMBA171206_3C_1_CEMBA171206_3C_3_A10_AD001_indexed

source_name: Mouse primary motor cortex

tag: age

tag: genome build

tag: library strategy


title: Rectum tissue isolated from African Green Monkey at Day 9 in Animal ID AGM127


title: K562_AB-12209_H3K4me3_ICeChIP

description:
Sample 10
ICeChIP-seq conducted on K562 cells against H3K4me3 using the antibody  Abcam ab12209, Lot GR275790-1, anti-H3K4me3.

tag: chip antibody


title: K562_AB-8580_H3K4me3_ICeChIP

description:
Sample 11
ICeChIP-seq conducted on K562 cells against H3K4me3 using the antibody  Abcam ab8580, Lot GR190229-1, anti-H3K4me3.


title: Dropseq-T2D1

source_name: Diabetic donor 1


title: Rectum tissue isolated from African Green Monkey at Day 9 in Animal ID AGM133


title: Combination, 2w_1

city: SAN DIEGO

first: gaowei

last: wang

state: CALIFORNIA

address: 3393 Lebon DR, APT 202

institute: University of California San Diego

data_processing:
Raw data quality control using QCfastq
Sequenced reads were mapped to mm9 whole genome using star
Fragments Per Kilobase Million(FPKM) were calculated using cuffdiff
Genome_build: mm9
Supplementary_files_format_and_content: csv files include FPKM values for each sample

source_name: CD45 positive cells in Liver

extract_protocol:
CD45 positive immune cells were isolated by isolated magnetic beads separation (MACS) using PE-conjugated CD45 mAb and PE microbeads. Total RNAs was extracted and 1 ug of total RNA was used for the construction of sequencing libraries.. cDNA library was prepared using Illumina TruSeq Stranded mRNA Library Prep Kit (RS-122-2101, Illumina).
RNA libraries were prepared for sequencing using standard Illumina protocols

treatment_protocol: All the mice were hydrodynamic transfected of human c-Myc and N-ras genes at day 0. Control group: PBS was administrated i.p. at day 14,16,18,20 and 22; IgG(200ug ) was administrated at day 17, 19, 21. PolyIC group: polyIC (4mg/kg in PBS) was administrated i.p. at day 14,16,18,20 and 22; IgG(200ug) was administrated at day 17, 19, 21. anti-PD-L1 group: PBS was administrated i.p. at day 14,16,18,20 and 22; PD-L1 Ab (200ug) was administrated at day 17, 19, 21. Combination group: polyIC (4mg/kg in PBS) was administrated i.p. at day 14,16,18,20 and 22; PD-L1 Ab (200ug) was administrated at day 17, 19, 21. All the mice were euthanized at day 24 and the livers were isolated.

tag: strain

value: Liver

tag: time point

value: post hydrodynamic transfection and treatment day 22

tag: transfection

value: Transfection of c-Myc/N-Ras plus polyIC and PD-L1 Ab treatment

value: CD45 positive immune cell


title: Combination, 2w_0

tag: tissue


title: Combination, 2w_2


title: only PD-L1, 2w_2

value: Transfection of c-Myc/N-Ras plus PD-L1 Ab treatment


title: Dropseq-H1

source_name: Non-diabetic donor 1


title: Rectum tissue isolated from African Green Monkey at Day 6 in Animal ID AGM19

value: Day 6


title: 20B_tumor vs reference

description:
raw data file: US22502676_252336313663_S01_CGH_105_Dec08_1_3.txt
raw data file: US22502676_252336313663_S01_CGH_105_Dec08_1_2.txt

source_name: tumor

tag: histology

value: adenocarcinoma

source_name: reference


title: Dropseq-H4

source_name: Non-diabetic donor 4


title: K562_EMD-05-1338_H3K4me2_ICeChIP

description:
Sample 8
ICeChIP-seq conducted on K562 cells against H3K4me2 using the antibody  EMD Millipore 05-1338, Lot 2757107, anti-H3K4me2.

tag: cell line


title: Rectum tissue isolated from African Green Monkey at Day 6 in Animal ID AGM23


title: K562_TF-710796_H3K4me2_ICeChIP

description:
Sample 9
ICeChIP-seq conducted on K562 cells against H3K4me2 using the antibody  Thermo Fisher 710796, Lot QL230606, anti-H3K4me2.

tag: amplification cycles


title: only PD-L1, 2w_1

tag: cell type


title: Rectum tissue isolated from African Green Monkey at Day 6 in Animal ID AGM18


title: 20A_tumor vs reference

value: large cell carcinoma


title: K562_CST-9725_H3K4me2_ICeChIP

description:
Sample 7
ICeChIP-seq conducted on K562 cells against H3K4me2 using the antibody  Cell Signaling Technologies 9725, Lot 9, anti-H3K4me2.


