title: hpf14-61

description:
processed data file:
hpf14.txt
hpf14-S155


title: hpf14-64

status: Public on Jun 15 2017

type: SRA

city: New York

first: wei

last: wang

state: NY

address: 100 washington square east

department: Biology

country: USA

institute: New York University

description:
processed data file:
hpf14.txt
hpf14-S158

library_source: transcriptomic

data_processing:
For each demultiplexed single cell RNA-seq library, sequencing reads were mapped to Ciona genome (Joined-scaffold (KH), http://ghost.zool.kyotou.ac.jp/datas/JoinedScaffold.zip) using TopHat 2.0.12 (Kim et al., 2013; Trapnell et al., 2012) with parameter: --no-coverage-search. Cufflinks 2.2.0 (Trapnell et al., 2012) was used to calculate the Fragments Per Kilobase of transcript per Million mapped reads (FPKM) using KH gene models (ver.2013, http://ghost.zool.kyoto-u.ac.jp/datas/KH.KHGene.2013.gff3.zip) with default parameters.
Genome_build: Ciona genome (Joined-scaffold (KH), http://ghost.zool.kyotou.ac.jp/datas/JoinedScaffold.zip)
Supplementary_files_format_and_content: tab-delimited text files include raw FPKMs for each gene per cell

molecule: polyA RNA

organism: Ciona robusta

source_name: ScRNA-seq of FACS purified cardiopharyngeal progenitors_14hpf

extract_protocol:
Sample dissociation and FACS were performed essentially as described (Christiaen et al., 2009c; Razy-Krajka et al., 2014). Embryos and larvae were harvested at 12, 14, 16, 18 and 20hpf in 5ml borosilicate glass tubes (Fisher Scientific, Waltham, MA. Cat.No. 14-961-26) and washed with 2ml calcium- and magnesium-free artificial seawater (CMF-ASW: 449 mM NaCl, 33 mM Na2SO4, 9 mM KCl, 2.15 mM NaHCO3, 10 mM Tris-Cl pH 8.2, 2.5 mM EGTA). Embryos and larvae were dissociated in 2ml 0.2% trypsin (w/v, Sigma, T- 4799) ASW by pipetting with glass Pasteur pipettes. The dissociation was stopped by adding 2ml filtered ice cold 0.05% BSA CMF-ASW. The dissociated cells were passed through 40µm cell-strainer and collected in 5ml polystyrene round-bottom tube (Corning Life Sciences, Oneonta, New York. REF 352235). Cells were collected by centrifugation at 800g for 3 min at 4°C, followed by two rounds of washing with ice cold 0.05% BSA CMF-ASW. Cell suspensions were filtered again through a 40µm cell-strainer and stored on ice. Following dissociation, cell suspensions were used for sorting within 1 hour. The B7.5 lineage cells were labeled by Mesp>tagRFP reporter. Contaminating B-line mesenchyme cells were counter-selected using  MyoD905>EGFP as described (Christiaen et al., 2008; Razy-Krajka et al., 2014).  The TVC-specific Hand-r>tagBFP reporter was used in a 3-color FACS scheme for positive co-selection of TVC-derived cells, in order to minimize the effects of mosaicism. Dissociated cell were loaded in the BD FACS AriaTM cell sorter.  488 nm laser, FITC filter was used for EGFP; 407 nm laser, 561 nm laser, DsRed filter was used for tagRFP and Pacific BlueTM filter was used for tagBFP. The nozzle size was 100 µm. tagRFP +, tagBFP + and EGFP – cells were collected for downstream RNA sequencing analysis.
Reverse transcription and cDNA amplification were carried out using modified Smart-seq2 protocol (Picelli et al., 2013). Single cells were sorted by FACS as described above into 96-well plates and collected in 3.4 µl RT buffer (0.5 µl 10µM 3’ RT Primer ( 5' - AAG CAG TGG TAT CAA CGC AGA GTA C T30 VN - 3’), 0.5µl 10µM dNTP Mix, 0.5 µl 4U/µl RNase Inhibitor, 1 µl Maxima RT Buffer, 0,9 µl nuclease-free water) in each well. Plates were stored at -80°C  or processed immediately.  Plates were incubated at 72°C for 3min and chilled on the ice to denature the template RNA. 2µl  RT reaction mixture (0.5µl 10um TSO primer (5′-AGACGTGTGCTCTTCCGATCTNNNNNrGrGrG-3′), 0.925µl 5M Betaine, 0.4 100mM MgCl2, 0.125µl 40U/ RNAase inhibitor, 0.05µl 200 U/µl Maxima H Minus Reverse Transcriptase) were added to each well. Reverse transcription was carried out by incubating the plate at 42°C for 90 min, followed by 10 cycles of (50 °C for 2 min, 42 °C for 2 min) and heat inactivation at 70° for 15 min. 7µl PCR amplification mixture (0.25µl 10 μM PCR primer (5′AGACGTGTGCTCTTCCGATCT-3′), 6.25µl KAPA HIFI ReadyMix, 0.5µl nuclease-free water) were added to each well. PCR amplification was carried out with a denaturation at 98 °C for 3 min, followed by 21 cycles of (98 °C for 15 s, 67 °C for 20 s, and 72 °C for 6 min) and a final extension at 72° for 5 min. PCR products were purified by adding 10µl (0.8×) Agencourt AMPureXP SPRI beads (Beckman-Coulter) to each well, followed by 5min incubation and two rounds of wash using 100µl freshly prepared  70% Ethanol at room temperature. Purified cDNA were eluted in 20µl TE. The concentration of amplified cDNA was measured across the entire plate using Picogreen assay. The concentration of amplified cDNA was in a 0.5–2 ng/μl range.  Fragment size distribution was checked for randomly selected wells with High-Sensitivity Bioanalyzer Chip (Agilent), the expected size average should be ~2 kb. For each sample, the amplified cDNA were normalized to a working concentration ranging from 0.1 to 0.2 ng/µl with TE buffer. 1.25 µl of diluted cDNA from each well were used for library preparation. Single cell libraries were prepared using the Nextera XT DNA Sample Kit (Illumina) according to manufacturer's instructions.  After library amplification, 2.5μl from each well were pooled into a single 1.5-ml microcentrifuge tube, purified using Agencourt AMPure XP beads and eluted with 30μl TE buffer.  1μl purified library was used to measure the fragment size distribution using the Agilent HS DNA BioAnalyzer chip and another 1μl of the purified library was loaded into Qubit fluorometer to estimate library concentration according to the manufacturer's instructions.  Libraries were sequenced on an Illumina HiSeq 2500 sequencer to obtain paired-end 25bp reads.

tag: genotype/variation

value: Wild type

tag: developmental stage

value: Late tailbud II

tag: cell type

value: Trunk ventral cells

tag: cell subtype

value: Multipotent cardiopharyngeal progenitor cells


title: hpf14-56

description:
processed data file:
hpf14.txt
hpf14-S150


title: hpf14-53

description:
processed data file:
hpf14.txt
hpf14-S148


title: hpf14-47

description:
processed data file:
hpf14.txt
hpf14-S142


title: hpf14-51

description:
processed data file:
hpf14.txt
hpf14-S146


title: hpf14-49

description:
processed data file:
hpf14.txt
hpf14-S144


title: hpf14-58

description:
processed data file:
hpf14.txt
hpf14-S152


title: hpf14-44

description:
processed data file:
hpf14.txt
hpf14-S13


title: hpf14-57

description:
processed data file:
hpf14.txt
hpf14-S151


title: hpf14-43

description:
processed data file:
hpf14.txt
hpf14-S139


title: hpf14-52

description:
processed data file:
hpf14.txt
hpf14-S147


title: hpf14-71

description:
processed data file:
hpf14.txt
hpf14-S164


title: hpf14-54

description:
processed data file:
hpf14.txt
hpf14-S149


title: hpf14-50

description:
processed data file:
hpf14.txt
hpf14-S145


title: hpf14-48

description:
processed data file:
hpf14.txt
hpf14-S143


title: hpf14-42

description:
processed data file:
hpf14.txt
hpf14-S138


title: hpf14-46

description:
processed data file:
hpf14.txt
hpf14-S141


title: hpf14-45

description:
processed data file:
hpf14.txt
hpf14-S140


title: hpf14-41

description:
processed data file:
hpf14.txt
hpf14-S137


title: hpf14-39

description:
processed data file:
hpf14.txt
hpf14-S135


title: hpf14-55

description:
processed data file:
hpf14.txt
hpf14-S14


title: hpf14-38

description:
processed data file:
hpf14.txt
hpf14-S134


title: hpf14-33

description:
processed data file:
hpf14.txt
hpf14-S12


title: hpf14-35

description:
processed data file:
hpf14.txt
hpf14-S131


title: hpf14-37

description:
processed data file:
hpf14.txt
hpf14-S133


title: hpf14-32

description:
processed data file:
hpf14.txt
hpf14-S129


title: hpf14-29

description:
processed data file:
hpf14.txt
hpf14-S126


title: hpf14-30

description:
processed data file:
hpf14.txt
hpf14-S127


title: hpf14-28

description:
processed data file:
hpf14.txt
hpf14-S125


title: hpf14-31

description:
processed data file:
hpf14.txt
hpf14-S128


title: hpf14-24

description:
processed data file:
hpf14.txt
hpf14-S121


title: hpf14-25

description:
processed data file:
hpf14.txt
hpf14-S122


title: hpf14-23

description:
processed data file:
hpf14.txt
hpf14-S120


title: hpf14-22

description:
processed data file:
hpf14.txt
hpf14-S11


title: hpf14-21

description:
processed data file:
hpf14.txt
hpf14-S119


title: hpf14-20

description:
processed data file:
hpf14.txt
hpf14-S118


title: hpf14-19

description:
processed data file:
hpf14.txt
hpf14-S117


title: hpf14-40

description:
processed data file:
hpf14.txt
hpf14-S136


title: hpf14-18

description:
processed data file:
hpf14.txt
hpf14-S116


title: hpf14-36

description:
processed data file:
hpf14.txt
hpf14-S132


title: hpf14-17

description:
processed data file:
hpf14.txt
hpf14-S115


title: hpf14-14

description:
processed data file:
hpf14.txt
hpf14-S112


title: hpf14-13

description:
processed data file:
hpf14.txt
hpf14-S111


title: hpf14-15

description:
processed data file:
hpf14.txt
hpf14-S113


title: hpf14-16

description:
processed data file:
hpf14.txt
hpf14-S114


title: hpf14-34

description:
processed data file:
hpf14.txt
hpf14-S130


title: hpf14-11

description:
processed data file:
hpf14.txt
hpf14-S10


title: hpf14-12

description:
processed data file:
hpf14.txt
hpf14-S110


title: hpf14-10

description:
processed data file:
hpf14.txt
hpf14-S109


title: hpf14-7

description:
processed data file:
hpf14.txt
hpf14-S106


title: hpf14-26

description:
processed data file:
hpf14.txt
hpf14-S123


title: hpf14-9

description:
processed data file:
hpf14.txt
hpf14-S108


title: hpf14-27

description:
processed data file:
hpf14.txt
hpf14-S124


title: hpf14-6

description:
processed data file:
hpf14.txt
hpf14-S105


title: hpf14-5

description:
processed data file:
hpf14.txt
hpf14-S104


title: hpf14-3

description:
processed data file:
hpf14.txt
hpf14-S102


title: hpf14-2

description:
processed data file:
hpf14.txt
hpf14-S101


title: hpf14-1

description:
processed data file:
hpf14.txt
hpf14-S100


title: hpf12-192

description:
processed data file:
hpf12.txt
hpf12_p4_9_R1

source_name: ScRNA-seq of FACS purified cardiopharyngeal progenitors_12hpf

value: Late tailbud I


title: hpf12-191

description:
processed data file:
hpf12.txt
hpf12_p4_96_R1


title: hpf12-189

description:
processed data file:
hpf12.txt
hpf12_p4_94_R1


title: hpf12-188

description:
processed data file:
hpf12.txt
hpf12_p4_93_R1


title: hpf12-190

description:
processed data file:
hpf12.txt
hpf12_p4_95_R1


title: hpf12-187

description:
processed data file:
hpf12.txt
hpf12_p4_92_R1


title: hpf12-186

description:
processed data file:
hpf12.txt
hpf12_p4_91_R1


title: hpf12-181

description:
processed data file:
hpf12.txt
hpf12_p4_87_R1


title: hpf12-184

description:
processed data file:
hpf12.txt
hpf12_p4_8_R1


title: hpf12-183

description:
processed data file:
hpf12.txt
hpf12_p4_89_R1


title: hpf12-179

description:
processed data file:
hpf12.txt
hpf12_p4_85_R1


title: hiPSC_Astrocytes2_activated_rep2

city: Freiburg im Breisgau

first: Galina

last: Erikson

address: Stübeweg 51

country: Germany

institute: Max Planck Institute of Immunobiology and Epigenetics

description:
mRNA
Processed data file: raw.txt
Processed data file: fpkm.txt

data_processing:
Sequenced reads were quality tested using FASTQC v0.11.2 using the default parameters.
Alignment to the mm10 genome was performed using the STAR aligner version 2.4.0k using the --outFilterIntronMotifs RemoveNoncanonical parameter
Gene expression was quantified using the HOMER analysis suite and differential gene expression was carried out using the edgeR package v3.6.8 using duplicates to compute within-group dispersion. FDR < 0.05 and log2 fold change greater than 0.5.
Genome_build: mm10 (GRCm38)
Supplementary_files_format_and_content: raw.txt: Tab-delimited text file includes raw counts.
Supplementary_files_format_and_content: fpkm.txt: Tab-delimited text file includes fpkm normalized table.

molecule: total RNA

organism: Homo sapiens

source_name: skin biopsy

growth_protocol: The growth protocols have been described in our study (Santos et al.)

extract_protocol:
in vitro derivation from human iPSC.
RNA libraries were prepared for sequencing using standard Illumina protocols.

