title: Aact_24h_1

city: Braunschweig

first: Szymon

middle: Piotr

last: Szafranski

address: Inhoffenstr. 7

department: Research Group Microbial Communication

institute: Helmholtz Centre for Infection Research

description:
mRNA was enriched
Aact_RNAseq.xlsx

data_processing:
Sequence reads after library adapter clipping were aligned with the reference using Bowtie 2 (Langmead et al., 2012). Afterwards, HTSeq-count was used to count the mapped reads per gene (http://www-huber.embl.de/users/anders/HTSeq). Differential gene expression was calculated using R package edgeR (Robinson et al., 2010). Genes showing differential expression in dual-species biofilms (all time points compared to the 4 h time point, log2 fold change < - 2 and > 2; p < 0.01, were sorted into clusters according to their expression profile using the c-means algorithm (Kumar et al., 2007). Rockhopper software for RNAseq data analysis was used to identify novel transcripts and determine operons (McClure et al., 2013).
Genome_build: The S. mutans UA159 and A. act. HK1651 reference genomes (GenBank accession numbers NC_004350.2 and CP007502.1, respectively) were used for RNA-seq read alignment (Ajdic et al., 2002;Najer, 2002).
Supplementary_files_format_and_content: In two files: Aact_RNAseq.xlsx and Smut_RNAseq.xlsx there are processed data for A.act. and S.mutans, respectively. Log fold changes (dual vs single, and all time points vs 4 h), FDR (p values), and mean log CPM values are given for every gene. Additional information for genes are also provided (start, end, name, locus, product, strand,  clustering based on expression changes in time).

organism: Aggregatibacter actinomycetemcomitans

source_name: In vitro biofilm, static in 24-well plate

growth_protocol: Overnight cultures of both organisms were centrifuged (5000 rpm, 10 min, 4°C) and suspended in artificial saliva (modified from Wong and Sissons 2001;  without mucin, haemin replaced with 1.2 µM FeCl3 ) and inoculated at OD600 of 0.1 (equivalent of roughly 10 to the power 7 CFU). Cultivation of biofilms was performed in 24 Well Cell Culture Plates (Greiner Bio-One, Frickenhausen, Germany). The plates were incubated aerobically (5% CO2) at 37°C. Biofilms were allowed to develop for 4, 6, 8, 10, 12 and 24 h. After removal of the supernatants, the single and dual species biofilms were covered with 300 µl RNA protect (Qiagen, Hilden, Germany). The biofilms were scraped off with sterile one-way cell scrapers, transferred to sterile Eppendorf tubes and centrifuged (4°C, 5 min at 13,000 rpm). The pellets were stored at -70°C until RNA extraction.

extract_protocol:
For RNA isolation the pellets were resuspended in lysis buffer containing 10 mM Tris, 1 mM EDTA, pH 8.0, 2.5 mg/ml lysozyme (Sigma-Aldrich, Taufkirchen, Germany) and 50 U/ml mutanolysin (Sigma-Aldrich, Taufkirchen, Germany) and incubated at 25°C for 1.5 h at 350 rpm. Subsequently TRIzol reagent (Invitrogen, Carlsbad, CA, USA) was added, samples were well mixed and incubated at room temperature for 5 min. Next the cells were vortexed for 10 times 30 s with 1 min intervals on ice in the presence of 50 µg sterile, acid-washed glass beads (diameter 106 µm; Sigma-Aldrich, Taufkirchen, Germany). Samples were briefly centrifuged and supernatants were vortexed with ice cooled 1-bromo-3-chloropropane (Sigma-Aldrich, Taufkirchen, Germany) for 15 s followed by 5 min incubation at room temperature. Next, samples were centrifuged (4°C, 15 min at 13,000 rpm) and RNA in collected aqueous phase was precipitated with 1 volume of ice cooled isopropanol and suspended in water. After removal of DNA with RNase free DNaseI (Qiagen, Hilden, Germany) using the in-solution digestion protocol, RNA samples were purified using the RNeasy mini kit (Qiagen, Hilden, Germany) according to the manufacturer`s instructions with the additional DNaseI on-column digestion. Bacterial rRNAs were depleted using MicrobExpress (Ambion, Austin, TX).
mRNA was converted to cDNA using a ScriptSeq v2 RNA-Seq (Epicentre Biotechnologies, Madison, Wisconsin, USA).