title: Dropseq-H2

source_name: Non-diabetic donor 2


title: Rectum tissue isolated from African Green Monkey at Day 5 in Animal ID AGM136

value: Day 5


title: Rectum tissue isolated from African Green Monkey at Day 55 in Animal ID AGM132

value: Day 55


title: Rectum tissue isolated from African Green Monkey at Day 53 in Animal ID AGM131

value: Day 53


title: only poly-IC, 2w_2

value: Transfection of c-Myc/N-Ras plus polyIC treatment


title: only poly-IC, 2w_1


title: Rectum tissue isolated from African Green Monkey at Day 4 in Animal ID AGM135

value: Rectum

value: Day 4


title: only poly-IC, 2w_0


title: K562_AB-7766_H3K4me2_ICeChIP

description:
Sample 6
ICeChIP-seq conducted on K562 cells against H3K4me2 using the antibody  Abcam ab7766, Lot GR289627-1, anti-H3K4me2.


title: K562_TF-710795_H3K4me1_ICeChIP

description:
Sample 5
ICeChIP-seq conducted on K562 cells against H3K4me1 using the antibody  Thermo Fisher 710795, Lot QL230605, anti-H3K4me1.


title: Control_2

value: Transfection of c-Myc/N-Ras


title: Rectum tissue isolated from African Green Monkey at Day 4 in Animal ID AGM128


title: Control_1


title: Rectum tissue isolated from African Green Monkey at Day 4 in Animal ID AGM125


title: Rectum tissue isolated from African Green Monkey at Day 48 in Animal ID AGM130

value: Day 48


title: K562_EPG-A-4031-050_H3K4me1_ICeChIP

description:
Sample 4
ICeChIP-seq conducted on K562 cells against H3K4me1 using the antibody  EpiGentek A-4031-050, Lot 606359, anti-H3K4me1.


title: Rectum tissue isolated from African Green Monkey at Day 46 in Animal ID AGM129

description: RNA isolated from Rectum from African Green Monkey

value: Day 46


title: Control_0

value: Liver


title: K562_CST-5326BF_H3K4me1_ICeChIP

description:
Sample 3
ICeChIP-seq conducted on K562 cells against H3K4me1 using the antibody  Cell Signaling Technologies 5326 BF, Lot 2, anti-H3K4me1.


title: K562_AB-8895_H3K4me1_ICeChIP

description:
Sample 1
ICeChIP-seq conducted on K562 cells against H3K4me1 using the antibody  Abcam ab8895, Lot GR305231-1, anti-H3K4me1.


title: FS116_TREM2+/+

city: Los Angeles

first: Giovanni

last: Coppola

address: 1524 Gonda, 695 Charles Young Drive South

department: Psychiatry and Neurology

institute: UCLA

data_processing:
Aligned to mm10 using STAR spliced read aligner
Count read using HT-seq and using mm10 RefSeq as reference
Normalization (TMM), quantitation (FPKM) and differential expression analysis using edgeR and limma voom.
Genome_build: mm10
Supplementary_files_format_and_content: FPKM values

extract_protocol:
Total RNA was extracted using RNeazy kit (Qiagen)accordance with the prescribed protocol provided with the kit.
Libraries were prepared using the Lexogen QuantSeq 3' mRNA-Seq Library Prep Kit and sequenced on an Illumina HiSeq4000 sequencer.

tag: genotype

value: M

value: 8- to 9-months

value: cortical brain tissue


title: K562_AM-39635_H3K4me1_ICeChIP

description:
Sample 2
ICeChIP-seq conducted on K562 cells against H3K4me1 using the antibody  Active Motif 39635, Lot 30615011, anti-H3K4me1.


title: FS115_TREM2+/+

value: F


title: FS114_TREM2+/+


title: R24Sil_RGn14_RM_SIV_SC_D14_1_NCMono_RLT_mRNA_Seq_S15

description: Sorted Non-classical Monocytes  Infected with SIV at Day 14

data_processing:
Constructed libraries were sequenced on an NextSeq 500 Illumina platform, producing 2x75nt stranded paired end reads (52 Gb). Image analysis, base calling, and error estimation were performed using Illumina Analysis Pipeline (version 2.8).
Quality control of the primary sequencing data was performed using FastQC (version 0.11.3) and included adapter trimming using cutadapt (version 1.8.3).
Reads were aligned using STAR (version 2.4.2)
Ensembl build Mmul_1 using illumina's igenomes resource
We mapped reads to the genome Macaca mulatta genome (build Mmul_1) with STAR. After mapping, we assigned aligned read counts from BAM files to exons and genes using the python package HT-Seq (version 0.6.1).  HT-Seq provided the most accurate way of aligning read counts to overlapping exons. Reads that mapped to multiple positions were removed.
Genome_build: Macaca mulatta Ensembl (build Mmul_1) from illumina's igenomes resource: https://support.illumina.com/sequencing/sequencing_software/igenome.html
Genome_build: Chlorocebus_sabeus 1.0 from https://www.ncbi.nlm.nih.gov/assembly/GCA_000409795.1/
Supplementary_files_format_and_content: Two raw count matrix of files generated with ht-seq is attached

organism: Macaca mulatta

extract_protocol:
mRNA was isolated using the QIAGEN miRNeasy Kit.
Whole transcriptome libraries were constructed using the TruSeq Stranded Total RNA with Ribo-Zero Gold (Illumina, San Diego, CA) according to the manufacturer’s guide.  Libraries were quality controlled and quantitated using the BioAnalzyer 2100 system and qPCR (Kapa Biosystems, Woburn, MA).