value: induced pluripotent stem cells-derived astrocytes

tag: genotype

value: neurotypical

tag: treatment


title: hpf12-180

description:
processed data file:
hpf12.txt
hpf12_p4_86_R1


title: hpf12-182

description:
processed data file:
hpf12.txt
hpf12_p4_88_R1


title: hpf12-185

description:
processed data file:
hpf12.txt
hpf12_p4_90_R1


title: hpf12-175

description:
processed data file:
hpf12.txt
hpf12_p4_81_R1


title: hpf12-177

description:
processed data file:
hpf12.txt
hpf12_p4_83_R1


title: hpf12-178

description:
processed data file:
hpf12.txt
hpf12_p4_84_R1


title: hpf12-173

description:
processed data file:
hpf12.txt
hpf12_p4_7_R1


title: hpf12-174

description:
processed data file:
hpf12.txt
hpf12_p4_80_R1


title: hpf14-8

description:
processed data file:
hpf14.txt
hpf14-S107


title: hpf12-172

description:
processed data file:
hpf12.txt
hpf12_p4_79_R1


title: hpf12-171

description:
processed data file:
hpf12.txt
hpf12_p4_78_R1


title: hpf14-4

description:
processed data file:
hpf14.txt
hpf14-S103


title: hpf12-170

description:
processed data file:
hpf12.txt
hpf12_p4_77_R1


title: hpf12-160

description:
processed data file:
hpf12.txt
hpf12_p4_68_R1


title: hpf12-167

description:
processed data file:
hpf12.txt
hpf12_p4_74_R1


title: hpf12-159

description:
processed data file:
hpf12.txt
hpf12_p4_67_R1


title: hpf12-168

description:
processed data file:
hpf12.txt
hpf12_p4_75_R1


title: hpf12-165

description:
processed data file:
hpf12.txt
hpf12_p4_72_R1


title: hpf12-161

description:
processed data file:
hpf12.txt
hpf12_p4_69_R1


title: hpf12-166

description:
processed data file:
hpf12.txt
hpf12_p4_73_R1


title: hpf12-162

description:
processed data file:
hpf12.txt
hpf12_p4_6_R1


title: hpf12-164

description:
processed data file:
hpf12.txt
hpf12_p4_71_R1


title: hpf12-176

description:
processed data file:
hpf12.txt
hpf12_p4_82_R1


title: hpf12-158

description:
processed data file:
hpf12.txt
hpf12_p4_66_R1


title: hpf12-155

description:
processed data file:
hpf12.txt
hpf12_p4_63_R1


title: hpf12-153

description:
processed data file:
hpf12.txt
hpf12_p4_61_R1


title: hpf12-157

description:
processed data file:
hpf12.txt
hpf12_p4_65_R1


title: hpf12-154

description:
processed data file:
hpf12.txt
hpf12_p4_62_R1


title: hpf12-169

description:
processed data file:
hpf12.txt
hpf12_p4_76_R1


title: hpf12-163

description:
processed data file:
hpf12.txt
hpf12_p4_70_R1


title: hpf12-152

description:
processed data file:
hpf12.txt
hpf12_p4_60_R1


title: hpf12-150

description:
processed data file:
hpf12.txt
hpf12_p4_59_R1


title: hpf12-151

description:
processed data file:
hpf12.txt
hpf12_p4_5_R1


title: hpf12-149

description:
processed data file:
hpf12.txt
hpf12_p4_58_R1


title: hpf12-148

description:
processed data file:
hpf12.txt
hpf12_p4_57_R1


title: hpf12-146

description:
processed data file:
hpf12.txt
hpf12_p4_55_R1


title: hpf12-145

description:
processed data file:
hpf12.txt
hpf12_p4_54_R1


title: hpf12-144

description:
processed data file:
hpf12.txt
hpf12_p4_53_R1


title: hpf12-142

description:
processed data file:
hpf12.txt
hpf12_p4_51_R1


title: hpf12-147

description:
processed data file:
hpf12.txt
hpf12_p4_56_R1


title: hpf12-143

description:
processed data file:
hpf12.txt
hpf12_p4_52_R1


title: hpf12-141

description:
processed data file:
hpf12.txt
hpf12_p4_50_R1


title: hpf12-156

description:
processed data file:
hpf12.txt
hpf12_p4_64_R1


title: hpf12-139

description:
processed data file:
hpf12.txt
hpf12_p4_49_R1


title: hpf12-137

description:
processed data file:
hpf12.txt
hpf12_p4_47_R1


title: hpf12-132

description:
processed data file:
hpf12.txt
hpf12_p4_42_R1


title: hpf12-138

description:
processed data file:
hpf12.txt
hpf12_p4_48_R1


title: hpf12-133

description:
processed data file:
hpf12.txt
hpf12_p4_43_R1


title: hpf12-134

description:
processed data file:
hpf12.txt
hpf12_p4_44_R1


title: hpf12-140

description:
processed data file:
hpf12.txt
hpf12_p4_4_R1


title: hpf12-136

description:
processed data file:
hpf12.txt
hpf12_p4_46_R1


title: hpf12-135

description:
processed data file:
hpf12.txt
hpf12_p4_45_R1


title: hpf12-125

description:
processed data file:
hpf12.txt
hpf12_p4_36_R1


title: hpf12-127

description:
processed data file:
hpf12.txt
hpf12_p4_38_R1


title: hpf12-115

description:
processed data file:
hpf12.txt
hpf12_p4_27_R1


title: hpf12-119

description:
processed data file:
hpf12.txt
hpf12_p4_30_R1


title: hpf12-126

description:
processed data file:
hpf12.txt
hpf12_p4_37_R1


title: hpf12-131

description:
processed data file:
hpf12.txt
hpf12_p4_41_R1


title: hpf12-130

description:
processed data file:
hpf12.txt
hpf12_p4_40_R1


title: hpf12-123

description:
processed data file:
hpf12.txt
hpf12_p4_34_R1


title: hpf12-129

description:
processed data file:
hpf12.txt
hpf12_p4_3_R1


title: hpf12-116

description:
processed data file:
hpf12.txt
hpf12_p4_28_R1


title: hpf12-118

description:
processed data file:
hpf12.txt
hpf12_p4_2_R1


title: hpf12-111

description:
processed data file:
hpf12.txt
hpf12_p4_23_R1


title: hpf12-117

description:
processed data file:
hpf12.txt
hpf12_p4_29_R1


title: hpf12-113

description:
processed data file:
hpf12.txt
hpf12_p4_25_R1


title: hpf12-128

description:
processed data file:
hpf12.txt
hpf12_p4_39_R1


title: hpf12-121

description:
processed data file:
hpf12.txt
hpf12_p4_32_R1


title: hpf12-114

description:
processed data file:
hpf12.txt
hpf12_p4_26_R1


title: hpf12-120

description:
processed data file:
hpf12.txt
hpf12_p4_31_R1


title: hpf12-124

description:
processed data file:
hpf12.txt
hpf12_p4_35_R1


title: hpf12-112

description:
processed data file:
hpf12.txt
hpf12_p4_24_R1


title: hpf12-109

description:
processed data file:
hpf12.txt
hpf12_p4_21_R1


title: hpf12-122

description:
processed data file:
hpf12.txt
hpf12_p4_33_R1


title: hpf12-107

description:
processed data file:
hpf12.txt
hpf12_p4_1_R1


title: hpf12-108

description:
processed data file:
hpf12.txt
hpf12_p4_20_R1


title: hpf12-104

description:
processed data file:
hpf12.txt
hpf12_p4_17_R1


title: hpf12-103

description:
processed data file:
hpf12.txt
hpf12_p4_16_R1


title: hpf12-102

description:
processed data file:
hpf12.txt
hpf12_p4_15_R1


title: hpf12-110

description:
processed data file:
hpf12.txt
hpf12_p4_22_R1


title: hpf12-100

description:
processed data file:
hpf12.txt
hpf12_p4_13_R1


title: hpf12-105

description:
processed data file:
hpf12.txt
hpf12_p4_18_R1


title: hpf12-101

description:
processed data file:
hpf12.txt
hpf12_p4_14_R1


title: hpf12-99

description:
processed data file:
hpf12.txt
hpf12_p4_12_R1


title: hpf12-96

description:
processed data file:
hpf12.txt
hpf12_p3_9_R1


title: hpf12-86

description:
processed data file:
hpf12.txt
hpf12_p3_88_R1


title: hpf12-75

description:
processed data file:
hpf12.txt
hpf12_p3_78_R1


title: hpf12-76

description:
processed data file:
hpf12.txt
hpf12_p3_79_R1


title: hpf12-97

description:
processed data file:
hpf12.txt
hpf12_p4_10_R1


title: hpf12-89

description:
processed data file:
hpf12.txt
hpf12_p3_90_R1


title: hpf12-98

description:
processed data file:
hpf12.txt
hpf12_p4_11_R1


title: hpf12-93

description:
processed data file:
hpf12.txt
hpf12_p3_94_R1


title: hpf12-95

description:
processed data file:
hpf12.txt
hpf12_p3_96_R1


title: hpf12-85

description:
processed data file:
hpf12.txt
hpf12_p3_87_R1


title: hpf12-84

description:
processed data file:
hpf12.txt
hpf12_p3_86_R1


title: hpf12-94

description:
processed data file:
hpf12.txt
hpf12_p3_95_R1


title: hpf12-92

description:
processed data file:
hpf12.txt
hpf12_p3_93_R1


title: hpf12-90

description:
processed data file:
hpf12.txt
hpf12_p3_91_R1


title: hpf12-78

description:
processed data file:
hpf12.txt
hpf12_p3_80_R1


title: hpf12-83

description:
processed data file:
hpf12.txt
hpf12_p3_85_R1


title: hpf12-77

description:
processed data file:
hpf12.txt
hpf12_p3_7_R1


title: hpf12-88

description:
processed data file:
hpf12.txt
hpf12_p3_8_R1


title: hpf12-82

description:
processed data file:
hpf12.txt
hpf12_p3_84_R1


title: hpf12-106

description:
processed data file:
hpf12.txt
hpf12_p4_19_R1


title: hpf12-74

description:
processed data file:
hpf12.txt
hpf12_p3_77_R1


title: hpf12-81

description:
processed data file:
hpf12.txt
hpf12_p3_83_R1


title: hpf12-87

description:
processed data file:
hpf12.txt
hpf12_p3_89_R1


title: hpf12-73

description:
processed data file:
hpf12.txt
hpf12_p3_76_R1


title: hpf12-91

description:
processed data file:
hpf12.txt
hpf12_p3_92_R1


title: hpf12-80

description:
processed data file:
hpf12.txt
hpf12_p3_82_R1


title: hpf12-70

description:
processed data file:
hpf12.txt
hpf12_p3_73_R1


title: hpf12-71

description:
processed data file:
hpf12.txt
hpf12_p3_74_R1


title: hpf12-72

description:
processed data file:
hpf12.txt
hpf12_p3_75_R1


title: hpf12-68

description:
processed data file:
hpf12.txt
hpf12_p3_71_R1


title: hpf12-67

description:
processed data file:
hpf12.txt
hpf12_p3_70_R1


title: hpf12-62

description:
processed data file:
hpf12.txt
hpf12_p3_66_R1


title: hpf12-52

description:
processed data file:
hpf12.txt
hpf12_p3_57_R1


title: hpf12-66

description:
processed data file:
hpf12.txt
hpf12_p3_6_R1


title: hpf12-59

description:
processed data file:
hpf12.txt
hpf12_p3_63_R1


title: hpf12-61

description:
processed data file:
hpf12.txt
hpf12_p3_65_R1


title: hpf12-65

description:
processed data file:
hpf12.txt
hpf12_p3_69_R1


title: hpf12-63

description:
processed data file:
hpf12.txt
hpf12_p3_67_R1


title: hpf12-53

description:
processed data file:
hpf12.txt
hpf12_p3_58_R1


title: hpf12-58

description:
processed data file:
hpf12.txt
hpf12_p3_62_R1


title: hpf12-60

description:
processed data file:
hpf12.txt
hpf12_p3_64_R1


title: hpf12-64

description:
processed data file:
hpf12.txt
hpf12_p3_68_R1


title: hpf12-55

description:
processed data file:
hpf12.txt
hpf12_p3_5_R1


title: hpf12-54

description:
processed data file:
hpf12.txt
hpf12_p3_59_R1


title: hpf12-69

description:
processed data file:
hpf12.txt
hpf12_p3_72_R1


title: hpf12-51

description:
processed data file:
hpf12.txt
hpf12_p3_56_R1


title: hpf12-56

description:
processed data file:
hpf12.txt
hpf12_p3_60_R1


title: hpf12-50

description:
processed data file:
hpf12.txt
hpf12_p3_55_R1


title: two-weeks seedling plants

city: Shanghai

first: yiwen

last: Deng

state: Shanghai

address: 300 Fenglin Road

department: Institute of Plant Physiology and Ecology

country: China

institute: Shanghai Institute for Biological Sciences, Chinese Academy of Sciences

data_processing:
Illumina Casava1.7 software used for basecalling.
Sequenced reads were trimmed for adaptor sequence, and masked for low-complexity or low-quality sequence, then mapped to the rice reference genome Nipponbare genome using software SOAP  with parameters -v 0 -r 2 -M 0 -a clean.fa -D ref_genome.fa.index -o match_genome.soap
Genome_build: Os-Nipponbare-Reference-IRGSP-1.0 pseudomolecules
Supplementary_files_format_and_content: abundance measurements

organism: Oryza sativa

source_name: leaf

growth_protocol: The variety NIL-Pigm (the isogenic lines harboring Pigm dependent on the recurrent variety NIPB) and NIPB plants were grown in field in Shanghai station, the total RNA samples of leaf were isolated using the two-week old seedlings.