tag: biofilm

value: single-species

tag: time (h)

tag: replica


title: Smut_24h_1

status: Public on Mar 31 2017

type: SRA

country: Germany

description:
mRNA was enriched
Smut_RNAseq.xlsx

library_source: transcriptomic

molecule: total RNA

organism: Streptococcus mutans


title: 5FU_Day14_55

city: Cambridge

first: Evangelia

last: Diamanti

address: Wellcome Trust / MRC Building, Hills Road

department: Haematology

country: United Kingdom

institute: University of Cambridge

data_processing:
RNA-seq reads were aligned to mm10 using GSNAP (version 2015-09-29) with parameters (-B 5 -t 24 -n 1 -Q -N 1).
Reads in features were counted with htseq-count (HTSeq version 0.5.3p3) with the parameter (-s no).
Genome_build: mm10
Supplementary_files_format_and_content: HT-Seq counts (.txt)

organism: Mus musculus

source_name: Bone Marrow

extract_protocol:
Six- to 12-weeks-old C57Bl/6 wilt type mice were treated with a single dose of 150 mg/kg 5-FU (Sigma-Aldrich) or with PBS control and were analyzed at different time points (0, 8, 10, 12 and 14 days) post-treatment. Bone marrow (BM) cells were obtained by either crushing leg, arm and pelvic bones or just flushing leg bones in Hanks’ Buffered Saline Solution (HBSS) containing 2% heat-inactivated FBS (Sigma-Aldrich). HBSS/2% FBS was used for all incubation and wash steps. Erythrocytes and contaminating bone material were removed by ACK lysis followed by centrifugation on a Ficoll gradient (Histopaque 1119, Sigma-Aldrich) for crushed bones. BM cells were first pre-enriched for c-Kit+ cells using c-Kit microbeads (Miltenyi Biotec, 130-091-224) and MACS Separation LS Columns (Miltenyi Biotec, 130-042-401), and incubated with purified, unconjugated-lineage antibodies (CD3 from Biolegend, and CD4, CD5, CD8, B220, Ter119, Mac-1 and Gr-1 from eBioscience) followed by goat anti-rat-PE-Cy5 (Invitrogen, A10691) and subsequently blocked with purified rat IgG (Sigma-Aldrich). Cells were then stained with c-Kit-APC-eFluor780 (eBioscience, 47-1171-82), Sca-1-PB (BioLegend, 108120), CD48-A647 (BioLegend, 103416), CD150-PE (BioLegend, 115904), CD34-FITC (eBioscience, 11-0341-85) and FcγR-PerCP-eFluor710 (eBioscience, 46-0161-82) after 5-FU treatment. Single GMPs (lin-/c-kit+/sca-1-/cd34+/fcgr+) were directly sorted into individual wells of a 96-well PCR plate containing 2.3 µl of 0.2% Triton-X100 (Sigma-Aldrich, Cat. no. 93443) and 2.3 U of SUPERase-In RNase Inhibitor (Ambion, Cat no. AM2694). cDNA was obtained and amplified following the described SMARTSEQ2 protocol (Picelli et al., 2014). Libraries were prepared for sequencing from 125 ng of cDNA using the Illumina Nextera XT DNA preparation kit. Pooled libraries were run on the Illumina Hi-Seq 2500.
Libraries were prepared for sequencing from 125 ng of cDNA using the Illumina Nextera XT DNA preparation kit.