tag: infected

value: Infected with SIV

tag: animal id

tag: timepoint

value: Day 14

tag: library batch

value: Non-classical Monocytes


title: Rectum tissue isolated from African Green Monkey at Day 3 in Animal ID AGM15

value: Day 3


title: R24Sil_RGn14_RM_SIV_SC_D7_2_NCMono_RLT_mRNA_Seq_S14

description: Sorted Non-classical Monocytes  Infected with SIV at Day 7

value: Day 7


title: R24Sil_RGn14_RM_SIV_SC_D28_1_NCMono_RLT_mRNA_Seq_S16

description: Sorted Non-classical Monocytes  Infected with SIV at Day 28

value: Day 28


title: Rectum tissue isolated from African Green Monkey at Day 3 in Animal ID AGM13


title: R24Sil_RGn14_RM_Uninf_SC_Dminus29_1_NCMono_RLT_mRNA_Seq_S13

description: Sorted Non-classical Monocytes  Uninfected at Day minus29

value: Uninfected

value: Day minus29


title: only PD-L1, 2w_0

value: post hydrodynamic transfection and treatment day 22

value: Transfection of c-Myc/N-Ras plus PD-L1 Ab treatment

value: CD45 positive immune cell


title: R24Sil_RGn14_RM_SIV_SC_D28_1_Mono_RLT_mRNA_Seq_S12

description: Sorted Monocytes  Infected with SIV at Day 28

source_name: Sorted Monocytes  Infected with SIV

value: Monocytes


title: R24Sil_RGn14_RM_SIV_SC_D14_1_Mono_RLT_mRNA_Seq_S11

description: Sorted Monocytes  Infected with SIV at Day 14


title: FS112_P301S+ TREM2-/-

description: P301S+ TREM2-/-

tag: gender

tag: age


title: Rectum tissue isolated from African Green Monkey at Day 3 in Animal ID AGM122

value: Rectum


title: R24Sil_RGn14_RM_SIV_SC_D7_1_Mono_RLT_mRNA_Seq_S10

description: Sorted Monocytes  Infected with SIV at Day 7


title: Rectum tissue isolated from African Green Monkey at Day 2 in Animal ID AGM126

value: Day 2


title: FS111_P301S+ TREM2-/-

description: P301S+ TREM2-/-

value: M

value: 8- to 9-months


title: R24Sil_RGn14_RM_Uninf_SC_Dminus29_2_Mono_RLT_mRNA_Seq_S9

description: Sorted Monocytes  Uninfected at Day minus29

source_name: Sorted Monocytes  Uninfected


title: Rectum tissue isolated from African Green Monkey at Day 2 in Animal ID AGM124


title: Rectum tissue isolated from African Green Monkey at Day 2 in Animal ID AGM123


title: R24Sil_ROl14_RM_SIV_SC_D28_2_NCMono_RLT_mRNA_Seq_S8

source_name: Sorted Non-classical Monocytes  Infected with SIV


title: FS110_P301S+ TREM2-/-

description: P301S+ TREM2-/-

value: F

value: cortical brain tissue


title: Rectum tissue isolated from African Green Monkey at Day 1 in Animal ID AGM25

value: Day 1


title: FS109_P301S+ TREM2-/-

description: P301S+ TREM2-/-


title: FS108_P301S+ TREM2+/-

description: P301S+ TREM2+/-


title: R24Sil_ROl14_RM_SIV_SC_D14_1_NCMono_RLT_mRNA_Seq_S7

value: Infected with SIV

value: Day 14

value: Non-classical Monocytes


title: Rectum tissue isolated from African Green Monkey at Day 1 in Animal ID AGM12


title: Rectum tissue isolated from African Green Monkey at Day 12 in Animal ID AGM24

value: Day 12


title: R24Sil_ROl14_RM_Uninf_SC_Dminus21_1_NCMono_RLT_mRNA_Seq_S5

description: Sorted Non-classical Monocytes  Uninfected at Day minus21

value: Uninfected

value: Day minus21


title: FS106_P301S+ TREM2+/-

description: P301S+ TREM2+/-


title: FS113_TREM2+/+


title: Rectum tissue isolated from African Green Monkey at Day 12 in Animal ID AGM17


title: R24Sil_ROl14_RM_SIV_SC_D7_1_Mono_RLT_mRNA_Seq_S2

value: Day 7


title: R24Sil_ROl14_RM_SIV_SC_D14_1_Mono_RLT_mRNA_Seq_S3


title: Rectum tissue isolated from African Green Monkey at Day 12 in Animal ID AGM21


title: Rectum tissue isolated from African Green Monkey at Day 1 in Animal ID AGM14


title: FS105_P301S+ TREM2+/-

description: P301S+ TREM2+/-


title: Rectum tissue isolated from African Green Monkey at Day 12 in Animal ID AGM134


title: FS103_P301S+ TREM2+/+

description: P301S+ TREM2+/+


title: FS104_P301S+ TREM2+/+

description: P301S+ TREM2+/+


title: R24Sil_ROl14_RM_Uninf_SC_Dminus21_1_Mono_RLT_mRNA_Seq_S1

description: Sorted Monocytes  Uninfected at Day minus21


title: FS102_P301S+ TREM2+/+

description: P301S+ TREM2+/+


title: Rectum tissue isolated from African Green Monkey at Baseline in Animal ID AGM15