extract_protocol:
RNA was extracted with Trizol reagent, followed by clean-up and DNase I treatment with QIAGEN RNeasy mini kit in accordance with the prescribed protocol provided with the kit. Quality control was performed with Agilent Bioanalyser.
RNA libraries were prepared for sequencing using standard Illumina protocols

tag: development stage

value: seedling

tag: tissue

value: leaf

tag: cultivar

value: NIL-Pigm


title: hpf12-49

description:
processed data file:
hpf12.txt
hpf12_p3_54_R1


title: hpf12-48

description:
processed data file:
hpf12.txt
hpf12_p3_53_R1


title: CME181

city: Potsdam

first: Thomas

last: Lufkin

address: 8 Clarkson Avenue

institute: Clarkson University

library_source: genomic

data_processing:
BED_Alignment: Sequence reads were obtained and mapped to the Mouse genomes (CME018,CME019 mapped to mm8, CME179,CME180,CME181,CME182 mapped to mm9) using the Illumina Genome Analyzer Pipeline (Eland). All uniquely mapped reads that are with two or fewer mismatches were retained.
LIFTOVER_TXT_Peaks: (Treatment: CME018;  Control: CME019) Peak detection was performed with the Model-based Analysis of ChIP-Seq (MACS ver. 1.4) algorithm (http://liulab.dfci.harvard.edu/MACS/), and the called peaks are lifted over from mm8 to mm9 using UCSC liftover tools (http://genome.ucsc.edu/) .
TXT_Peaks: (Treatment: CME180;  Control: CME179), (Treatment: CME182;  Control: CME181) Peak detection was performed with the Model-based Analysis of ChIP-Seq (MACS ver. 1.4) algorithm (http://liulab.dfci.harvard.edu/MACS/).

molecule: genomic DNA

organism: Mus musculus

source_name: Sox6 ChIP-seq IgG Control

extract_protocol: Lysates  was obtained from sonicated nuclei and protein-DNA complexes were immunoprecipitated with respective antibodies.  Libraries were prepared according to Illumina's instructions accompanying the DNA Sample Kit (Part# 0801-0303). Briefly, DNA was end-repaired using a combination of T4 DNA polymerase, E. coli DNA Pol I large fragment (Klenow polymerase) and T4 polynucleotide kinase. The blunt, phosphorylated ends were treated with Klenow fragment (32 to 52 exo minus) and dATP to yield a protruding 3- 'A' base for ligation of Illumina's adapters which have a single 'T' base overhang at the 3’ end. After adapter ligation DNA was PCR amplified with Illumina primers for 15 cycles and library fragments of ~200 bp (insert plus adaptor and PCR primer sequences) were band isolated from an agarose gel. The purified DNA was captured on an Illumina flow cell for cluster generation. Libraries were sequenced on the Genome Analyzer following the manufacturer's protocols.

tag: Stage

value: E13.5 embryos

tag: strain

value: CD1 wildtype

value: Limbs and tails

tag: chip antibody

value: rabbit IgG


title: CME180

source_name: Sox5 ChIP-seq

value: CD1 HA-tagged transgenic mice

value: anti-HA


title: CME182

source_name: Sox6 ChIP-seq


title: CME179

source_name: Sox5 ChIP-seq negative control


title: hpf12-57

description:
processed data file:
hpf12.txt
hpf12_p3_61_R1


title: hpf12-47

description:
processed data file:
hpf12.txt
hpf12_p3_52_R1


title: hpf12-46

description:
processed data file:
hpf12.txt
hpf12_p3_51_R1


title: hpf12-45

description:
processed data file:
hpf12.txt
hpf12_p3_50_R1


title: HEK293T AlkB demethylase-treated one-pot charging; replicate 3

city: Chicago

first: Tao

last: Pan

state: Illinois

address: 929 E. 57th Street

department: Biochemistry and Molecular Biology

institute: University of Chicago

data_processing:
Raw reads were processed using Trimmomatic v 0.32n and custom Python scripts to fully remove traces of adapter.
Processed reads were aligned in Bowtie 1.0 according to the parameters of -t -v 3 -m 10 --best --strata -p 12
Fragments shorter than 18 base pairs during alignment were removed during subsequent analysis.
Further processing using custom C and Python scripts were used to map reads onto tRNA isodecoders/rRNAs for visualization and modification index analysis.
Lastly, charging ratios were determined using strict read criteria to determine whether an aligned read was both A or C ending as well as aligned to the 3' end 20 nucleotides.
Genome_build: hg19
Supplementary_files_format_and_content: Individual alignment files mapping tRNA to isodecoder with read counts, as well as an aggregate spreadsheet containing map information and tRNA charging ratios

growth_protocol: Human embryonic kidney HEK293T (CRL-11268) cells from ATCC were grown in DMEM supplemented with 10% FBS, 1% 100× penicillin-streptomycin and passaged 7 times.

extract_protocol:
RNA was extracted once the cells reached 70-90% confluency at P7. RNA was isolated using Trizol, ensuring that the pH< 5 throughout the isolation to prevent hydrolysis of the aminoacyl bond. After ethanol precipitation, total RNA was resuspended in 10 mM NaOac/HOAc pH 4.8, 1 mM EDTA.Total RNA isolated  was mixed with prepared tRNA standards (final concentration = 0.03 pmol/µg total RNA). Periodate oxidation of total RNA (final concentration = 1 µg/µL) was carried out in 100 mM NaOAc/HOAc, pH 4.8, freshly prepared 50 mM NaIO4 at room temperature for 30 minutes. The reaction was quenched with 100 mM glucose for 5 minutes at room temperature. Any unquenched periodate was removed by Micro Bio-Spin P-6 Columns (Bio-Rad) and two ethanol precipitations. For β-elimination and deacylation, total RNA (final concentration = 1 µg/µL) was treated in 60 mM sodium borate buffer, pH 9.5 at 45°C for 90 min. RNA was purified by Micro Bio-Spin P-6 Columns before ethanol precipitation. To purify tRNA, total RNA was then run on an 8% denaturing polyacryimide gel (7M Urea, 1X TBE), and tRNA was excised and eluted from the gel in 50 mM KOAc, 200 mM KCl. After ethanol precipitation, short RNA oligo standards were added in different proportions (final concentration = 0.4 pmol total / µg tRNA) to ensure that TGIRT extension from T- and G-ending primers are equivalent. In the first biological replicate, the oligos were added in a ratio of 75% 3'-CCA, 25% 3'-CC. In the second biological replicate, oligos were added in equal ratios (50% 3'-CCA, 50% 3'-CC). In the third biological replicate, oligos were added in a ratio of 25% 3'-CCA, 75% 3'-CC. The tRNA and model oligo mixture was used as input for DM-tRNA-seq.
DM-tRNA-seq was carried out as previously described (Zheng 2015) with minor modifications. First, tRNA was demethylated under previously optimized conditions. 60 pmol of total tRNA was treated with 120 pmol WT AlkB and 240 pmol D135S AlkB in 25 mM MES, pH 5.0, 300 mM KCl, 2 mM MgCl2, 2 mM ascorbic acid, 300 µM α-ketoglutarate, 50 µM (NH4)2Fe(SO4)2 in the presence of RNAsin at room temperature for 2 hours. Zymo RNA Clean and Concentrator was used to remove the demethylases and clean up the reaction. Next, end repair (2'-3'-cyclic phosphate removal) was performed. 1 µM total tRNA was treated with T4 PNK (Affymetrix) (final concentration = 0.2 U/µL) at 37°C for 30 min. Zymo RNA Clean and Concentrator was used to clean up the reaction. For cDNA synthesis, TGIRT primer was 5' labeled with T4 PNK. 4 pmol of each 5' labeled TGIRT primer (T-ending: 5' GATCGTCGGACTGTAGAACTAGACGTGTGCTCTTCCGATCTTTCAGGCATTAGGCTCAAAGT, G-ending: 5' GATCGTCGGACTGTAGAACTAGACGTGTGCTCTTCCGATCTTTCAGGCATTAGGCTCAAAGG) was annealed to 8 pmol complementary RNA (5' CUUUGAGCCUAAUGCCUGAAAGAUCGGAAGAGCACACGUCUAGUUCUACAGUCCGACGAUC/3SpC3/) in 100 mM TrisHCl, pH 7.5, 0.5 mM EDTA at 82°C for 2min, then slow cooled to room temp. 4 pmol tRNA was then added. The tRNA/primer mixture (200 nM tRNA, 200 nM each primer) was pre-incubated at room temp for 30min in 100 mM TrisHCl, pH 7.5, 450 mM NaCl, 5 mM MgCl2, 5 mM DTT with 500 nM TGIRT (InGex, Inc.). dNTPs were added to a final concentration of 1 mM to initiate the reaction. The reverse transcription was performed at 60°C for 60min. The reactions were terminated with additions of NaOH to 0.25 M and incubation at 95°C for 3min. The reaction was neutralized with 0.25 M HCl. An equal volume of 50% Formamide, 4.5M Urea, 50mM EDTA, 0.05% Bromophenol blue, 0.05% xylene cyanol was added, and the mixture was heated at 95°C for 15 min. cDNAs were then purified by denaturing 10% PAGE (7M Urea, 1X TBE), and extended products were cut and eluted from the gel overnight in 50 mM KOAc, 200 mM KCl. Purified cDNA was ethanol precipitated with addition of linear acrylamide (Thermo) to 20 µg/mL. Purified cDNA was then circularized using CircLigase II (Epicentre) at 60°C overnight. After inactivation at 80°C for 10 min, samples were phenol/chloroform extracted and precipitated. PCR library preparation for Illumina sequencing was performed using Phusion Master Mix (Thermo) for 12 PCR cycles (98°C 5s, 60°C 10s, 72°C 10s). Samples were sequenced on Illumina HiSeq system.

tag: cell line

value: human embryonic kidney cells

tag: treated with

value: treatement with AlkB-derived enzyme mixture

tag: standard addition

value: RNA oligos added:25% CCA-ending,75% CC-ending

tag: molecule subtype

value: tRNA


title: hpf12-43

description:
processed data file:
hpf12.txt
hpf12_p3_49_R1

value: Wild type

value: Trunk ventral cells

value: Multipotent cardiopharyngeal progenitor cells


title: hpf12-44

description:
processed data file:
hpf12.txt
hpf12_p3_4_R1

value: Late tailbud I


title: hpf12-38

description:
processed data file:
hpf12.txt
hpf12_p3_44_R1


title: HEK293T AlkB demethylase-treated one-pot charging; replicate 2

value: RNA oligos added:50% CCA-ending, 50% CC-ending


title: hpf12-40

description:
processed data file:
hpf12.txt
hpf12_p3_46_R1


title: hpf12-41

description:
processed data file:
hpf12.txt
hpf12_p3_47_R1


title: hpf12-39

description:
processed data file:
hpf12.txt
hpf12_p3_45_R1


title: hpf12-36

description:
processed data file:
hpf12.txt
hpf12_p3_42_R1


title: HEK293T AlkB demethylase-treated one-pot charging; replicate 1

value: RNA oligos added:75% CCA-ending, 25% CC-ending


title: hpf12-35

description:
processed data file:
hpf12.txt
hpf12_p3_41_R1


title: hpf12-33

description:
processed data file:
hpf12.txt
hpf12_p3_3_R1


title: hpf12-32

description:
processed data file:
hpf12.txt
hpf12_p3_39_R1


title: hpf12-34

description:
processed data file:
hpf12.txt
hpf12_p3_40_R1


title: HEK293T AlkB demethylase-untreated one-pot charging; replicate 3

value: none (no treatment control)


title: CME019

source_name: Sox9 ChIP-seq IgG control

value: E13.5 embryos

value: goat IgG


title: HEK293T AlkB demethylase-untreated one-pot charging; replicate 2


title: hpf12-28

description:
processed data file:
hpf12.txt
hpf12_p3_35_R1


title: hpf12-30

description:
processed data file:
hpf12.txt
hpf12_p3_37_R1


title: CME018

source_name: Sox9 ChIP-seq

value: CD1 wildtype

value: Limbs and tails


title: hpf12-31

description:
processed data file:
hpf12.txt
hpf12_p3_38_R1


title: hpf12-29

description:
processed data file:
hpf12.txt
hpf12_p3_36_R1


title: hpf12-27

description:
processed data file:
hpf12.txt
hpf12_p3_34_R1


title: hpf12-26

description:
processed data file:
hpf12.txt
hpf12_p3_33_R1


title: hpf12-23

description:
processed data file:
hpf12.txt
hpf12_p3_30_R1


title: hpf12-25

description:
processed data file:
hpf12.txt
hpf12_p3_32_R1


title: hpf12-42

description:
processed data file:
hpf12.txt
hpf12_p3_48_R1


title: hpf12-24

description:
processed data file:
hpf12.txt
hpf12_p3_31_R1


title: hpf12-22

description:
processed data file:
hpf12.txt
hpf12_p3_2_R1


title: hpf12-37

description:
processed data file:
hpf12.txt
hpf12_p3_43_R1


title: HEK293T AlkB demethylase-untreated one-pot charging; replicate 1


title: hpf12-21

description:
processed data file:
hpf12.txt
hpf12_p3_29_R1


title: hpf12-20

description:
processed data file:
hpf12.txt
hpf12_p3_28_R1


title: Deproteinized DNA, genome-wide ATAC transposition background

city: oxford

first: Jim

last: Hughes

address: Weatherall Institute Of Molecular Me

department: MHU

country: United Kingdom

institute: University of Oxford

description: Genome-wide library of Tn5 transposed fragments

data_processing:
Alignment to the genome Bowtie(1.1.2); default with -m set to 2
Trim_galore(0.3.1) (to remove sequencing adaptors from unmapped reads)
Flashing reads using flash(1.2.8); default with -m 9 -x 0.125 (to combine short overlapping, unmapped reads)
Realignment of former unmapped reads using Bowtie as described before
Removal of PCR duplicates, using samtools(0.1.19)
Excluding regions highly affecting uniqueness and mappability using bedtools(2.17.0)
Merge bam files from same sample and same sequencing instrument model, keeping R1 and R2 separate.
Merge bam files from same sample for coverage estimation. Estimate coverage by aligned reads in a moving 300 bp window with a moving increment of 30 bp using custom perl script.
Genome_build: hg18
Supplementary_files_format_and_content: wig, window averaged read coverage

source_name: Erythroid cells

extract_protocol:
Human: Primary erythroid stem cell progenitors were isolated from peripheral blood, using CD34 coupled magnetic beads. Cells were expanded for 7days in low Epo conditions (0.5 IU/ml)) and then transfered for differentiation in high Epo (3.0 IU/ml) medium. On day 10, cells were washed in cold PBS, and counted.  70,000  cells were used for the ATAC-seq protocol. Nuclei were isolated by lysing the cells and transposition with Tn5 transposase, DNA extraction and sequencing was performed as published by Buenostro et al 2013.
ATAC-seq was performed as previously published (Buenrostro:2013). Nuclei of 70000 lysed cells were isolated and transposition with Tn5 transposase (Nextera, Illumina) was performed for 30 minutes at 37 °C. The DNA was extracted using a MinElute kit (Qiagen). Libraries were then amplified and barcoded using the NEBNext 2xMastermix (NEB) and the custom ATAC-seq primers published by Buenrostro and colleagues. ATAC-seq libraries profiles were visualized using D1000 tape on the Tapestation (Agilent) and quantified using the universal library quantification kit (KAPA Biosystems). For the ATAC-seq background 100 ng of genomic DNA was incubated with the Tn5 transposase and the library was made following the protocol described before. The strategy was to isolate open chromatin fragments specifically as they underly regulatory genomic regions. These are short stretches of DNA, with low abundance in the genome and thus require to be processed efficiently. The fragments can then be multiplexed and sequenced.