tag: strain

tag: cell type

value: GMP

tag: treatment

tag: time


title: Smut_24h_2

description:
mRNA was enriched
Smut_RNAseq.xlsx


title: Smut_08h_2

description:
mRNA was enriched
Smut_RNAseq.xlsx


title: Aact_08h_2

description:
mRNA was enriched
Aact_RNAseq.xlsx


title: Aact_08h_1

description:
mRNA was enriched
Aact_RNAseq.xlsx


title: BA-4_60

extract_protocol:
C57Bl/6 Scl-tTA::TRE-BCR/ABL (BA)  mice as previously described (Reynaud, D et al., Cancer Cell, 2011), were bred in the presence of 20mg/L doxycycline in their drinking water, and induced for CML-like MPN development at 5 weeks of age by doxycycline withdrawal. BA and control mice were analyzed 5-6 weeks after BCR/ABL induction. Bone marrow (BM) cells were obtained by either crushing leg, arm and pelvic bones or just flushing leg bones in Hanks’ Buffered Saline Solution (HBSS) containing 2% heat-inactivated FBS (Sigma-Aldrich). HBSS/2% FBS was used for all incubation and wash steps. Erythrocytes and contaminating bone material were removed by ACK lysis followed by centrifugation on a Ficoll gradient (Histopaque 1119, Sigma-Aldrich) for crushed bones. BM cells were first pre-enriched for c-Kit+ cells using c-Kit microbeads (Miltenyi Biotec, 130-091-224) and MACS Separation LS Columns (Miltenyi Biotec, 130-042-401), and incubated with purified, unconjugated-lineage antibodies (CD3 from Biolegend, and CD4, CD5, CD8, B220, Ter119, Mac-1 and Gr-1 from eBioscience) followed by goat anti-rat-PE-Cy5 (Invitrogen, A10691) and subsequently blocked with purified rat IgG (Sigma-Aldrich). Cells were then stained with c-Kit-APC-eFluor780 (eBioscience, 47-1171-82), Sca-1-PB (BioLegend, 108120), CD48-A647 (BioLegend, 103416), CD150-PE (BioLegend, 115904), CD34-FITC (eBioscience, 11-0341-85) and FcγR-PerCP-eFluor710 (eBioscience, 46-0161-82). Single GMPs (lin-/c-kit+/sca-1-/cd34+/fcgr+) were directly sorted into individual wells of a 96-well PCR plate containing 2.3 µl of 0.2% Triton-X100 (Sigma-Aldrich, Cat. no. 93443) and 2.3 U of SUPERase-In RNase Inhibitor (Ambion, Cat no. AM2694). cDNA was obtained and amplified following the described SMARTSEQ2 protocol (Picelli et al., 2014). Libraries were prepared for sequencing from 125 ng of cDNA using the Illumina Nextera XT DNA preparation kit. Pooled libraries were run on the Illumina Hi-Seq 4000.
Libraries were prepared for sequencing from 125 ng of cDNA using the Illumina Nextera XT DNA preparation kit.