value: Baseline


title: R24Sil_RMe14_RM_SIV_SC_D7_1_NCMono_RLT_mRNA_Seq_S14


title: FS101_P301S+ TREM2+/+

description: P301S+ TREM2+/+


title: Rectum tissue isolated from African Green Monkey at Baseline in Animal ID AGM25


title: R24Sil_RMe14_RM_SIV_SC_D7_1_Mono_RLT_mRNA_Seq_S10


title: R24Sil_RMe14_RM_Uninf_SC_Dminus22_2_NCMono_RLT_mRNA_Seq_S13

description: Sorted Non-classical Monocytes  Uninfected at Day minus22

value: Day minus22


title: Rectum tissue isolated from African Green Monkey at Baseline in Animal ID AGM14


title: FS107_P301S+ TREM2+/-

description: P301S+ TREM2+/-


title: Rectum tissue isolated from African Green Monkey at Baseline in Animal ID AGM13


title: R24Sil_ROl14_RM_SIV_SC_D28_1_Mono_RLT_mRNA_Seq_S4

value: Day 28

value: Monocytes


title: R24Sil_ROl14_RM_SIV_SC_D7_1_NCMono_RLT_mRNA_Seq_S6


title: Rectum tissue isolated from African Green Monkey at Baseline in Animal ID AGM12

value: Rectum


title: R24Sil_RMe14_RM_SIV_SC_D14_combined_NCMono_RLT_mRNA_Seq_S15

value: combined


title: R24Sil_RMe14_RM_Uninf_SC_Dminus22_1_Mono_RLT_mRNA_Seq_S9

description: Sorted Monocytes  Uninfected at Day minus22


title: R24Sil_RMe14_RM_SIV_SC_D28_combined_NCMono_RLT_mRNA_Seq_S16


title: Rectum tissue isolated from African Green Monkey at Baseline in Animal ID AGM123


title: Rectum tissue isolated from African Green Monkey at Baseline in Animal ID AGM124


title: R24Sil_RMe14_RM_SIV_SC_D14_1_Mono_RLT_mRNA_Seq_S11


title: Rectum tissue isolated from African Green Monkey at Baseline in Animal ID AGM122


title: R24Sil_RCt14_RM_SIV_SC_D7_1_NCMono_RLT_mRNA_Seq_S6


title: R24Sil_RMe14_RM_SIV_SC_D28_2_Mono_RLT_mRNA_Seq_S12


title: R24Sil_RCt14_RM_SIV_SC_D7_1_Mono_RLT_mRNA_Seq_S2


title: Rectum tissue isolated from African Green Monkey at Baseline in Animal ID AGM126


title: R24Sil_RCt14_RM_Uninf_SC_Dminus28_1_NCMono_RLT_mRNA_Seq_S5

description: Sorted Non-classical Monocytes  Uninfected at Day minus28

value: Day minus28


title: R24Sil_RCt14_RM_Uninf_SC_Dminus28_1_Mono_RNAzol_mRNA_Seq_S1

description: Sorted Monocytes  Uninfected at Day minus28


title: R24Sil_RCt14_RM_SIV_SC_D28_1_NCMono_RLT_mRNA_Seq_S8


title: N18

city: Shanghai

first: Xiao-yan

last: dong

state: Shanghai

address: NO 24, Alley 1400, West Beijing Road

department: Department of respiration

institute: Shanghai Children's Hospital

hyb_protocol: The total 25 μl mixture from Hy3TM-labeled samples with 25 μl hybridization buffer were denatured at 95°C for 2 min, incubated in ice for 2 min, and then hybridized to the microarray at 56°C for 16–20 h with a 12-Bay Hybridization System (Nimblegen, Madison, WI, USA).

scan_protocol: After hybridization, the slides were prepared, washed with a wash buffer kit (Exiqon, Vedbaek, Denmark) and dried by centrifugation at 400 rpm for 5 min. The slides were then scanned by the Axon GenePix 4000B microarray scanner (Axon Instruments, Foster City, CA, USA).

data_processing: After low intensity miRNAs filtering, raw signal intensities are normalized in Median method. (miRNAs that intensities>=30 in all samples are chosen for calculating normalization factor)

source_name: control_peripheral blood

label_protocol: After RNA was isolated from the samples, the miRCURY™ Hy3™/Hy5™ Power labeling kit (Exiqon, Vedbaek, Denmark) was used according to the manufacturer’s guidelines for miRNA labeling. Briefly, 1g of each sample was 3'-end-labeled with Hy3TM fluorescent dye using T4 RNA ligase by the following procedure: RNA in 2.0 μl of water was combined with 1.0 μl of CIP buffer and CIP (Exiqon, Vedbaek, Denmark). The mixture was incubated for 30 min at 37°C and was terminated at 95°C for 5 min. Then, 3.0 μl of labeling buffer, 1.5 μl of fluorescent label (Hy3TM), 2.0 μl of DMSO and 2.0 μl of labeling enzyme were added to the mixture. The labeling reaction was incubated for 1 h at 16°C and was terminated by incubation at 65°C for 15 min. After termination, the Hy3TM-labeled samples were hybridized on the miRCURYTM LNA Array (v.18.0) according to the array manual.