value: blood

tag: genes analysed

value: Genome-wide


title: hpf12-19

description:
processed data file:
hpf12.txt
hpf12_p3_27_R1

tag: genotype/variation


title: hpf12-18

description:
processed data file:
hpf12.txt
hpf12_p3_26_R1

tag: developmental stage


title: hpf12-17

description:
processed data file:
hpf12.txt
hpf12_p3_25_R1

tag: cell type


title: hpf12-16

description:
processed data file:
hpf12.txt
hpf12_p3_24_R1

tag: cell subtype


title: Primary erythroid cells, growth protocol 2_ATAC


title: hpf12-15

description:
processed data file:
hpf12.txt
hpf12_p3_23_R1


title: hpf12-14

description:
processed data file:
hpf12.txt
hpf12_p3_22_R1


title: hpf12-13

description:
processed data file:
hpf12.txt
hpf12_p3_21_R1


title: hpf12-12

description:
processed data file:
hpf12.txt
hpf12_p3_20_R1


title: Deproteinized DNA, genome-wide DNaseI digestion background (human)

description: Genome-wide library of DNase I digested fragments

data_processing:
Alignment to the genome Bowtie(1.1.2); default with -m set to 2
Trim_galore(0.3.1) (to remove sequencing adaptors from unmapped reads)
Flashing reads using flash(1.2.8); default with -m 9 -x 0.125 (to combine short overlapping, unmapped reads)
Realignment of formerunmapped reads using Bowtie as described before
Removal of PCR duplicates, using samtools(0.1.19)
Excluding regions highly affecting uniqueness and mappability using bedtools(2.17.0)
Merge bam files from same sample and same sequencing instrument model, keeping R1 and R2 separate.
Merge bam files from same sample for coverage estimation. Estimate coverage by aligned reads in a moving 300 bp window with a moving increment of 30 bp using custom perl script.
Genome_build: hg18/mm9
Supplementary_files_format_and_content: bigWig, window averaged read coverage

extract_protocol:
Mouse: A single cell suspension was made by gently dissociating the PHZ spleens and passing it through a 70um filter.  Cells were washed in cold PBS, and counted. 50million cells were used for the DNase-Seq protocol. Nuclei were isolated by lysing the cells and digested with increasing concentrations of DNaseI to isolate open chromatin fragments as previously published (Hosseini, 2014).
Human: Primary erythroid stem cell progenitors were isolated from peripheral blood, using CD34 coupled magnetic beads. Cells were expanded for 7days in low Epo conditions (0.5 IU/ml)) and then transfered for differentiation in high Epo (3.0 IU/ml) medium. On day 13, cells were washed in cold PBS, and counted. 50million cells were used for the DNase-Seq protocol. Nuclei were isolated by lysing the cells and digested with increasing concentrations of DNaseI to isolate open chromatin fragments as previously published (Hosseini, 2014, DOI 10.1371/journal.pgen.1003570).
The DNase-Seq protocol was perfromed as previously published (Hosseini 2014). DNA was purified using a phenol choroform extraction and the optimal disgests were selected for library preparation using NEB Library Preparation kit. Libraries were amplified and barcoded using the NEBNext 2xMastermix (NEB) and TruSeq oligos. A size selection step for small fragments was performed. DNase-Seq libraries profiles were visualized using D1000 tape on the Tapestation (Agilent). The libraries were quantified using the universal library quantification kit (KAPA Biosystems). Samples were sequenced on Illumina platform using: 150bp paired end reads (MiSeq), 75bp paired end reads (HiSeq) or 40/75 bp (NextSeq) paired end reads. For the background libraries, 100ng of genomic DNA was incubated with DNase for 3 minutes and then libraries were created to test sequence for sequence bias.
The strategy was to isolate open chromatin fragments specifically as they underly regulatory genomic regions. These are short stretches of DNA, with low abundance in the genome and thus require to be processed efficiently. The fragments can then be multiplexed and sequenced.

treatment_protocol: Mouse: Phenylhydrazine (PHZ) injections, 4 in the course of 6 days.


title: hpf12-11

description:
processed data file:
hpf12.txt
hpf12_p3_1_R1


title: hpf12-10

description:
processed data file:
hpf12.txt
hpf12_p3_19_R1


title: hpf12-9

description:
processed data file:
hpf12.txt
hpf12_p3_18_R1


title: Primary erythroid cells, biological replicate 2, growth protocol 2 (human)


title: hpf12-7

description:
processed data file:
hpf12.txt
hpf12_p3_16_R1


title: hpf12-8

description:
processed data file:
hpf12.txt
hpf12_p3_17_R1


title: hpf12-6

description:
processed data file:
hpf12.txt
hpf12_p3_15_R1


title: hpf12-5

description:
processed data file:
hpf12.txt
hpf12_p3_14_R1


title: hpf12-4

description:
processed data file:
hpf12.txt
hpf12_p3_13_R1


title: hpf12-3

description:
processed data file:
hpf12.txt
hpf12_p3_12_R1


title: hpf12-1

description:
processed data file:
hpf12.txt
hpf12_p3_10_R1


title: hpf12-2

description:
processed data file:
hpf12.txt
hpf12_p3_11_R1


title: Rpl10a siRNA_combined_heavy_RNA-seq_rep1

city: Stanford

first: Zhen

last: Shi

state: California

address: 279 Campus Drive

institute: Stanford

description: RNA (with rRNA depletion)

data_processing:
For Ribo-Seq, sequencing reads were parsed by cutadapt to remove the 3’ adapter sequence and reads with good sequencing qualities (ascii quality score>33) were kept. A layered alignment employing Bowtie 2 was performed to first discard reads mapping to rRNA, tRNA or snRNA sequences. Non-rRNA/tRNA/snRNA reads were then aligned against the canonical isoform of UCSC known gene transcripts (mm10). Mapped footprints were assigned to a specific position based on the A sites. The position of A site in relative to the 5’ end of each read is calculated as follows: 29-30 nt: +15; 31-33 nt: +16 and 34-35 nt: +17. For each gene, the total number of ribosome footprints mapping to the CDS excluding the first 15 or last 5 codons were counted. The density of ribosome footprints was calculated as Reads Per Kilobase per Million mapped reads (RPKM). Genes having ≥5 RPKM were kept for further analysis.
For polysome profiling, the paired-end reads were aligned to the mouse genome using the STAR RNA-Seq aligner (Dobin et al., 2013), and the total number of reads uniquely mapped to each genes were counted. We then calculated the amount of mRNAs in the combined medium and heavy polysome fractions compared to all other fractions as an estimate of their translation activities.
Genome_build: mm10
Supplementary_files_format_and_content: Text files include normalized read values for protein-coding genes

source_name: Embryonic stem cells (ESCs)

growth_protocol: E14 mouse embryonic stem cells (ESCs) were grown in knockout DMEM (Invitrogen 10829-018) with 15% ESC grade FBS (EMD Millipore ES-009-B), 1x L-glutamine (EMD Millipore TMS-002-C), 1x ES-grade Penicillin/streptomycin (EMD Millipore TMS-AB2-C), 1x non-essential amino acid (EMD Millipore TMS-001-C), 0.1% 2-mercaptoethanol, 1x Leukemia Inhibitory Factor (LIF, EMD Millipore ESG1107) in 37 °C, 5% CO2 incubator. 1 hr prior to harvest, cells were incubated with fresh mESC media and were at ~60-70% confluency.

extract_protocol:
The RNA-Seq libraries following gradient fractionation were prepared using the TruSeq Stranded mRNA Library Prep Kit (Illumina) by the Stanford Functional Genomics Facility and sequenced on a NextSeq sequencer (Illumina) with 2 x 75 bp read length.
The media was then aspirated and cells were scraped from the plate in cold PBS with 100 μg/ml cycloheximide. After centrifugation, the cell pellet was re-suspended in cold lysis buffer (20 mM Tris pH 7.5, 150 mM NaCl, 15 mM MgCl2, 100 μg/ml cycloheximide, 1 mM DTT, 0.5% Triton X-100, 8% glycerol, 20 U/ml TURBO™ DNase, 1x Combined Protease and Phosphatase Inhibitor (Thermo 78443)) and incubated for 30 min at 4°C with occasional vortex. The lysate was clarified by sequential centrifugation for 5 min at 1,800 x g and 10,000 x g at 4°C to remove nucleus and mitochondria. A 250 μl of lysate (with RNA concentration of ~ 1 μg/μl) was then treated with RNase A/T1 mix (0.5 mg RNase A (Life Technologies AM2272) and 1,000 U RNase T1 (Life Technologies 2280)) for 30 min at room temperature to digest mRNAs not protected by the ribosome. The digestion was stopped by adding 4.5 μl of SUPERase-In™ RNase Inhibitor (20 U/μl, Ambion AM2696). For the total Ribo-Seq, lysate was then loaded onto a 1 M sucrose cushion. Ribosomes were pelleted by centrifugation at 70, 000 rpm for 4 hr at 4°C. Pellet were re-suspended in TRIzol® for RNA extraction. For the RPL10A-Ribo-Seq and RPS25-Ribo-Seq, ribosomes containing RPS25 or RPL10A were enriched by IP using anti-FLAG® M2 magnetic beads (Sigma) on a head-over-tail rotator at 4 °C for 2 hrs. For RPL22-Ribo-Seq, ribosomes containing RPL22 were enriched by IP by incubating with anti-HA antibody (ab9110) on a head-over-tail rotator at 4 °C for 4 hrs and and then Dynabeads Protein A (Life Technologies, 10001D) at 4 °C for 1 hr. Beads were washed 5 min for 3 times in washing buffer (20 mM Tris pH 7.5, 200 mM NaCl, 15 mM MgCl2, 100 μg/ml cycloheximide, 0.5% Triton X-100). After the final wash, liquid was aspirated and immunoprecipitated ribosomes were eluted in TRIzol® for RNA extraction. Polysome profiling was performed comparing the control siRNA and Rpl10a knockdown. After polysome fractionation, RNA was extracted using Acid-Phenol:Chloroform, pH 4.5 (with IAA, 125:24:1) (Ambion, AM9722). We collected RNA from combined light fractions containing the free RNPs, 40S/60S ribosomal free subunits, 80S/monosome as well as light polysomes (2-3 ribosomes on a mRNA molecule), and combined heavy polysome fractions containing the most actively translating ribosomes (≥4 ribosomes along a mRNA molecule).

treatment_protocol: Gene knockdown was achieved by transfecting mESCs with small double-stranded interfering RNAs (siRNA) with Lipofectamine 2000 (Invitrogen, 11668). siRNA used for targeting Rpl10a: SASI_Mm01_00200342 (Sigma). MISSION siRNA Universal Negative Control 2 (Sigma, SIC002) was used as a control. Cells were briefly treated with 100 μg/ml cycloheximide (Sigma) in mESC media for 1 min prior to harvest.