value: BCR/ABL induction


title: 5FU_Day14_61


title: Smut_08h_1

description:
mRNA was enriched
Smut_RNAseq.xlsx


title: BA-4_5


title: BA-4_59


title: BA-4_58


title: Aact_24h_2

description:
mRNA was enriched
Aact_RNAseq.xlsx


title: BA-4_57


title: 5FU_Day14_52


title: 5FU_Day14_53


title: 5FU_Day14_4


title: 5FU_Day14_50


title: BA-4_56


title: BA-4_64


title: BA-4_54


title: 5FU_Day14_54


title: 5FU_Day14_51


title: 5FU_Day14_49


title: BA-4_55


title: 5FU_Day14_48


title: BA-4_51


title: 5FU_Day14_47


title: BA-4_50


title: BA-4_49


title: BA-4_48


title: 5FU_Day14_44


title: 5FU_Day14_46


title: BA-4_4


title: 5FU_Day14_43


title: BA-4_46


title: 5FU_Day14_41


title: BA-4_53


title: 5FU_Day14_40


title: 5FU_Day14_3


title: BA-4_52


title: 5FU_Day14_45


title: BA-4_44


title: 5FU_Day14_39


title: BA-4_42


title: BA-4_43


title: BA-4_47


title: BA-4_41


title: 5FU_Day14_37


title: 5FU_Day14_35


title: 5FU_Day14_34


title: 5FU_Day14_42


title: 5FU_Day14_31


title: BA-4_45


title: BA-4_40


title: BA-4_3


title: 5FU_Day14_32


title: BA-4_37


title: BA-4_38


title: BA-4_36


title: 5FU_Day14_2


title: BA-4_34


title: 5FU_Day14_38


title: 5FU_Day14_30


title: BA-4_39


title: BA-4_33


title: BA-4_35


title: 5FU_Day14_29


title: 5FU_Day14_27


title: 5FU_Day14_26


title: 5FU_Day14_36


title: BA-4_31


title: BA-4_30


title: 5FU_Day14_25


title: 5FU_Day14_24


title: 5FU_Day14_23


title: BA-4_2


title: BA-4_28


title: 5FU_Day14_22


title: 5FU_Day14_21


title: 5FU_Day14_28


title: 5FU_Day14_20


title: BA-4_26


title: 5FU_Day14_1


title: BA-4_25


title: BA-4_29


title: BA-4_22


title: BA-4_21


title: 5FU_Day14_19


title: BA-4_32


title: 5FU_Day14_17


title: BA-4_23


title: 5FU_Day14_15


title: BA-4_24


title: 5FU_Day14_18


title: BA-4_1


title: 5FU_Day14_13


title: BA-4_20


title: 5FU_Day14_14


title: 5FU_Day14_33


title: BA-4_27


title: BA-4_18


title: 5FU_Day14_12


title: BA-4_17


title: 5FU_Day14_11


title: BA-4_16


title: BA-4_14


title: 5FU_Day12_9


title: 5FU_Day12_96


title: BA-4_15


title: 5FU_Day12_95


title: BA-4_13


title: 5FU_Day14_10


title: 5FU_Day14_16


title: BA-4_19


title: 5FU_Day12_92


title: BA-4_12


title: BA-4_11


title: BA-2_96


title: BA-2_9


title: BA-2_95


title: 5FU_Day12_93


title: 5FU_Day12_94


title: BA-4_10


title: 5FU_Day12_89


title: 5FU_Day12_8


title: BA-2_94


title: 5FU_Day12_90


title: BA-2_92


title: 5FU_Day12_91


title: 5FU_Day12_88


title: BA-2_93


title: 5FU_Day12_86


title: 5FU_Day12_87


title: BA-2_91


title: BA-2_90


title: BA-2_8


title: 5FU_Day12_83


title: BA-2_88


title: 5FU_Day12_82


title: BA-2_87


title: 5FU_Day12_85


title: BA-2_86


title: 5FU_Day12_7


title: 5FU_Day12_80


title: BA-2_84


title: 5FU_Day12_81


title: BA-2_85


title: 5FU_Day12_79


title: BA-2_83


title: BA-2_82


title: BA-2_81


title: 5FU_Day12_75


title: 5FU_Day12_77


title: 5FU_Day12_74


title: 5FU_Day12_78


title: 5FU_Day12_76


title: BA-2_79


title: BA-2_7


title: 5FU_Day12_71


title: BA-2_77


title: 5FU_Day12_72


title: BA-2_78


title: 5FU_Day12_84


title: 5FU_Day12_70


title: BA-2_89


title: BA-2_74


title: 5FU_Day12_6


title: 5FU_Day12_69


title: 5FU_Day12_66