extract_protocol: Twelve pairs of gender- and age-matched Group D dust mite-induced asthma children and Group N normal control children were included in the initial microarray-based discovery analysis. Five ml of EDTA-anticoagulant peripheral blood was collected from each child. Total RNA was isolated by using the TRIzol (Invitrogen, Grand Island, NY, USA) and the miRNeasy mini kits (Qiagen, Chatsworth, CA, USA) according to the manufacturer’s instructions. This protocol efficiently recovered all RNA species, including miRNAs. The quality and quantity of RNAs collected from all participants were measured with a nanodrop spectrophotometer ND-1000 (Nanodrop Technologies, Wilmington, DE, USA), and RNA integrity was determined by gel electrophoresis.

tag: subject status

value: normal control child

value: female

tag: age(year)

value: peripheral blood


title: N21

value: male


title: R24Sil_AGM91_AGM_SIV_SC_D7_1_Mono_RLT_RNA_mRNA_Seq_S8

value: Infected with SIV


title: R24Sil_AGM91_AGM_SIV_SC_D7_1_NCMono_RLT_RNA_mRNA_Seq_S11

value: Non-classical Monocytes


title: N19


title: N17


title: R24Sil_AGM91_AGM_SIV_SC_D28_1_NCMono_RLT_RNA_mRNA_Seq_S13


title: R24Sil_RCt14_RM_SIV_SC_D14_1_NCMono_RLT_mRNA_Seq_S7

value: Day 14


title: N15


title: N16


title: R24Sil_AGM91_AGM_SIV_SC_D28_1_Mono_RLT_RNA_mRNA_Seq_S10


title: R24Sil_RCt14_RM_SIV_SC_D14_1_Mono_RLT_mRNA_Seq_S3


title: R24Sil_AGM91_AGM_SIV_SC_D14_1_NCMono_RLT_RNA_mRNA_Seq_S12


title: N7


title: R24Sil_AGM91_AGM_SIV_SC_D14_1_Mono_RLT_RNA_mRNA_Seq_S9


title: R24Sil_AGM90_AGM_Uninf_SC_Dminus28_1_NCMono_RLT_RNA_mRNA_Seq_S4

source_name: Sorted Non-classical Monocytes  Uninfected

value: Uninfected


title: N6


title: R24Sil_AGM90_AGM_Uninf_SC_Dminus28_1_Mono_RLT_RNA_mRNA_Seq_S1


title: IF_24hr_185

city: Eugene

first: Amy

middle: K

last: Webster

state: Oregon

address: Institute of Ecology and Evolution

institute: University of Oregon

data_processing:
Bowtie version 1.1.2 was used to map reads to the WS210 version of the genome.
Count files were generated with HTSeq using the GTF file used in Maxwell et al., 2012. This file includes features of the WS210 genome in addition to features of WS220 mapped back to WS210.
edgeR was used on count tables for differential expression analysis. The count tables for the two batches were combined for filtering. The  count table was first filtered to contain those annotated as genes, reducing the number of genes to 22581.
Genes were further filtered by including only genes with counts per million (CPM) > 1 across 4 libraries. This resulted in 18190 genes. These were used for pairwise comparisons using the exact test in edgeR
Genome_build: WS210
Supplementary_files_format_and_content: Count files include the sequence identifier for each gene and the number of counts mapping to that gene (not normalized across libraries). An excel file of differential expression analyses is included.

organism: Caenorhabditis elegans

source_name: L1 larvae

growth_protocol: LRB269 maintained at 20C was chunked, then bleached three days later to obtain embryos. Embryos were resuspended in 10 mL S-basal at a density of 1 embryo/uL. For ivermectin samples, ivermectin was added at a concentration of 5 ng/mL. These were then placed on the carousel at 22C. After 24 hours, HB101 was added at a concentration of 25 mg/mL to the food samples

extract_protocol:
Total RNA was extracted using TRIzol with minor modifications to the manufacturer's protocol. 1mL of TRIzol was used per sample. Samples were homogenized using 100uL of acid-washed sand.
Libraries were prepared using the NEB Next Ultra Non-directional mRNA-seq Kit.

treatment_protocol: 1 hour after HB101 is added (25 hours post bleach), the 1hr samples are collected. 24 hours after HB101 is added, the 24hr samples are collected (48 hours post bleach). Fed samples were washed quickly 3-4X with S-basal to remove bacteria prior to collection. Collected samples were flash frozen in liquid nitrogen and stored at -80C until RNA isolation