value: E14 ESC

tag: genotype

value: Wild type mESCs treated with Rpl10a siRNA


title: Rpl10a siRNA_combined_light_RNA-seq_rep2

description: RNA (with rRNA depletion)


title: Rpl10a siRNA_combined_light_RNA-seq_rep1

description: RNA (with rRNA depletion)


title: Control siRNA_combined_heavy_RNA-seq_rep2

description: RNA (with rRNA depletion)

value: Wild type mESCs treated with control siRNA


title: Rpl10a siRNA_combined_heavy_RNA-seq_rep2

description: RNA (with rRNA depletion)


title: Control siRNA_combined_light_RNA-seq_rep2

description: RNA (with rRNA depletion)


title: Deproteinized DNA, genome-wide DNaseI digestion background (mouse)


title: RPL22-Ribo-Seq (RPL22-3xHA cells)_rep1

description: Ribosome footprints

data_processing:
Library strategy: Ribo-seq
For Ribo-Seq, sequencing reads were parsed by cutadapt to remove the 3’ adapter sequence and reads with good sequencing qualities (ascii quality score>33) were kept. A layered alignment employing Bowtie 2 was performed to first discard reads mapping to rRNA, tRNA or snRNA sequences. Non-rRNA/tRNA/snRNA reads were then aligned against the canonical isoform of UCSC known gene transcripts (mm10). Mapped footprints were assigned to a specific position based on the A sites. The position of A site in relative to the 5’ end of each read is calculated as follows: 29-30 nt: +15; 31-33 nt: +16 and 34-35 nt: +17. For each gene, the total number of ribosome footprints mapping to the CDS excluding the first 15 or last 5 codons were counted. The density of ribosome footprints was calculated as Reads Per Kilobase per Million mapped reads (RPKM). Genes having ≥5 RPKM were kept for further analysis.
For polysome profiling, the paired-end reads were aligned to the mouse genome using the STAR RNA-Seq aligner (Dobin et al., 2013), and the total number of reads uniquely mapped to each genes were counted. We then calculated the amount of mRNAs in the combined medium and heavy polysome fractions compared to all other fractions as an estimate of their translation activities.
Genome_build: mm10
Supplementary_files_format_and_content: Text files include normalized read values for protein-coding genes

extract_protocol:
All Ribo-Seq libraries were prepared as described in Ingolia, N. T., Brar, G. A., Rouskin, S., McGeachy, A. M. & Weissman, J. S. The ribosome profiling strategy for monitoring translation in vivo by deep sequencing of ribosome-protected mRNA fragments. Nat Protoc 7, 1534–1550 (2012).
The media was then aspirated and cells were scraped from the plate in cold PBS with 100 μg/ml cycloheximide. After centrifugation, the cell pellet was re-suspended in cold lysis buffer (20 mM Tris pH 7.5, 150 mM NaCl, 15 mM MgCl2, 100 μg/ml cycloheximide, 1 mM DTT, 0.5% Triton X-100, 8% glycerol, 20 U/ml TURBO™ DNase, 1x Combined Protease and Phosphatase Inhibitor (Thermo 78443)) and incubated for 30 min at 4°C with occasional vortex. The lysate was clarified by sequential centrifugation for 5 min at 1,800 x g and 10,000 x g at 4°C to remove nucleus and mitochondria. A 250 μl of lysate (with RNA concentration of ~ 1 μg/μl) was then treated with RNase A/T1 mix (0.5 mg RNase A (Life Technologies AM2272) and 1,000 U RNase T1 (Life Technologies 2280)) for 30 min at room temperature to digest mRNAs not protected by the ribosome. The digestion was stopped by adding 4.5 μl of SUPERase-In™ RNase Inhibitor (20 U/μl, Ambion AM2696). For the total Ribo-Seq, lysate was then loaded onto a 1 M sucrose cushion. Ribosomes were pelleted by centrifugation at 70, 000 rpm for 4 hr at 4°C. Pellet were re-suspended in TRIzol® for RNA extraction. For the RPL10A-Ribo-Seq and RPS25-Ribo-Seq, ribosomes containing RPS25 or RPL10A were enriched by IP using anti-FLAG® M2 magnetic beads (Sigma) on a head-over-tail rotator at 4 °C for 2 hrs. For RPL22-Ribo-Seq, ribosomes containing RPL22 were enriched by IP by incubating with anti-HA antibody (ab9110) on a head-over-tail rotator at 4 °C for 4 hrs and and then Dynabeads Protein A (Life Technologies, 10001D) at 4 °C for 1 hr. Beads were washed 5 min for 3 times in washing buffer (20 mM Tris pH 7.5, 200 mM NaCl, 15 mM MgCl2, 100 μg/ml cycloheximide, 0.5% Triton X-100). After the final wash, liquid was aspirated and immunoprecipitated ribosomes were eluted in TRIzol® for RNA extraction. Polysome profiling was performed comparing the control siRNA and Rpl10a knockdown. After polysome fractionation, RNA was extracted using Acid-Phenol:Chloroform, pH 4.5 (with IAA, 125:24:1) (Ambion, AM9722). We collected RNA from combined light fractions containing the free RNPs, 40S/60S ribosomal free subunits, 80S/monosome as well as light polysomes (2-3 ribosomes on a mRNA molecule), and combined heavy polysome fractions containing the most actively translating ribosomes (≥4 ribosomes along a mRNA molecule).

value: Bearing 1 allele of Rpl22-3xHA at endogeneous locus

tag: ip antibody

value: anti-HA antibody (Abcam , catalog # ab9110)


title: Total Ribo-Seq (RPL22-3xHA cells)_rep1

description: Ribosome footprints


title: RPL22-Ribo-Seq (RPL22-3xHA cells)_rep2

description: Ribosome footprints


title: A128T-IN-mock

city: Saint Louis

first: Sebla

middle: Bulent

last: Kutluay

state: Missouri

address: Washington University School of Medicine

description: CLIP-seq with NC

data_processing:
CLIP-seq reads were trimmed off of the adapters and collapsed using FASTX-toolkit
Resulting reads were mapped to the viral genome using bowtie
Samtools mpileup and custom scripts were used to generate read density (i.e. number of reads that map to each nucleotide of the viral genome) graphs on viral RNAs.
Genome_build: HIV-1 NL4-3 strain
Supplementary_files_format_and_content: tab-delimited files showing read density on each nucleotide of the viral genome as well as nucleotide substitutions.

organism: Human immunodeficiency virus 1

source_name: HIV-1 NL4-3 virions

growth_protocol: Cells transfected with HIV-1 NL4-3-based proviral plasmids were utilized for harvesting virus particles.

extract_protocol:
Virions collected from cells were pelleted on sucrose cushions, lysed in detergent containing buffers and IN-RNA complexes were immunoprecipitated from these lysates.
Libraries were constructed based on previously published methodologies as explained before (Kutluay et al,m 2014, Cell). The 3' adapter is common to all samples and has the sequence: TCG TAT GCC GTC TTC TGC TTG. The 5' adapters are:Sample 1 (rGUU CAG AGU UCU ACA GUC CGA CGA UC GAU NNN UC); Sample 2 ( rGUU CAG AGU UCU ACA GUC CGA CGA UC GUG NNN UC); Sample 3 ( rGUU CAG AGU UCU ACA GUC CGA CGA UC UAG NNN UC); Sample 4 (rGUU CAG AGU UCU ACA GUC CGA CGA UC AUC NNN UC).

treatment_protocol: Prior to isolation of HIV-1 particles, cells were treated with 4-thiouridine to enhance UV crosslinking. Some samples were treated with allosteric integrase inhibitor BI-B2.

tag: antibody

value: monoclonal anti-NC


title: A128T-IN(+)BI-B2

institute: Washington University School of Medicine

description: CLIP-seq with NC


title: WT(+)BI-B2

description: CLIP-seq with NC


title: Primary erythroid cells, biological replicate 1, growth protocol 1 (human)

tag: tissue


title: Total Ribo-Seq (Rps25-3xFLAG cells)_rep1

description: Ribosome footprints

data_processing:
library strategy: Ribo_seq
Sequencing reads were parsed by cutadapt to remove the 3’ adapter sequence and reads with good sequencing qualities (ascii quality score>33) were kept.
A layered alignment employing Bowtie 2 was performed to first discard reads mapping to rRNA, tRNA or snRNA sequences. Non-rRNA/tRNA/snRNA reads were then aligned against the canonical isoform of UCSC known gene transcripts (mm10).
Mapped footprints were assigned to a specific position based on the A sites. The position of A site in relative to the 5’ end of each read is calculated as follows: 29-30 nt: +15; 31-33 nt: +16 and 34-35 nt: +17.
For each gene, the total number of ribosome footprints mapping to the CDS excluding the first 15 or last 5 codons were counted. The density of ribosome footprints was calculated as Reads Per Kilobase per Million mapped reads (RPKM). Genes having ≥5 RPKM were kept for further analysis.
Genome_build: mm10
Supplementary_files_format_and_content: Text file includes UCSC_IDs and RPKM values for each protein-coding gene

extract_protocol:
The media was then aspirated and cells were scraped from the plate in cold PBS with 100 μg/ml cycloheximide. After centrifugation, the cell pellet was re-suspended in cold lysis buffer (20 mM Tris pH 7.5, 150 mM NaCl, 15 mM MgCl2, 100 μg/ml cycloheximide, 1 mM DTT, 0.5% Triton X-100, 0.1 mg/ml heparin, 8% glycerol, 20 U/ml TURBO™ DNase, 1x Combined Protease and Phosphatase Inhibitor (Thermo 78443)) and incubated for 30 min at 4°C with occasional vortex. The lysate was clarified by sequential centrifugation for 5 min at 1,800 x g and 10,000 x g at 4°C to remove nucleus and mitochondria. A 250 μl of lysate (with RNA concentration of ~ 1 μg/μl) was then treated with RNase A/T1 mix (0.5 mg RNase A (Life Technologies AM2272) and 1,000 U RNase T1 (Life Technologies 2280)) for 30 min at room temperature to digest mRNAs not protected by the ribosome. The digestion was stopped by adding 4.5 μl of SUPERase-In™ RNase Inhibitor (20 U/μl, Ambion AM2696). For the total Ribo-Seq, lysate was then loaded onto a 1 M sucrose cushion. Ribosomes were pelleted by centrifugation at 70, 000 rpm for 4 hr at 4°C. After a brief wash, pellet were re-suspended in TRIzol® for RNA extraction. For the RPL10A-Ribo-Seq and RPS25-Ribo-Seq, ribosomes containing RPS25 or RPL10A were enriched by IP using anti-FLAG® M2 magnetic beads (Sigma) on a head-over-tail rotator at 4 °C for 2 hrs. Beads were washed 5 min for 3 times in washing buffer (20 mM Tris pH 7.5, 200 mM NaCl, 15 mM MgCl2, 100 μg/ml cycloheximide, 0.5% Triton X-100). After the final wash, liquid was aspirated and immunoprecipitated ribosomes were eluted in TRIzol® for RNA extraction.
All Ribo-Seq libraries were prepared as described in Ingolia, N. T., Brar, G. A., Rouskin, S., McGeachy, A. M. & Weissman, J. S. The ribosome profiling strategy for monitoring translation in vivo by deep sequencing of ribosome-protected mRNA fragments. Nat Protoc 7, 1534–1550 (2012)

treatment_protocol: Cells were briefly treated with 100 μg/ml cycloheximide (Sigma) in mESC media for 1 min prior to harvest.

value: Bearing 1 allele of Rps25-3xFLAG at endogeneous locus


title: Total Ribo-Seq (Rps25-3xFLAG cells)_rep2

description: Ribosome footprints


title: RPL10A-Ribo-Seq (3xFLAG-Rpl10a cells)_rep1

description: Ribosome footprints

value: Bearing 1 allele of 3xFLAG-Rpl10a at endogeneous locus

value: anti-FLAG M2 magnetic beads (Sigma, Catalog# M8823, Lot# SLBL1128V)


title: Liver

city: Guangzhou

first: Jing

middle: E

last: Ma

state: Guangdong

address: xingangxi

institute: Guangdong Institute of Applied Biological Resources

data_processing:
Primary sequencing data (raw reads) from the Illumina HiSeqTM 2000 were subjected to quality control (QC) through in-house perl scripts to determine whether a resequencing step was needed. Followed as: a) remove reads with adaptors, b) remove reads in which unknown bases (N) are more than 5% ( Parameters:-n 0.05), c) remove low quality reads (we define the low quality read as the percentage of base which quality is lesser than 10 is greater than 20% in a read) (Parameters:-l 10 -q 0.2).
The FPKM (Fragments Per Kilobase of transcript per Million fragments mapped) value of the transcripts from the RNA-Seq results, which had been calculated by the followed computational formula, FPKM=(cDNA Fragments)/(Mapped Fragments (Millions)  × Transcript Length (kb) ). Note: “cDNA Fragments” represents the number of fragments on a transcript, “Mapped Fragments (Millions)” represents the total number of fragments aligned to the referred genome in the unit of 106 bp, “Transcript Length (kb)” represents the length of a transcript in the unit of 103 bp.
Genome_build: ManJav1.0
Supplementary_files_format_and_content: FPKMs

organism: Manis javanica

source_name: Liver

extract_protocol:
Total RNA of the three tissues from three samples was extracted from tissues with RNAiso reagent (Takara, Otsu, Japan) and was treated with DNase I (Takara).
RNA libraries were prepared for sequencing using standard Illumina protocols

value: Adult

tag: gender

value: female

value: Liver


title: Small intestine

source_name: Small intestine

value: Intestine


title: RPL10A-Ribo-Seq (3xFLAG-Rpl10a cells)_rep2

description: Ribosome footprints


title: RPS25-Ribo-Seq (Rps25-3xFLAG cells)_rep2

description: Ribosome footprints


title: Salivary glands

source_name: Salivary glands

value: Salivary glands


title: RPS25-Ribo-Seq (Rps25-3xFLAG cells)_rep1

description: Ribosome footprints


title: Control siRNA_combined_heavy_RNA-seq_rep1

description: RNA (with rRNA depletion)


title: Total Ribo-Seq (3xFLAG-Rpl10a cells)_rep2

description: Ribosome footprints


title: Control siRNA_combined_light_RNA-seq_rep1

description: RNA (with rRNA depletion)


title: Total Ribo-Seq (RPL22-3xHA cells)_rep2

description: Ribosome footprints


title: Mouse erytrhoid cells from knockout of CTCF binding site -HS38 and -HS39 replicate 2

status: Public on Jun 14 2017

city: Oxford

first: Jelena

middle: M

last: Telenius

address: MRC Weatherall Institute of Molecular Medicine University of Oxford John Radcliffe Hospital Headington

department: Weatherall Institute of Molecular Medicine (WIMM)

institute: Oxford University

description: globin-depleted polyA+ RNA

data_processing:
Trim_galore (to remove sequencing adaptors)
FLASH (to reconstruct paired end reads into single reads where possible)
Alignment of RNA-seq data with STAR to mm9, using the following parameters: --outFilterType BySJout --outFilterMultimapNmax 20 --alignSJoverhangMin 8 --alignSJDBoverhangMin 1 --outFilterMismatchNmax 999
Deeptools bamcoverage was used (options: --filterRNAstrand, --normalizeUsingRPKM) to visualise total and strand-specific RNA-seq aligments in UCSC genome browser
Genome_build: mm9
Supplementary_files_format_and_content: wig of RPKM