title: 5FU_Day12_67


title: BA-2_73


title: BA-2_70


title: BA-2_71


title: 5FU_Day12_65


title: 5FU_Day12_73


title: 5FU_Day12_68


title: BA-2_72


title: BA-2_69


title: 5FU_Day12_62


title: 5FU_Day12_61


title: BA-2_68


title: BA-2_75


title: 5FU_Day12_63


title: BA-2_66


title: 5FU_Day12_60


title: BA-2_64


title: BA-2_63


title: 5FU_Day12_59


title: BA-2_76


title: BA-2_65


title: BA-2_6


title: 5FU_Day12_58


title: 5FU_Day12_57


title: 5FU_Day12_56


title: 5FU_Day12_55


title: BA-2_60


title: BA-2_62


title: BA-2_59


title: 5FU_Day12_53


title: 5FU_Day12_52


title: 5FU_Day12_64


title: BA-2_67


title: BA-2_58


title: 5FU_Day12_51


title: BA-2_57


title: BA-2_56


title: 5FU_Day12_5


title: 5FU_Day12_50


title: 5FU_Day12_4


title: 5FU_Day12_49


title: 5FU_Day12_47


title: 5FU_Day12_48


title: BA-2_55


title: BA-2_53


title: BA-2_54


title: BA-2_61


title: BA-2_52


title: 5FU_Day12_46


title: 5FU_Day12_54


title: 5FU_Day12_45


title: BA-2_4


title: BA-2_51


title: 5FU_Day12_43


title: 5FU_Day12_42


title: BA-2_48


title: BA-2_49


title: BA-2_47


title: 5FU_Day12_41


title: 5FU_Day12_40


title: BA-2_50


title: BA-2_46


title: BA-2_44


title: 5FU_Day12_39


title: 5FU_Day12_38


title: BA-2_41


title: 5FU_Day12_36


title: BA-2_40


title: 5FU_Day12_44


title: 5FU_Day12_34


title: BA-2_39


title: 5FU_Day12_33


title: BA-2_36


title: BA-2_37


title: 5FU_Day12_30


title: BA-2_35


title: 5FU_Day12_31


title: 5FU_Day12_28


title: BA-2_34


title: 5FU_Day12_29


title: 5FU_Day12_2


title: BA-2_32


title: BA-2_42


title: 5FU_Day12_3


title: BA-2_43


title: BA-2_45


title: 5FU_Day12_35


title: 5FU_Day12_25


title: 5FU_Day12_27


title: BA-2_38


title: BA-2_5


title: BA-2_31


title: BA-2_30


title: BA-2_29


title: BA-2_2


title: BA-2_33


title: BA-2_28


title: 5FU_Day12_37


title: BA-2_26


title: 5FU_Day12_21


title: 5FU_Day12_32


title: BA-2_80


title: 5FU_Day12_22


title: 5FU_Day12_26


title: BA-2_24


title: BA-2_23


title: BA-2_25


title: 5FU_Day12_18


title: BA-2_3


title: BA-2_27


title: 5FU_Day12_1


title: BA-2_22


title: 5FU_Day12_17


title: BA-2_21


title: BA-2_20


title: 5FU_Day12_15


title: 5FU_Day12_16


title: BA-2_1


title: 5FU_Day12_14


title: 5FU_Day12_23


title: BA-2_19


title: 5FU_Day12_20


title: 5FU_Day12_12


title: BA-2_18


title: 5FU_Day12_19


title: 5FU_Day12_11


title: BA-2_14


title: BA-2_16


title: BA-2_15


title: BA-2_17


title: 5FU_Day10_9


title: 5FU_Day10_96


title: 5FU_Day12_10


title: BA-2_12


title: BA-2_13


title: BA-2_11


title: 5FU_Day10_93


title: 5FU_Day10_94


title: 5FU_Day10_90


title: 5FU_Day10_92


title: 5FU_Day10_91


title: BA-1_9


title: BA-1_96


title: BA-2_10


title: BA-1_95


title: 5FU_Day12_13


title: 5FU_Day10_89


title: 5FU_Day10_88


title: BA-1_94


title: BA-1_93


title: BA-1_92


title: 5FU_Day10_95


title: 5FU_Day10_87


title: BA-1_91


title: 5FU_Day10_86


title: 5FU_Day10_85


title: BA-1_90


title: BA-1_8


title: 5FU_Day10_8


title: BA-1_88


title: 5FU_Day10_83


title: 5FU_Day10_84


title: BA-1_89


title: BA-1_87


title: 5FU_Day10_82


title: BA-1_86


title: 5FU_Day10_80


title: 5FU_Day10_81


title: BA-1_84


title: 5FU_Day10_7