tag: food or no food

value: food

tag: 1hr or 24 hr

tag: ivermectin or no ivermectin

value: ivermectin

tag: strain


title: IF_24hr_181


title: IF_24hr_186


title: R24Sil_RCt14_RM_SIV_SC_D28_1_Mono_RLT_mRNA_Seq_S4


title: IF_1hr_186


title: R24Sil_AGM90_AGM_SIV_SC_D7_1_NCMono_RLT_RNA_mRNA_Seq_S5

value: Day 7


title: R24Sil_AGM90_AGM_SIV_SC_D7_1_Mono_RLT_RNA_mRNA_Seq_S2


title: R24Sil_AGM90_AGM_SIV_SC_D28_1_NCMono_RLT_RNA_mRNA_Seq_S6


title: IF_1hr_185


title: R24Sil_AGM90_AGM_SIV_SC_D28_1_Mono_RLT_RNA_mRNA_Seq_S3


title: N3

tag: tissue


title: N5


title: N4


title: IF_24hr_183


title: RC_08: ASML_cld7mutPalm reconstituted

first: Jan

last: Provaznik

state: Baden-Wuerttemberg

address: Meyerhofstr. 1

department: Genomics Core Facility

institute: EMBL Heidelberg

hyb_protocol: Hybridization was done as described in miRNA Microarray System protocol v 1.7 from Agilent Technologies

scan_protocol: Scanning was done on Agilent Microarray Scanner G2565BA, Agilent Scan Control Software,

data_processing:
IPA was used for miRNA analysis and for correlating miRNA with protein expression after predicted targets were selected by miRNA databases (http://www.microrna.org, http://www.targetscan.org). For pathway analysis PANTHER (http://pantherdb.org), KEGG (http://www.kegg.jp), Reactome (https://reactome.org), and STRING (http://string-db.org) databases were used
after normalization, data were compared between identical non-transfected versus transfected cells and exosomes, respectively. In most instances, 2-fold differences were considered significant. In exosome preparations, 1.5-fold differences were occassionally included. For processed data, the programm suggestions were followed.

source_name: exosome

label_protocol: Labeling was done as described in miRNA Microarray System protocol v 1.7 from Agilent Technologies

growth_protocol:
The rat pancreatic adenocarcinoma lines ASML and AS spontaneously arisen in the BDX rat strain are maintained in RPMI1640/10%FCS/L-glutamine/antibiotics. Confluent cells are trypsinized and split.
ASML-EpC kd,ASML-cld7 kd, ASML-CD151 kd, ASML-Tspan8 kd, ASML-CD44v6 kd and AS-CD9 kd lines were derived from transfection with shRNA (Qiagen, Hilden, Germany) and maintained in RPMI1640/10%FCS/pyruvate/L-glutamine/antibiotics containing 0.5µg/ml G418. Single cell kd clones were derived by 2 rounds of single cell cloning. (EpC: EpCAM, cld7: claudin7)
ASML-EpCAM mutAG and ASML-cld7mutPalm were derived from ASML-EpC kd, respectively ASML-cld7 kd cells by transfection with mutated EpCAM (exchange of AG in the binding site of EpC to cld7) or mutation in the palmitoylation site of cld7 shRNA (Qiagen, Hilden, Germany). Cells were maintained in RPMI1640/10%FCS/pyruvate/L-glutamine/antibiotics containing 0.5µg/ml G418, 120µg hygromycin. Single cell kd clones were derived by 2 rounds of single cell cloning.

extract_protocol: Cell and exosome mRNA/microRNA was extracted using the miRNeasyMinikit following the supplier's suggestion (Qiagen, Hildesheim, Germany). Exosomes were pretreated with RNAse to avoid contamination by attached free mRNA.

treatment_protocol: Cells have not undergone any special treatment. All exosome preparation were derived from culture supernatant after cells have been cultured in serum-free medium for 48h-72h. Culture supernatant was collected and cleared (2x10min, 500g, 1x20min, 2000g, 1x30min, 10000g), filter sterizied (0.2µm) and were centrifuged (120min, 100000g). The pellet was washed (PBS, 120min, 100000g), resuspended in 40% sucrose overlaid by a discontinuous sucrose gradient (30%-5%) and centrifuged (16h, 100000g). Exosomes were collected from the 10%-5% sucrose interface (light density fractions, d: 1.15-1.56g/ml).

value: rat pancreatic adenocarcinoma

tag: cell line

value: ASML

tag: condition

value: cld7mutPalm reconstituted


title: RC_07: ASML_cld7kd


title: R24Sil_AGM91_AGM_Uninf_SC_Dminus27_1_Mono_RLT_RNA_mRNA_Seq_S7

description: Sorted Monocytes  Uninfected at Day minus27

value: Day minus27

tag: cell type

value: Monocytes


title: I_24hr_183

value: no food


title: I_24hr_186


title: I_24hr_185


title: HXP

source_name: dust mite_peripheral blood

value: dust mite-induced asthma child


title: CZP


title: I_24hr_181


title: SXJ

value: female


title: YZZ

value: peripheral blood


title: N2

value: normal control child


title: RC_04: ASML_CD44v6 Kd

value: CD44v6 Kd


title: IF_1hr_181


title: RC_05: ASML_EpCAM Kd

value: EpCAM Kd


title: RC_06: ASML_EpCAM mutAG reconstituted

value: EpCAM mutAG reconstituted


title: dCD-R1-mESC_Input

description:
Sample 24
Input chromatin used for ICeChIP-seq in Sample 23.