growth_protocol: C57BL/6 mice were treated with phenylhydrazine (three doses of 40 mg/g body weight given 12 hours apart; mice were sacrificed after five days)

extract_protocol:
A single cell suspension was made by gently dissociating the spleen of a phenyhydrazine treated mouse and passing it through a 70um filter. Ter119 selection was performed by staining the cells with phycoerythrin conjugated anti-ter119 antibodies (BD biosciences). The cells were subsequently washed and then incubated with MACS anti-phycoerythrin beads. Cells were then separated using an auto-MACS (Miltenyi Biotec).
Globin-zero Gold kit (Illumina) followed by NEBNext poly(A) isolation module and NEBnext ultra library prep kit

treatment_protocol: Ter119+ cells were isolated from mouse spleens

value: Phenylhydrazine treated spleen

value: genome-wide


title: Mouse erytrhoid cells from knockout of CTCF binding site -HS38 and -HS39 replicate 3

description: globin-depleted polyA+ RNA


title: Mouse wild type erythroid cells replicate 1

description: globin-depleted polyA+ RNA


title: Mouse D39 mutant erythroid cells H3K4me3 ChIP biological replicate 2

data_processing:
Trim_galore (to remove sequencing adaptors)
FLASH (to reconstruct paired end reads into single reads where possible, options: -m 9 –x 0.125)
Alignment of ChIP-seq data with bowtie 1.1.2 to mm9 (options: -m 2, -k 1, --best)
Samtools rmdup (to remove PCR duplicates)
Deeptools bamcoverage was used (options: --normalizeUsingRPKM) to normalise the reads for RPKM, and generate the bigWig file for visualisation
Genome_build: mm9
Supplementary_files_format_and_content: bigWig of RPKM

source_name: Ter119+ Erythroid cells

extract_protocol:
A single cell suspension was made by gently dissociating the spleen of a phenyhydrazine treated mouse and passing it through a 70um filter. Ter119 selection was performed by staining the cells with phycoerythrin conjugated anti-ter119 antibodies (BD biosciences). The cells were subsequently washed and then incubated with MACS anti-phycoerythrin beads. Cells were then separated using an auto-MACS (Miltenyi Biotec).
NEBnext ultra library prep kit

value: anti_H3K4me3, Abcam ChIP Grade (ab8580)


title: Mouse D39 mutant erythroid cells H3K4me3 ChIP biological replicate 1


title: Mouse D39 mutant erythroid cells CTCF ChIP biological replicate 1

value: anti-CTCF, Millipore 07-729


title: Total Ribo-Seq (3xFLAG-Rpl10a cells)_rep1

description: Ribosome footprints

value: E14 ESC


title: Mouse wild type erythroid cells replicate 2

description: globin-depleted polyA+ RNA


title: Mouse erytrhoid cells from knockout of CTCF binding site -HS29 replicate 3

description: globin-depleted polyA+ RNA


title: Mouse erytrhoid cells from knockout of CTCF binding site -HS38 and -HS39 replicate 1

description: globin-depleted polyA+ RNA


title: Mouse erytrhoid cells from knockout of CTCF binding site -HS29 replicate 1

description: globin-depleted polyA+ RNA


title: Mouse erytrhoid cells from knockout of CTCF binding site -HS29 replicate 2

description: globin-depleted polyA+ RNA


title: WT-mock

description: CLIP-seq with NC


title: Mouse D39 mutant erythroid cells CTCF ChIP biological replicate 2


title: Mouse D38 mutant erythroid cells H3K4me3 ChIP biological replicate 2


title: Mouse D3839 mutant erythroid cells H3K4me3 ChIP biological replicate 1


title: Mouse D3839 mutant erythroid cells CTCF ChIP biological replicate 2


title: Mouse D3839 mutant erythroid cells H3K4me3 ChIP biological replicate 2


title: Mouse D3839 mutant erythroid cells H3K27me3 ChIP biological replicate 2

value: anti-H3K27me3 (C36B11), Cell signalling #9733


title: Mouse D38 mutant erythroid cells CTCF ChIP biological replicate 2


title: Mouse wild type erythroid cells replicate 3

description: globin-depleted polyA+ RNA


title: Mouse D3839 mutant erythroid cells PolII ChIP biological replicate 1

value: anti-RNA PolII, Santa Cruz sc-56767


title: Mouse D38 mutant erythroid cells CTCF ChIP biological replicate 1


title: Mouse D29 mutant erythroid cells H3K4me3 ChIP biological replicate 2


title: Mouse D3839 mutant erythroid cells CTCF ChIP biological replicate 1


title: Mouse D3839 mutant erythroid cells H3K27me3 ChIP biological replicate 1


title: Mouse D38 mutant erythroid cells H3K4me3 ChIP biological replicate 1


title: Mouse D3839 mutant erythroid cells Ezh2 ChIP biological replicate 2

value: anti-Ezh2 (D2C9), Cell signalling #5246


title: Mouse D3839 mutant erythroid cells Ezh2 ChIP biological replicate 1


title: Mouse D29 mutant erythroid cells CTCF ChIP biological replicate 2


title: Mouse D29 mutant erythroid cells H3K4me3 ChIP biological replicate 1


title: Mouse D29 mutant erythroid cells CTCF ChIP biological replicate 1


title: Mouse wild type erythroid cells RNA Polymerase II ChIP biological replicate 2


title: Mouse wild type erythroid cells H3K4me3 ChIP biological replicate 2


title: Mouse wild type erythroid cells H3K27me3 ChIP biological replicate 1


title: Mouse wild type erythroid cells Ezh2 ChIP biological replicate 1


title: Mouse wild type erythroid cells CTCF ChIP biological replicate 1


title: Mouse wild type erythroid cells Ezh2 ChIP biological replicate 2


title: Mouse wild type erythroid cells CTCF ChIP biological replicate 2


title: Mouse D39 mutant erythroid cells ATAC-seq biological replicate 1

extract_protocol:
A single cell suspension was made by gently dissociating the spleen of a phenyhydrazine treated mouse and passing it through a 70um filter. Ter119 selection was performed by staining the cells with phycoerythrin conjugated anti-ter119 antibodies (BD biosciences). The cells were subsequently washed and then incubated with MACS anti-phycoerythrin beads. Cells were then separated using an auto-MACS (Miltenyi Biotec).
Illumina Nextera DNA library kit, subjected to ATAC-seq (Assay for Transposase-Accessible Chromatin)


title: Mouse D3839 mutant erythroid cells PolII ChIP biological replicate 2


title: Mouse D39 mutant erythroid cells ATAC-seq biological replicate 2


title: Mouse D3839 mutant erythroid cells ATAC-seq biological replicate 2


title: Mouse D3839 mutant erythroid cells ATAC-seq biological replicate 1


title: Mouse D29 mutant erythroid cells ATAC-seq biological replicate 1


title: ChIP-seq, BAT NCoR, 29C/5pm

city: Cincinnati

first: Hee-Woong

last: Lim

state: Ohio

address: 3333 Burnet Ave. MLC 7024

department: Division of Biomedical Informatics

institute: Cincinnati Children's Hospital Medical Center

data_processing:
ChIP-seq: All ChIP-seq libraries were sequenced on HiSeq 2500 (single-end 50bp or 100bp) or NextSeq 500 (single-end 75). Before alignment, all the reads were adjusted to 50bp by trimming 3’-end to avoid any bias due to different read lengths. Sequencing reads were aligned to the UCSC mm9 using Bowtie. For HDAC3 ChIP-seq data, each of WT and HDAC3-KO aligned reads were downsampled to 20 million reads to adjust read-depth bias and avoid peak saturation, and any duplicate reads were removed except for one. All the bigwig files for ChIP-seq data were generated using Homer and bedGraphToBigWig command. Two biological replicates of H3K27ac ChIP-seq were pooled to make single bigwig file in each condition.
GRO-seq: GRO-seq libraries were sequenced twice for better coverage, and sequencing replicates were pooled for each condition. Before alignment, sequencing reads were cleaned by trimming off low quality base, adapter and tailing poly-A sequences using cutadapt16. Trimmed reads of at least 25bp were selected and aligned to UCSC mm9 using bowtie with an option ‘--best --strata -m 1 -v 3’. Aligned reads within extremely high signal regions such as rRNA, snoRNA, snRNA and tRNA were removed before downstream analysis to minimize read-depth bias. To visualize transcriptional activity in genome browser, bigwig files were generated for (+) and (−) strands separately, where all the reads were extended to 150bp for smooth profile and (−) strand signal were presented as negative values.
Genome_build: mm9
Supplementary_files_format_and_content: Read-per-million (RPM) normalized stack-height profile in bigWig format

source_name: Brown adipose tissue

growth_protocol: Male wild-type and HDAC3 KO mice maintained on a C57BL/6 background maintained on a 12 hour dark /light cycle and were sacrificed at 5PM (ZT10) at 12 weeks of age for the collection of interscapular brown adipose tissue.

extract_protocol:
ChIP-seq: Interscapular BAT was isolated and processed separately for each sacrificed mouse. The tissues were immediately minced, crosslinked in 1 % formaldehyde/PBS, and stored at -80 C. For each fat pad, material from 3-4 mice was used for sequencing. To prepare ChIP extracts, frozen tissue was thawed, immediately crosslinked, and suspended ChIP buffer (50 mM HEPES/NaOH pH 7.5, 140 mM NaCl, 1 mM EDTA, 1% Triton X-100, 0.1% Na-Deoxycholate and Complete protease inhibitor) supplemented with 0.1% SDS. Samples were incubated at 4 C for 10 min, and subjected to microtip probe sonication under conditions optimized for IP efficiency. Extracts were cleared of insoluble lipid by microfuge centrifugation at full speed followed by extract transfer to a new tube. The process was repeated until minimal lipid accumulated at the top of the extract after centrifugation. GRO-seq:The nuclear run-on assay was performed as previously described (Core et al., 2008; Step et al., 2014; Wang et al., 2011). Nuclei extraction: Mouse tissues were harvested at Zeitgeber Time 10 and washed with cold swelling buffer (10 mM Tris pH 7.5, 2 mM MgCl2, 3 mM CaCl2, 2 U/ml Superase-In). Nuclear extract was prepared by dounce homogenization in cold swelling buffer and filtered using the cell strainer (100 µm, BD Biosciences). Lysate was centrifuged at 400g for 10 min, then resuspended in lysis buffer (swelling buffer with 10% glycerol and 1% Igepal) and incubated on ice for 5 min. Nuclei were washed twice with lysis buffer, and resuspended in freezing buffer (50 mM Tris pH 8.3, 40% glycerol, 5 mM MgCl2, 0.1 mM EDTA). Nuclear Run-On (NRO): Nuclei were mixed with an equal volume of run-on buffer (10 mM Tris pH 8.0, 5 mM MgCl2, 1 mM DTT, 300 mM KCl, 200 U/ml Superase-In, 1% Sarkosyl, 500 μM ATP, GTP and Br-UTP, 2 μM CTP) and incubated for 5 min at 30°C. Nuclear RNA was extracted with Trizol (Invitrogen) and treated with DNase (Ambion). RNA was hydrolyzed using fragmentation reagents (Ambion) for 13 min at 70°C and purified through a Micro Bio-Spin Column P-30 (Bio-Rad). RNA was treated with T4 polynucleotide kinase (New England Biolabs) for 1h at 37°C, then with an additional T4 polynucleotide kinase for 1h more, and inactivated for 5 min at 75°C. NRO-RNA immuno-precipitation: Anti-BrdU agarose beads (Santa Cruz) were rotated for 1h in blocking buffer (0.25x SSPE, 38 mM NaCl, 1 mM EDTA, 0.05% Tween-20, 0.1% PVP, and 0.1% BSA). Then NRO-RNA was rotated with beads for 1h, followed by washes twice in binding buffer (0.25x SSPE, 38 mM NaCl, 1 mM EDTA, 0.05% Tween-20), twice in low salt buffer (0.2x SSPE, 1 mM EDTA, 0.05% Tween-20), once in high salt buffer (0.2x SSPE, 1 mM EDTA, 0.05% Tween-20, 138 mM NaCl), and twice in TET buffer (TE pH 7.4, 0.05% Tween-20). BrdU-labeled RNA was eluted from the beads four times for 10 min with 100 μl elution buffer pre-heated to 42°C. RNA was precipitated by adding 300 mM NaCl, 1 µl of glycogen and 2.5 volumes of ethanol and incubated overnight at -20°C. RNA was resuspended in water (with 1 U/µl Superase-In and 0.05% Tween-20), denatured for 3 min at 65°C, and treated with poly(A)-polymerase (New England Biolabs) for 30 min at 37°C.
ChIP-seq: ChIP-Seq libraries were produced and sequenced according to Illumina protocols. Libraries were multiplexed for sequencing.GRO-seq: Synthesis of cDNA from RNA was performed as described previously (Wang et al., 2011) using oNTI223 primer (5′-/5Phos/GA TCG TCG GAC TGT AGA ACT CT/idSp/CAA GCA GAA GAC GGC ATA CGA TTT TTT TTT TTT TTT TTT TTV N-3′) where the “5Phos” indicates 5′ phosphorylation, “idSp” indicates the abasic dSpacer furan, and “VN” indicates degenerate nucleotides. The reaction was treated with exonuclease I (Fermentas) for 15 min at 37°C, followed by 100 mM NaOH for 20 min at 98°C, and neutralized with 100 mM HCl. cDNA was denatured at 70°C for 3 min, and run on 10% TBE-urea gel, then products from 105-400 nucleotides size were excised and eluted from shredded gel pieces for 4h in TE + 0.1% Tween and precipitated by adding 1 µl of glycogen, 300 mM NaCl and 2 volumes of ethanol and incubated overnight at -20°C. cDNA was resuspended into circularization reaction and circularized with CircLigase (Epicentre) for 1 hour at 60°C, denatured for 10 min at 80°C, and relinearized by adding relinearization mix (25 mM KCl, 500 µM DTT) and APE I (15U; New England Biolabs). Linearized DNA was amplified by PCR using Phusion Hot Start II Kit, according to manufacturer’s instructions. The oligonucleotide primers oNTI200 (5′-CAA GCA GAA GAC GGC ATA-3′) and oNTI201 (5′-AAT GAT ACG GCG ACC ACC GAC AGG TTC AGA GTT CTA CAG TCC GAC G-3′) were used for amplification. The PCR product was run on a 10% TBE gel and products from 150-305 nucleotides size were eluted from shredded gel pieces for 4h in TE + 0.1% Tween + 150 mM NaCl and purified using ChIP DNA Clean and Concentrator Kit (Genesee Scientific). Libraries were sequenced on an Illumina HiSeq2000 with sequencing primer 5′-CGA CAG GTT CAG AGT TCT ACA GTC CGA CGA TC-3′.