title: BA-1_85


title: BA-1_82


title: 5FU_Day10_79


title: 5FU_Day10_76


title: 5FU_Day10_78


title: 5FU_Day10_75


title: BA-1_80


title: BA-1_7


title: 5FU_Day10_74


title: BA-1_78


title: BA-1_79


title: BA-1_77


title: 5FU_Day10_73


title: BA-1_81


title: 5FU_Day10_69


title: BA-1_76


title: 5FU_Day10_70


title: 5FU_Day10_77


title: BA-1_74


title: BA-1_75


title: BA-1_83


title: 5FU_Day10_6


title: BA-1_73


title: BA-1_72


title: 5FU_Day10_66


title: 5FU_Day10_71


title: 5FU_Day10_72


title: BA-1_6


title: BA-1_70


title: BA-1_68


title: BA-1_67


title: 5FU_Day10_61


title: 5FU_Day10_60


title: 5FU_Day10_58


title: BA-1_63


title: 5FU_Day10_57


title: BA-1_62


title: BA-1_71


title: 5FU_Day10_67


title: 5FU_Day10_65


title: 5FU_Day10_63


title: BA-1_61


title: 5FU_Day10_68


title: 5FU_Day10_55


title: 5FU_Day10_64


title: 5FU_Day10_56


title: BA-1_60


title: 5FU_Day10_59


title: BA-1_69


title: BA-1_5


title: BA-1_66


title: BA-1_64


title: 5FU_Day10_52


title: BA-1_57


title: 5FU_Day10_51


title: 5FU_Day10_62


title: BA-1_65


title: BA-1_56


title: 5FU_Day10_5


title: 5FU_Day10_54


title: BA-1_58


title: 5FU_Day10_53


title: BA-1_59


title: 5FU_Day10_49


title: BA-1_55


title: BA-1_54


title: 5FU_Day10_48


title: BA-1_53


title: BA-1_52


title: 5FU_Day10_47


title: BA-1_4


title: BA-1_51


title: BA-1_50


title: 5FU_Day10_44


title: 5FU_Day10_46


title: BA-1_49


title: 5FU_Day10_42


title: BA-1_47


title: BA-1_48


title: 5FU_Day10_43


title: 5FU_Day10_41


title: 5FU_Day10_4


title: 5FU_Day10_45


title: 5FU_Day10_50


title: BA-1_45


title: 5FU_Day10_3


title: 5FU_Day10_39


title: BA-1_46


title: 5FU_Day10_40


title: BA-1_44


title: BA-1_43


title: 5FU_Day12_24


title: BA-1_42


title: 5FU_Day10_37


title: 5FU_Day10_36


title: 5FU_Day10_38


title: BA-1_41


title: BA-1_3


title: BA-1_39


title: 5FU_Day10_34


title: 5FU_Day10_33


title: 5FU_Day10_35


title: BA-1_40


title: BA-1_38


title: BA-1_37


title: 5FU_Day10_32


title: 5FU_Day10_31


title: BA-1_36


title: 5FU_Day10_30


title: BA-1_35


title: BA-1_34


title: BA-1_33


title: 5FU_Day10_29


title: 5FU_Day10_28


title: 5FU_Day10_27


title: BA-1_32


title: 5FU_Day10_26


title: BA-1_30


title: 5FU_Day10_25


title: BA-1_2


title: BA-1_28


title: 5FU_Day10_24


title: BA-1_31


title: 5FU_Day10_22


title: BA-1_29


title: BA-1_26


title: BA-1_24


title: 5FU_Day10_20


title: 5FU_Day10_1


title: 5FU_Day10_21


title: 5FU_Day10_19


title: 5FU_Day10_23


title: BA-1_23


title: 5FU_Day10_17


title: 5FU_Day10_2


title: BA-1_21


title: 5FU_Day10_15


title: 5FU_Day10_16


title: BA-1_20


title: 5FU_Day10_14


title: BA-1_19


title: BA-1_18


title: BA-1_17


title: 5FU_Day10_11


title: 5FU_Day10_12


title: BA-1_16


title: BA-1_15


title: BA-1_25


title: BA-1_27


title: 5FU_Day10_10


title: BA-1_14


title: 5FU_Day10_18


title: BA-1_12


title: 5FU_Day0_9


title: BA-1_11


title: 5FU_Day0_96


title: 5FU_Day0_95


title: 5FU_Day0_93


title: BA-1_13


title: BA-1_22


title: BA-1_10


title: 5FU_Day0_94


title: BA-1_1


title: 5FU_Day0_92


title: SmutAact_10h_2

description:
S.mutans, A. actinomycetemcomitans
mRNA was enriched
Aact_RNAseq.xlsx Smut_RNAseq.xlsx