data_processing:
To align reads, a reference genome was first created, consisting of the mouse genome (mm9) appended by the sequences of each of the nucleosome standard barcodes. Reads were then mapped to this reference genome using Bowtie2 using the sensitive preset and end-to-end alignment options.
Using SAMTools, any reads which were not paired, not mapped in a proper pair, or mapped with a map quality below 20 were then discarded to prevent low-quality mapping or sequencing reads from impacting downstream analyses. Reads were then flattened to create a single mapping from each matched pair of reads, and any mappings with lengths greater than 200bp were also discarded to ensure only mononucleosomes were being analyzed.
Bedgraphs of genome coverage were then generated using BEDTools, and IP and input genome coverage bedgraphs were merged using BEDTools7. The sum of reads across ladder members for each nucleosomal standard was computed for each sample and HMD bedgraphs were then generated from the merged bedgraphs using awk to apply the following formula: HMD (%) = 100% * (IPlocus/INlocus) / (IPbarcode/INbarcode). Error and 95% confidence intervals were computed with Poisson statistics and error propagation from the merged bedgraphs using awk to apply the following formula: 95CI Error (%) = 1.96 * HMD (%) * sqrt(1/IPlocus + 1/INlocus). Bigwig files were generated for visualization using the bedGraphToBigWig tool.
To compute average HMD across a series of intervals, a “double mapping” procedure was used. First, the HMD bedgraph was mapped onto 1bp windows made for each interval using BEDTools. Then, the mapped windows were mapped onto the original intervals using BEDTools. This procedure ensured that the degree of overlap of the interval with each value of the HMD bedgraph was accounted for in the mapping procedure.
Genome_build: mm9_bar_Ca1-17 (mm9 appended by synthetic DNA barcodes used with semisynthetic nucleosome standards)
Supplementary_files_format_and_content: Bedgraph files all have the form <chromosome> <start> <end> <value>, where the fourth field represents the value of the measurement. Bigwig files are bedgraph files in binary format, appropriate for viewing on a genome browser. Fasta file (with extension .fa) represents the reference genome used in this study, with the mouse genome (mm9) appended by the DNA barcode sequences used for the semisynthetic nucleosome standards.

source_name: Cell Culture

growth_protocol: dCD MLL3/4 R1 mESCs were cultured as previously described (Dorighi et al. 2017).