treatment_protocol: Male mice for thermoneutrality (29C) cohorts were allowed to acclimate for 2 weeks prior to tissue harvest. Mice were acclimated to 29C in environmental rodent incubator maintained on the same 12:12 hour dark/light cycle. Room temperature (22C) cohorts were maintained in standard housing room.

value: WT

tag: time

tag: temperature

value: Brown adipose tissue

value: Anti-NCoR [Custom rabbit antibody, GST fusion of NCoR C-term (a.a.1944-2453), Affinity purified]


title: ChIP-seq, BAT H3K27ac, Adiponectin-Cre HDAC3 floxed, 29C/5pm, replicate 2

description: biological replicates were pooled to make single bigwig file (linked to GEO Series entry GSE83927)

data_processing:
ChIP-seq: All ChIP-seq libraries were sequenced on HiSeq 2500 (single-end 50bp or 100bp) or NextSeq 500 (single-end 75). Before alignment, all the reads were adjusted to 50bp by trimming 3’-end to avoid any bias due to different read lengths. Sequencing reads were aligned to the UCSC mm9 using Bowtie. For HDAC3 ChIP-seq data, each of WT and HDAC3-KO aligned reads were downsampled to 20 million reads to adjust read-depth bias and avoid peak saturation, and any duplicate reads were removed except for one. All the bigwig files for ChIP-seq data were generated using Homer and bedGraphToBigWig command. Two biological replicates of H3K27ac ChIP-seq were pooled to make single bigwig file in each condition.
Genome_build: mm9
Supplementary_files_format_and_content: Read-per-million (RPM) normalized stack-height profile in bigWig format

extract_protocol:
ChIP-seq: Interscapular BAT was isolated and processed separately for each sacrificed mouse. The tissues were immediately minced, crosslinked in 1 % formaldehyde/PBS, and stored at -80 C. For each fat pad, material from 3-4 mice was used for sequencing. To prepare ChIP extracts, frozen tissue was thawed, immediately crosslinked, and suspended ChIP buffer (50 mM HEPES/NaOH pH 7.5, 140 mM NaCl, 1 mM EDTA, 1% Triton X-100, 0.1% Na-Deoxycholate and Complete protease inhibitor) supplemented with 0.1% SDS. Samples were incubated at 4 C for 10 min, and subjected to microtip probe sonication under conditions optimized for IP efficiency. Extracts were cleared of insoluble lipid by microfuge centrifugation at full speed followed by extract transfer to a new tube. The process was repeated until minimal lipid accumulated at the top of the extract after centrifugation. GRO-seq:The nuclear run-on assay was performed as previously described (Core et al., 2008; Step et al., 2014; Wang et al., 2011). Nuclei extraction: Mouse tissues were harvested at Zeitgeber Time 10 and washed with cold swelling buffer (10 mM Tris pH 7.5, 2 mM MgCl2, 3 mM CaCl2, 2 U/ml Superase-In). Nuclear extract was prepared by dounce homogenization in cold swelling buffer and filtered using the cell strainer (100 µm, BD Biosciences). Lysate was centrifuged at 400g for 10 min, then resuspended in lysis buffer (swelling buffer with 10% glycerol and 1% Igepal) and incubated on ice for 5 min. Nuclei were washed twice with lysis buffer, and resuspended in freezing buffer (50 mM Tris pH 8.3, 40% glycerol, 5 mM MgCl2, 0.1 mM EDTA).
ChIP-seq: ChIP-Seq libraries were produced and sequenced according to Illumina protocols. Libraries were multiplexed for sequencing.GRO-seq: Synthesis of cDNA from RNA was performed as described previously (Wang et al., 2011) using oNTI223 primer (5′-/5Phos/GA TCG TCG GAC TGT AGA ACT CT/idSp/CAA GCA GAA GAC GGC ATA CGA TTT TTT TTT TTT TTT TTT TTV N-3′) where the “5Phos” indicates 5′ phosphorylation, “idSp” indicates the abasic dSpacer furan, and “VN” indicates degenerate nucleotides. The reaction was treated with exonuclease I (Fermentas) for 15 min at 37°C, followed by 100 mM NaOH for 20 min at 98°C, and neutralized with 100 mM HCl. cDNA was denatured at 70°C for 3 min, and run on 10% TBE-urea gel, then products from 105-400 nucleotides size were excised and eluted from shredded gel pieces for 4h in TE + 0.1% Tween and precipitated by adding 1 µl of glycogen, 300 mM NaCl and 2 volumes of ethanol and incubated overnight at -20°C. cDNA was resuspended into circularization reaction and circularized with CircLigase (Epicentre) for 1 hour at 60°C, denatured for 10 min at 80°C, and relinearized by adding relinearization mix (25 mM KCl, 500 µM DTT) and APE I (15U; New England Biolabs). Linearized DNA was amplified by PCR using Phusion Hot Start II Kit, according to manufacturer’s instructions. The oligonucleotide primers oNTI200 (5′-CAA GCA GAA GAC GGC ATA-3′) and oNTI201 (5′-AAT GAT ACG GCG ACC ACC GAC AGG TTC AGA GTT CTA CAG TCC GAC G-3′) were used for amplification. The PCR product was run on a 10% TBE gel and products from 150-305 nucleotides size were eluted from shredded gel pieces for 4h in TE + 0.1% Tween + 150 mM NaCl and purified using ChIP DNA Clean and Concentrator Kit (Genesee Scientific). Libraries were sequenced on an Illumina HiSeq2000 with sequencing primer 5′-CGA CAG GTT CAG AGT TCT ACA GTC CGA CGA TC-3′.

tag: strain

value: Adiponectin-Cre, HDAC3 floxed

tag: chip antibody


title: ChIP-seq, BAT ERRa, Adiponectin-Cre HDAC3 floxed, 29C/5pm

value: Anti-ERRalpha [ABCAM, Cat# 16363, Lot# GR263385-1]


title: Exo NP rep2

city: Brisbane

first: Dominic

last: Guanzon

state: Queensland

address: Building 71/918 RBWH Herston

department: UQCCR

country: Australia

institute: University of Queensland

data_processing:
Remove index and adaptor sequences using TagCleaner (version 0.16)
Trim reads to 28 nucleotides using FASTX-Toolkit (version 0.0.13)
miRNAs were identified using the miRDeep2 program (version 2.0.0.7), using the deafult parameters with the exception of allowing for 1 mismatch and discarding read less than 16 nucleotides in length.
Raw miRNA counts were normalised and statistically anlaysed using the DESeq2 package (version 1.10.1) in R (version 3.2.2)
Genome_build: hg19
Supplementary_files_format_and_content: csv file containing raw miRNA counts

source_name: Plasma

growth_protocol: Trophoblast cultured at 8% or 1% oxygen in phenol red-free RPMI 1640 medium supplemented with 10% heat-inactivated fetal bovine serum, 1% non-essential amino acids, 1 mM sodium pyruvate, 100 U/mL penicillin, and 100 mg/mL streptomycin. Cultures were maintained at 37 degrees.

extract_protocol:
Exosomal RNA was extracted using the RNeasy Mini Kit 50 and TRIzol LS Reagent.
Libraries were constructed using the TruSeq® SmallRNA Library Prep Kit, following the manufacturer’s instructions. A total of 1 µg RNA or 100 to 300 ng of exosomal RNA was used as input for library construction.

tag: cell compartment

value: Exosome

value: plasma

tag: diagnosis

value: normal pregnancy


title: ChIP-seq, BAT H3K27ac, HDAC3 floxed, 29C/5pm, replicate 1

value: HDAC3 floxed


title: Mouse wild type erythroid cells RNA Polymerase II ChIP biological replicate 1

value: Phenylhydrazine treated spleen


title: Exo SPTB rep1

value: pre-term birth


title: ChIP-seq, BAT H3K27ac, HDAC3 floxed, 29C/5pm, replicate 2


title: Mouse wild type erythroid cells H3K27me3 ChIP biological replicate 2


title: Exo PE rep1

value: pre-eclampsia


title: Exo SPTB rep2


title: Exo NP rep3


title: ChIP-seq, BAT ERRa, HDAC3 floxed, 29C/5pm


title: Exo 1% O2 rep1

source_name: Trophoblast

value: extravillous trophoblast


title: upper leaf - plant 3

city: Pretoria

first: Dave

middle: Kenneth

last: Berger

state: Gauteng

address: Lynnwood Road

department: Plant and Soil Science

country: South Africa

institute: University of Pretoria

description:
BR3A
Maize leaf with minimal infection by C. zeina.
Processed data file: edgeR_BR_all.txt

data_processing:
Data were filtered to remove adapters and trimmed for quality using
Trimmomatic (v0.30; (Bolger et al., 2014); settings
TruSeq3-PE-2.fa:2:30:10 LEADING:3 SLIDINGWINDOW:4:15 MINLEN:36).
Filtered reads were aligned to a combined Zea mays genome (v5b.60 from the Phytozome v9.1 resource, Schaeffer et al., 2013) and the draft genome of Cercospora zeina (GenBank SUB212877) using Tophat2 with following setting: -p 4 -r 0 --mate-std-dev 200.
The longest transcript gene structure in the Z. mays annotation GFF3 file (retrieved from the Phytozome v9.1 resource) was used to estimate the transcript abundance in each locus by HTSeq.
Final data output include only values for Cercospora zeina genes.
Genome_build: Cercospora zeina draft genome
Supplementary_files_format_and_content: Tab-delimited text file includes raw counts for each gene.

organism: Zea mays

source_name: leaf, minimal C. zeina infection

growth_protocol: Maize B73 grown in the field in Greytown, Kwazulu-Natal, South Africa with natural infection of Cercospora zeina. Planted in December 2012.

extract_protocol:
Total RNA was extracted using Qiazol, treated with DNase, and purified with RNeasy Plant Mini kit according to manufacturer's instructions (Qiagen, Hilden, Germany). Total RNA was shown to be of good quality using an Agilent Bioanalyser (average RIN number = 7.7).
RNA libraries were prepared for sequencing using standard Illumina protocols.

tag: maize cultivar

value: upper leaf

tag: c. zeina infection

value: minimal

tag: plant id


title: upper leaf - plant 2

description:
BR2A
Maize leaf with minimal infection by C. zeina.
Processed data file: edgeR_BR_all.txt


title: Exo 8% O2 rep1


title: Mouse D29 mutant erythroid cells ATAC-seq biological replicate 2


title: Exo 1% O2 rep2


title: upper leaf - plant 1

description:
BR1A
Maize leaf with minimal infection by C. zeina.
Processed data file: edgeR_BR_all.txt


title: ChIP-seq, BAT H3K27ac, Adiponectin-Cre HDAC3 floxed, 29C/5pm, replicate 1


title: Dark grown, Spaceflight, PhyD Rep3

city: Gainesville

first: Robert

middle: J.

last: Ferl

state: Florida

address: 1301 Fifield Hall PO Box 110690

department: Horticultural Sciences

institute: University of Florida

description:
DK FLT PhyD Rep3
8. DK FLT Col-0 to DK FLT PhyD
14. DK FLT PhyD to DK GC PhyD
20. DK FLT PhyD to LT FLT PhyD

data_processing:
The paired end reads were mapped to the A. thaliana var. Columbia reference genome (TAIR10) using TopHat (Trapnell et al., 2009).
Multi-step process of transcriptome assembly and differential expression analysis was done using the Cufflinks tool (Trapnell et al., 2010; Trapnell et al., 2012). Reads that map to each transcript were counted and normalized based on fragment length and total reads. Normalized counts were expressed in terms of FPKM values (fragments per kilobase of transcript per million mapped fragments). FPKM is directly proportional to abundance of the transcript. The expression data was generated at a False Discovery Rate = 0.01 (FDR=0.01). At this FDR, a total of 33,234 genes were tested.
Genome_build: TAIR10
Supplementary_files_format_and_content: FPKM, fold change

organism: Arabidopsis thaliana

source_name: solid 0.5x MS media plates; grown in spaceflight, 11 day old plant harvested into KFT containing RNAlater fixative

growth_protocol: Three seed lines Wild-Type Wassilewskija (Ws), Columbia-0 (Col-0) and Col-0 PhyD (phyD) Mutants were tested for viability, sterility and ability to maintain dormancy before the launch. Tested batches of seeds were planted on phytagel plates as one genotype per plate for gene expression analysis in replicates of three. One set was planted for the flight and one for ground control. The plates were wrapped such that every surface of the plate was covered by two layers of Duvetyn Black-Out cloth (Seattle Fabrics) (Sng et al, 2014). The plates were stored 4° C until launch, and was then launched in a cold-stow bag to maintain the plates at 4° C until integration and activation on the ISS.