organism:
Aggregatibacter actinomycetemcomitans
Streptococcus mutans

value: dual-species


title: 5FU_Day10_13


title: SmutAact_10h_1


title: SmutAact_06h_2


title: 5FU_Day0_90


title: 5FU_Day0_8


title: SmutAact_06h_1


title: 5FU_Day0_89


title: 5FU_Day0_88


title: SmutAact_24h_1


title: 5FU_Day0_87


title: SmutAact_24h_2


title: SmutAact_12h_1


title: 5FU_Day0_86


title: SmutAact_12h_2


title: SmutAact_08h_1


title: 5FU_Day0_84


title: SmutAact_04h_1


title: 5FU_Day0_83


title: SmutAact_04h_2


title: 5FU_Day0_80


title: 5FU_Day0_82


title: SmutAact_08h_2


title: 5FU_Day0_79


title: 5FU_Day0_7


title: 5FU_Day0_74


title: 5FU_Day0_78


title: 5FU_Day0_77


title: 5FU_Day0_73


title: 5FU_Day0_75


title: 5FU_Day0_76


title: 5FU_Day0_91


title: 5FU_Day0_72


title: 5FU_Day0_71


title: 5FU_Day0_70


title: 5FU_Day0_6


title: 5FU_Day0_85


title: 5FU_Day0_81


title: 5FU_Day0_67


title: 5FU_Day0_66


title: 5FU_Day0_69


title: 5FU_Day0_62


title: 5FU_Day0_65


title: 5FU_Day0_63


title: 5FU_Day0_59


title: 5FU_Day0_60


title: 5FU_Day0_56


title: 5FU_Day0_57


title: 5FU_Day0_5


title: 5FU_Day0_64


title: 5FU_Day0_55


title: 5FU_Day0_47


title: 5FU_Day0_51


title: 5FU_Day0_49


title: 5FU_Day0_53


title: 5FU_Day0_50


title: 5FU_Day0_48


title: 5FU_Day0_4


title: 5FU_Day0_52


title: 5FU_Day0_45


title: 5FU_Day0_68


title: 5FU_Day0_44


title: 5FU_Day0_43


title: 5FU_Day0_40


title: 5FU_Day0_61


title: 5FU_Day0_42


title: 5FU_Day0_3


title: 5FU_Day0_38


title: 5FU_Day0_39


title: 5FU_Day0_34


title: 5FU_Day0_31


title: 5FU_Day0_37


title: 5FU_Day0_32


title: 5FU_Day0_54


title: 5FU_Day0_30


title: 5FU_Day0_35


title: 5FU_Day0_46


title: 5FU_Day0_29


title: 5FU_Day0_2


title: 5FU_Day0_23


title: 5FU_Day0_27


title: 5FU_Day0_24


title: 5FU_Day0_26


title: 5FU_Day0_25


title: 5FU_Day0_58


title: 5FU_Day0_41


title: 5FU_Day0_22


title: 5FU_Day0_20


title: 5FU_Day0_21


title: 5FU_Day0_1


title: 5FU_Day0_18


title: 5FU_Day0_17


title: 5FU_Day0_33


title: 5FU_Day0_14


title: 5FU_Day0_16


title: 5FU_Day0_36


title: 5FU_Day0_12


title: 5FU_Day0_28


title: 5FU_Day0_11


title: 5FU_Day0_13


title: 5FU_Day0_19


title: ATACseq_NCCprogenitors_1

city: Basel

first: Filippo

middle: M.

last: RIJLI

address: Maulbeerstrasse 66

country: Switzerland

institute: Friedrich Miescher Institute for Biomedical Research

library_source: genomic

data_processing:
Library strategy: ATAC-Seq
Illumina RTA 1.18.64 and bcl2fastq2 v2.17 was used for basecalling and demultiplexing.
Read pairs were aligned to the GRCm38/mm10 reference genome using the QuasR Bioconductor package (PMID: 25417205, version 1.12.0) by the qAlign function with parameters maxHits=10 (allowing up to 10 hits per read, reporting one randomly), which internally uses bowtie (version 1.1.1, PMID: 19261174).
Genome_build: GRCm38/mm10
Supplementary_files_format_and_content: wig files were generated using the qExportWig function from the QuasR package with default parameters except scaling=1e6 and binsize=100. The scores in the wig file therefore represent the number of alignment pairs per 100bp window and per million reads in the library.

molecule: genomic DNA

source_name: Cranial neural crest cells

extract_protocol:
To profile for open chromatin, we used the Assay for Transposase Accessible Chromatin (ATACseq) protocol, performed according to Buenrostro et al., 2015 with minor modifications.  The dissected samples were incubated in trypsin 0.5%/EDTA 1X at 37 °C for 10 minutes and rinsed in DMEM 1X/ FBS 10%. Cranial neural crest cells (CNCCs) were then dissociated with a pipette, filtered and collected by FACS. 50.000 CNCCs were used per transposition reaction. After sorting, CNCCs were pelleted by centrifugation (500 g for 5 minutes at 4°C), washed with cold 1X PBS buffer and pelleted again by centrifugation (500 g for 5 minutes at 4°C). They were then re-suspended in cold lysis buffer (10mM Tris-HCl pH 8.0; 10mM NaCl; 3mM MgCl2; 0.5% NP40) and pelleted by centrifugation (5 minutes, 500 g at 4°C). The supernatant was discarded.
The transposition reaction was performed as reported in Buenrostro et al., 2015, steps 6 – 12, using the Nextera Sample preparation kit from Illumina. PCR Amplification was done according to Buenrostro et al., 2015, steps 13 – 18. The indexing PCR primers mentioned in the supplemental material from Buenrostro et al., 2013 were used. Amplified libraries were cleaned up using AMPure XP beads in a DNA / beads ratio of 1 / 1.8. The Quality of the libraries and size distribution was assessed on an Agilent 2100 Bioanalyzer (Agilent Technologies). Sequencing was performed on an Illumina HiSeq 2500 machine (50 bp read length, paired-end) according to Illumina standards.