extract_protocol:
dCD MLL3/4 mouse embryonic cell pellets were washed twice with 5 mL of PBS, then washed twice with 5 mL of Buffer N (15 mM Tris-HCl pH 7.5, 15 mM NaCl, 60 mM KCl, 8.5% w/v Sucrose, 5 mM MgCl2, 1 mM CaCl2, 1 mM DTT, 200 uM PMSF, 50 ug/mL BSA, 1x Roche Protease Inhibitor Cocktail, Filter Sterilized), with each wash consisting of complete resuspension of the pellet, centrifugation at 500 g for 5 minutes at 4°C, and removal of supernatant. The washed pellet was then resuspended in at least 2 packed cell volumes (PCV) of Buffer N and mixed with 1 volume of Lysis Buffer (Buffer N supplemented with 0.6% NP-40 Substitute) and incubated on ice for 10 minutes to lyse cells. The crude nuclei were spun down at 500 g for 5 minutes at 4°C before being resuspended in at least 6 packed nuclear volumes (PNV) of Buffer N and applied to the top of 7.5 mL of Sucrose Cushion N (15 mM Tris-HCl pH 7.5, 15 mM NaCl, 60 mM KCl, 30% w/v Sucrose, 5 mM MgCl2, 1 mM CaCl2, 1 mM DTT, 200 uM PMSF, 50 ug/mL BSA, 1x Roche Protease Inhibitor Cocktail, Filter Sterilized) in a 15 mL centrifuge tube, then spun down at 500 g for 12 minutes at 4°C in a swinging-bucket rotor. The supernatant was discarded after centrifugation, and the pellet was resuspended in approximately 2 PNV of Buffer N. The nuclei were then quantified by diluting 2 uL of nuclei suspension into 48 uL of 2 M NaCl, water-bath sonicating to solubilize DNA, and spectroscopically measuring DNA concentration by Nanodrop measurement, where 1 A280nm¬ = 50 ng/uL chromatin in the measurement sample. After accounting for the 25-fold dilution of the measurement sample, the concentration of the nuclei was adjusted down to 1ug/uL of chromatin. Nuclei were aliquoted into 100 uL aliquots, flash frozen, and stored at -80°C until use. When nuclei aliquots were used, they were first thawed and spiked approximately 1 uL of each barcoded nucleosome standard per 50 ug of chromatin to be used. The nuclei suspension with spiked calibrants was then mixed with a pipette and transferred to a new tube, then warmed to 37°C for 2 minutes. After warming, 1 unit of micrococcal nuclease (MNase) per 4.375 ug of chromatin was added to the nuclei suspension and incubated at 37°C while shaking at 900 rpm for 12 minutes. After incubation, the digestion was stopped by adding 1/9 volume of 10x MNase Stop Buffer (100 mM EDTA, 100 mM EGTA, Filter Sterilized) while slowly vortexing, and nuclei were lysed by adding 5 M NaCl to a final concentration of 600 mM NaCl while slowly vortexing. 66 mg of HAP resin (BioRad, CHTTM Ceramic Hydroxyapatite, Type I, 20 um) per 100 ug of chromatin digested was rehydrated with 200 uL of HAP Buffer 1 (5 mM Sodium Phosphate pH 7.2, 600 mM NaCl, 1 mM EDTA, 200 uM PMSF, Filter Sterilized) per 100 ug of chromatin digested. The lysed nuclei were then spun down at 18,000 g for 1 minute to pellet insoluble nuclear debris, and the soluble fraction was added to the rehydrated HAP resin and incubated for 10 minutes on a rotator at 4°C. After incubation, the HAP resin slurry was added to a centrifugal filter unit (Millipore Ultrafree MC–HV Centrifugal Filter 0.45 um) and spun at 1000 g for 30 seconds at 4°C. The HAP resin left on the filter unit was then washed 4 times with 200 uL HAP Buffer 1 and 4 times with 200 uL HAP Buffer 2 (5 mM Sodium Phosphate pH 7.2, 100 mM NaCl, 1 mM EDTA, 200 uM PMSF, Filter Sterilized) by spinning at 1000 g for 30 seconds at 4°C. Into a clean tube, the HAP resin was eluted with three elutions of 100 uL HAP Elution Buffer (500 mM Sodium Phosphate pH 7.2, 100 mM NaCl, 1 mM EDTA, 200 uM PMSF, Filter Sterilized). The DNA content of the elution was then quantified spectroscopically with a Nanodrop, and the chromatin concentration was adjusted to 20 ng/uL with ChIP Buffer 1 (25 mM Tris pH 7.5, 5 mM MgCl2, 100 mM KCl, 10% v/v glycerol, 0.1% v/v NP-40 Substitute, Filter Sterilized) with 100 ug/mL of BSA. With the exception of the Koide Lab 304M3B antibody, every ICeChIP was conducted with 12.5 uL of Protein A Dynabeads (Invitrogen) and 3 ug of antibody. The Protein A Dynabeads for each ICeChIP were washed with 50 uL of ChIP Buffer 1 by use of a magnetic rack, then resuspended in 50 uL of ChIP Buffer 1. In a separate set of tubes, 3 ug of each antibody was diluted to 100 uL with ChIP Buffer 1. The antibody suspension and the Protein A Dynabeads suspension were combined and incubated on a rotator at 4°C for at least 1 hour, then washed with 200 uL of ChIP Buffer 1 by use of a magnetic rack and resuspended in 50 uL of ChIP Buffer 1. After antibody washing, 150 uL of the diluted chromatin elution was added to each antibody-bead conjugate and incubated for 15 minutes on a rotator at 4°C. After this incubation, the beads were washed twice with ChIP Buffer 2 (25 mM Tris pH 7.5, 5 mM MgCl2, 300 mM KCl, 10% v/v glycerol, 0.1% v/v NP-40 Substitute, Filter Sterilized) and once with ChIP Buffer 3 (10 mM Tris pH 7.5, 250 mM LiCl, 1 mM EDTA, 0.5% Sodium Deoxycholate, 0.5% v/v NP-40 Substitute, Filter Sterilized), with a wash consisting of removal of the existing supernatant by use of a magnetic rack, resuspension into 150 uL of buffer, transfer to a new siliconized tube, and incubation on the rotator for 10 minutes at 4°C. After these washes, the supernatant was removed, and the beads were resuspended in ChIP Buffer 1, transferred to a new siliconized tube, then rinsed once with 200 uL of TE before being resuspended in 50 uL of ChIP Elution Buffer and incubated at 55°C for 5 minutes. After incubation, the supernatant was transferred to a new set of siliconized tubes, and the beads were discarded. To each supernatant was then added 2 uL of 5 M NaCl, 1 uL of 500 mM EDTA, and 1 uL of 10 mg/mL Proteinase K. 15 uL of Input DNA was also diluted to 50 uL with 35 uL of ChIP Elution Buffer (50 mM Tris pH 7.5, 1 mM EDTA, 1% w/v SDS, Filter Sterilized) and was supplemented with 2 uL of 5 M NaCl, 1 uL of 500 mM EDTA, and 1 uL of 10 mg/mL Proteinase K. The IP elutions and diluted input were then incubated at 55°C for 2 hours for a Proteinase K digestion. After digestion, the DNA was purified by adding 1.5 volumes of Serapure HD (1:50 dilution of Sera-Mag SpeedBeads [Fisher], 20% PEG-8000, 2.5 M NaCl, 10 mM Tris pH 7.5, 1 mM EDTA, 0.05% Tween-20, Filter Sterilized prior to addition of SpeedBeads), incubating at room temperature for 15 minutes, then collecting the beads on a magnetic rack, washing twice with 150 uL of 70% ethanol, and eluting into 50 uL of water, which was then recovered and stored at -20°C.
DNA purified with Serapure was quantified using Quant-iT TM PicoGreen (Thermo Fisher) per manufacturer instructions. Up to 10 ng of each DNA sample for which libraries were to be prepared were then carried forward. Libraries were then generated from the NEBNext Ultra II DNA Library Prep kit (New England Biolabs) per manufacturer instructions. The DNA content of the library was then quantified and pooled for Illumina sequencing.

value: dCD MLL3/4 R1 mESC

tag: chip antibody

tag: amplification cycles


title: YSX

value: male


title: JY


title: I_1hr_186

value: ivermectin


title: IF_1hr_183

value: food


title: LYT