extract_protocol:
Seedlings stored in RNA later were recovered from -80°C freezer. Each tube containing the seedlings was allowed to thaw in the fridge overnight. After completely thawing the samples to room temperature, the seedlings were observed under the microscope. The preserved seedlings from the spaceflight and ground control harvests were dissected into distinct plant parts: leaf, hypocotyl, root tip and then the rest of the root. The root tip was defined as the last 2mm of the root for the light-grown plants and the last 1mm for the dark-grown plants. All the sections dissected were saved in RNA later and stored back to -80°C freezer.  Three independent roots were used as three biological replicates for the transcriptomic analysis. The root apex was dissected as 2mm root tip section for light grown samples and 1mm section for the dark grown samples. Dissected root tips were carefully transferred into pre-labeled micro-centrifuge tubes, flash frozen in liquid N. Total RNA was extracted from these frozen samples using Picopure RNA isolation kit (Arcturus®, Life technologies). RNA concentration was determined on a Qubit (Thermo Fisher/Life Technologies, Inc) and sample quality was assessed using the Agilent 2100 Bioanalyzer (Agilent Technologies, Inc.). Five ng RNA from each sample was used for low input array and RNA sequencing.
RNASeq Libraries Preparation using SMART-Seq V4 ultra low input RNA kit for sequencing combined with Illumina Nextera DNA Sample Preparation Kit  Five ng of total RNA was used for cDNA library construction using CloneTech SMART-Seq V4 ultra low input RNA kit for sequencing (Clontech Laboratories, Inc, cat#: 634890) according to manufacturer's protocol. Briefly, 1st strand cDNA was primed by the SMART-Seq v4 oligonucleotide and then base-pairs with these additional nucleotides, creating an extended template. The reverse transcriptase then switches templates and continues transcribing to the end of the oligonucleotide, resulting full-length cDNA that contains an anchor sequence that serves as a universal priming site for second strand synthesis. cDNA was amplified with primer II A for 9 PCR cycles. Then Illumina sequencing libraries were generated with 120 pg of cDNA using Illumina Nextera DNA Sample Preparation Kit (Cat#: FC-131-1024) according to manufacturer’s instructions. Briefly, 120 pg of cDNA was fragmented by tagementation reaction and then adapter sequences added onto template cDNA by PCR amplification. Libraries were quantitated by Bioanalyzer and qPCR (Kapa Biosystems, catalog number: KK4824). Finallly, the libraries were pooled equal molar concentration and sequenced by Illumina 2X75 NextSeq 500.
NextSeq500 sequencing: RNA seq  In preparation for sequencing, barcoded libraries were sized on the bioanalyzer, quantitated by QUBIT and qPCR (Kapa Biosystems, catalog number: KK4824). Individual samples were pooled equimolarly at 4 nM. This “working pool” was used as input in the NextSeq500 instrument sample preparation protocol (Illumina, Part # 15048776, Rev A). Typically, a 1.3 pM library concentration resulted in optimum clustering density in our instrument (i.e., ~200,000 clusters per mm2). Samples were sequenced on three flowcells, using a 2x75 cycles (paired-end) configuration. A typical sequencing run in the NextSeq500 produced 750-800 million paired-end read with a Q30>=85%. For RNA seq, around 40 million reads provided sufficient depth for transcriptome analysis.

treatment_protocol: The dormant plates were activated on station by removing the Black-Out cloth wrapping 12 days after launch. The plates were then placed on a fabric that was mounted in the US Laboratory module on the wall adjoining the MELFI freezer and secured using Velcro. The plants were allowed to grow on orbit for 11 days; some in the ambient light of ISS and some in the dark. The dark-grown plates were first activated by exposing the seeds to light for 4 hours, and then re-wrapped in Black-Out cloth for the duration of the growth period. A corresponding set of seedlings were grown as ground control in KSC.   At 11 days, seedlings were photographed, harvested into KFT containing RNAlater solutions and returned for post-flight analysis. All plates were harvested into KFTs with their counterpart (e.g. Light 1 was harvested with Dark 1). Once the plants were placed in the KFTs, the KFT was actuated with RNAlater to preserve the sample. At 24 hours post-harvest, KFTs were then transferred to MELFI, the -32°C freezer. Following SpaceX-3 splashdown in the Pacific Ocean, the KFTs transferred to the Cold Stowage charter plane at the Long Beach Airport, placed into an insulated shipper with dry ice, and flown to Johnson Space Center (JSC). The KFTs were then transferred via FedEx ground to the Kennedy Space Center. The KFTs were removed from dry ice and transferred to a -80°C freezer on May 22, 2014. The PIs retrieved the plant samples from the KFTs on May 29, 2014 and transferred the samples back to the University of Florida.  The harvested material was used to compare the transcriptomes of each genotype. The patterns of gene expression was compared among genotypes within a treatment (spaceflight or unit gravity) and between treatments (spaceflight versus ground control) for each genotype.

tag: cultivar

value: PhyD

value: Roots

tag: organism part

value: root tip - the last 1mm of the root for the dark grown plants

tag: age

value: 11 days old plants

tag: protocol

value: dark grown

tag: biosource type

value: RNAlater fixed_sample


title: Dark grown, Spaceflight, WS Rep3

description:
DK FLT WS Rep3
4. DK FLT Col-0 to DK FLT WS
10. DK FLT WS to DK GC WS
16. DK FLT WS to LT FLT WS

value: WS


title: Dark grown, Spaceflight, PhyD Rep1

description:
DK FLT PhyD Rep1
8. DK FLT Col-0 to DK FLT PhyD
14. DK FLT PhyD to DK GC PhyD
20. DK FLT PhyD to LT FLT PhyD


title: Dark grown, Spaceflight, PhyD Rep2

description:
DK FLT PhyD Rep2
8. DK FLT Col-0 to DK FLT PhyD
14. DK FLT PhyD to DK GC PhyD
20. DK FLT PhyD to LT FLT PhyD


title: Dark grown, Spaceflight, WS Rep2

description:
DK FLT WS Rep2
4. DK FLT Col-0 to DK FLT WS
10. DK FLT WS to DK GC WS
16. DK FLT WS to LT FLT WS


title: Dark grown, Spaceflight, Col-0 Rep3

description:
DK FLT Col-0 Rep3
4. DK FLT Col-0 to DK FLT WS
8. DK FLT Col-0 to DK FLT PhyD
12. DK FLT Col-0 to DK GC Col-0
18. DK FLT Col-0 to LT FLT Col-0


title: Exo NP rep1

value: Exosome


title: Dark grown, Spaceflight, Col-0 Rep2

description:
DK FLT Col-0 Rep2
4. DK FLT Col-0 to DK FLT WS
8. DK FLT Col-0 to DK FLT PhyD
12. DK FLT Col-0 to DK GC Col-0
18. DK FLT Col-0 to LT FLT Col-0


title: Exo 8% O2 rep2


title: Light grown, Spaceflight, WS Rep3

description:
LT FLT WS Rep3
2. LT FLT Col-0 to LT FLT WS
9. LT FLT WS to LT GC WS
16. DK FLT WS to LT FLT WS

value: root tip - the last 2mm of the root for the light grown plants

value: light grown


title: Light grown, Spaceflight, WS Rep2

description:
LT FLT WS Rep2
2. LT FLT Col-0 to LT FLT WS
9. LT FLT WS to LT GC WS
16. DK FLT WS to LT FLT WS


title: Light grown, Spaceflight, PhyD Rep3

description:
LT FLT PhyD Rep3
6. LT FLT Col-0 to LT FLT PhyD
13. LT FLT PhyD to LT GC PhyD
20. DK FLT PhyD to LT FLT PhyD


title: Light grown, Spaceflight, PhyD Rep2

description:
LT FLT PhyD Rep2
6. LT FLT Col-0 to LT FLT PhyD
13. LT FLT PhyD to LT GC PhyD
20. DK FLT PhyD to LT FLT PhyD


title: Light grown, Spaceflight, PhyD Rep1

description:
LT FLT PhyD Rep1
6. LT FLT Col-0 to LT FLT PhyD
13. LT FLT PhyD to LT GC PhyD
20. DK FLT PhyD to LT FLT PhyD


title: Mouse wild type erythroid cells H3K4me3 ChIP biological replicate 1

tag: antibody

value: anti_H3K4me3, Abcam ChIP Grade (ab8580)


title: Light grown, Spaceflight, Col-0 Rep3

description:
LT FLT Col-0 Rep3
2. LT FLT Col-0 to LT FLT WS
6. LT FLT Col-0 to LT FLT PhyD
11. LT FLT Col-0 to LT GC Col-0
18. DK FLT Col-0 to LT FLT Col-0

tag: tissue


title: Light grown, Spaceflight, Col-0 Rep1

description:
LT FLT Col-0 Rep1
2. LT FLT Col-0 to LT FLT WS
6. LT FLT Col-0 to LT FLT PhyD
11. LT FLT Col-0 to LT GC Col-0
18. DK FLT Col-0 to LT FLT Col-0


title: Dark grown, Ground Control, PhyD Rep3

description:
DK GC PhyD Rep3
7. DK GC Col-0 to DK GC PhyD
14. DK FLT PhyD to DK GC PhyD
19. DK GC PhyD to LT GC PhyD

source_name: solid 0.5x MS media plates; grown on the ground, 11 day old plant harvested into KFT containing RNAlater fixative


title: Dark grown, Spaceflight, Col-0 Rep1

description:
DK FLT Col-0 Rep1
4. DK FLT Col-0 to DK FLT WS
8. DK FLT Col-0 to DK FLT PhyD
12. DK FLT Col-0 to DK GC Col-0
18. DK FLT Col-0 to LT FLT Col-0


title: Light grown, Spaceflight, Col-0 Rep2

description:
LT FLT Col-0 Rep2
2. LT FLT Col-0 to LT FLT WS
6. LT FLT Col-0 to LT FLT PhyD
11. LT FLT Col-0 to LT GC Col-0
18. DK FLT Col-0 to LT FLT Col-0


title: Light grown, Spaceflight, WS Rep1

description:
LT FLT WS Rep1
2. LT FLT Col-0 to LT FLT WS
9. LT FLT WS to LT GC WS
16. DK FLT WS to LT FLT WS


title: Dark grown, Ground Control, PhyD Rep2

description:
DK GC PhyD Rep2
7. DK GC Col-0 to DK GC PhyD
14. DK FLT PhyD to DK GC PhyD
19. DK GC PhyD to LT GC PhyD


title: Dark grown, Ground Control, WS Rep3

description:
DK GC WS Rep3
3. DK GC Col-0 to DK GC WS
10. DK FLT WS to DK GC WS
15. DK GC WS to LT GC WS


title: Dark grown, Ground Control, PhyD Rep1

description:
DK GC PhyD Rep1
7. DK GC Col-0 to DK GC PhyD
14. DK FLT PhyD to DK GC PhyD
19. DK GC PhyD to LT GC PhyD


title: Dark grown, Ground Control, WS Rep1

description:
DK GC WS Rep1
3. DK GC Col-0 to DK GC WS
10. DK FLT WS to DK GC WS
15. DK GC WS to LT GC WS


title: Dark grown, Spaceflight, WS Rep1

description:
DK FLT WS Rep1
4. DK FLT Col-0 to DK FLT WS
10. DK FLT WS to DK GC WS
16. DK FLT WS to LT FLT WS


title: Dark grown, Ground Control, Col-0 Rep3

description:
DK GC Col-0 Rep3
3. DK GC Col-0 to DK GC WS
7. DK GC Col-0 to DK GC PhyD
12. DK FLT Col-0 to DK GC Col-0
17. DK GC Col-0 to LT GC Col-0


title: Dark grown, Ground Control, Col-0 Rep1

description:
DK GC Col-0 Rep1
3. DK GC Col-0 to DK GC WS
7. DK GC Col-0 to DK GC PhyD
12. DK FLT Col-0 to DK GC Col-0
17. DK GC Col-0 to LT GC Col-0


title: Light grown, Ground Control, PhyD Rep1

description:
LT GC PhyD Rep1
5. LT GC Col-0 to LT GC PhyD
13. LT FLT PhyD to LT GC PhyD
19. DK GC PhyD to LT GC PhyD


title: Light grown, Ground Control, PhyD Rep2

description:
LT GC PhyD Rep2
5. LT GC Col-0 to LT GC PhyD
13. LT FLT PhyD to LT GC PhyD
19. DK GC PhyD to LT GC PhyD


title: Light grown, Ground Control, PhyD Rep3

description:
LT GC PhyD Rep3
5. LT GC Col-0 to LT GC PhyD
13. LT FLT PhyD to LT GC PhyD
19. DK GC PhyD to LT GC PhyD


title: Light grown, Ground Control, WS Rep3

description:
LT GC WS Rep3
1. LT GC Col-0 to LT GC WS
9. LT FLT WS to LT GC WS
15. DK GC WS to LT GC WS


title: Dark grown, Ground Control, Col-0 Rep2

description:
DK GC Col-0 Rep2
3. DK GC Col-0 to DK GC WS
7. DK GC Col-0 to DK GC PhyD
12. DK FLT Col-0 to DK GC Col-0
17. DK GC Col-0 to LT GC Col-0


title: Light grown, Ground Control, Col-0 Rep3

description:
LT GC Col-0 Rep3
1. LT GC Col-0 to LT GC WS
5. LT GC Col-0 to LT GC PhyD
11. LT FLT Col-0 to LT GC Col-0
17. DK GC Col-0 to LT GC Col-0


title: Light grown, Ground Control, WS Rep2

description:
LT GC WS Rep2
1. LT GC Col-0 to LT GC WS
9. LT FLT WS to LT GC WS
15. DK GC WS to LT GC WS


title: Light grown, Ground Control, WS Rep1

description:
LT GC WS Rep1
1. LT GC Col-0 to LT GC WS
9. LT FLT WS to LT GC WS
15. DK GC WS to LT GC WS


title: Light grown, Ground Control, Col-0 Rep2

description:
LT GC Col-0 Rep2
1. LT GC Col-0 to LT GC WS
5. LT GC Col-0 to LT GC PhyD
11. LT FLT Col-0 to LT GC Col-0
17. DK GC Col-0 to LT GC Col-0