value: Cranial neural crest cells

tag: cell subpopulation

value: NCCprogenitors

tag: age

tag: genotype

value: WT


title: ATACseq_E10.5-Md_1

value: Md


title: ATACseq_E10.5-PA2-Ezh2cKO_1


title: ATACseq_E10.5-PA2_2


title: ATACseq_E10.5-FNP_2

value: FNP


title: ATACseq_E10.5-FNP-Ezh2cKO_1


title: ATACseq_E10.5-PA2_1


title: 5FU_Day0_15


title: ATACseq_E10.5-Md_2


title: ATACseq_E10.5-FNP_1


title: ATACseq_E10.5-Md-Ezh2cKO_1


title: ChIPseq_E10.5-FNP_H3K4me2_2

data_processing:
Illumina RTA 1.18.64 and bcl2fastq2 v2.17 was used for basecalling and demultiplexing.
Reads were aligned to the GRCm38/mm10 reference genome using the QuasR Bioconductor package (REF: PMID: 25417205, version 1.12.0) by the qAlign function with parameters maxHits=10 (allowing up to 10 hits per read, reporting one randomly), which internally uses bowtie (version 1.1.1, PMID: 19261174).
Genome_build: GRCm38/mm10
Supplementary_files_format_and_content: wig files were generated using the qExportWig function from the QuasR package with default parameters except scaling=1e6 and binsize=100. The scores in the wig file therefore represent the number of alignments per 100bp window and per million reads in the library.

extract_protocol:
For the single ChIP-Seq experiments: Embryonic tissues were micro-dissected at E8.5, E10.5 or E11.5, incubated in trypsin 0.5%/EDTA 1X at 37 °C for 10 minutes, rinsed (DMEM 1X/ FBS 10%, followed by PBS 1X), cross-linked with 1% formaldehyde in PBS 1X for 10 minutes at room temperature (RT) and quenched with 125 mM glycine for 5 minutes at RT. Cranial neural crest cells (CNCCs) were pelleted by centrifugation (2000 rpm, 10 minutes, 4°C) and rinsed three times in ice-cold PBS. CNCCs were collected by FACS and pelleted by centrifugation (2000 rpm, 10 minutes, 4°C). 1 millions of FACS-isolated CNCCs were used for each single ChIp-Seq experiments. CNCCs were incubated in 1 ml of Cell Lysis Buffer (0.5% NP40, 85 mM KCl, 5 mM PIPES pH8) for 10 min on ice. Nuclei were isolated by spinning the lysed CNCCs for 5 minutes at 2500 g, at 4°C. The supernatant was discarded and cell nuclei were re-suspended in Sonication Buffer (50 mM Tris HCl pH8, 10 mM EDTA, 1% SDS, 1X Protease Inhibitor Cocktail (PIC - Complete-EDTA free, Roche)). After 15 min of lysis on a tube shaker at 4°C, the DNA was sonicated using the Covaris machine (settings: cycles per burst 200, duty cycle 10%, peak incidence power 175 Watts for 2400 sec.) to obtain a mean DNA fragment size of 250 bp. After centrifugation (10 minutes, 10 000 rpm, 4°C), the supernatant was transferred in a new tube, diluted 10X with RIPA buffer (10 mM Tris-HCl pH7.5, 140 mM NaCl, 1 mM EDTA, 0.5 mM EGTA, 1% Triton X100, 0.1% SDS, 0,1% Na-deoxycholate, 1 mM PMSF, 20 mM NaB, 1X PIC) and incubated over-night at 4°C with 5 μl of antibody. The next day, 50 μl of protein G coupled to magnetic beads (Dynabeads Protein G, Invitrogen) were added and the incubation continued for 2 hours. The beads were then washed 3 times with RIPA buffer and once with TE buffer (10Mm Tris-HCl pH 8.0, 1mM EDTA). The protein-DNA complexes were eluted from the beads with 150 μl of Elution Buffer (20 mM Tris-HCl pH 7.5, 5 mM EDTA, 50 mM NaCl, 1% SDS, 50 mg/mL proteinase K) at 68 °C for 2 hours, to reverse cross-links and digest proteins. DNA was purified using MinElute PCR purification kit (Qiagen).
ChIP libraries were prepared using bar-coded adapters according to standard Illumina library preparation protocols, following the manufactorer’s instructions for the NEBNext® Ultra™ DNA Library Prep Kit for Illumina®, or the TruSeq® ChIP sample preparation for Illumina with and without gel-size selection (for the protocol used, see “library protocol” annotation of each sample). The Quality of the libraries and size distribution was assessed on an Agilent 2100 Bioanalyzer (Agilent Technologies). One to four samples with different barcodes were mixed at equimolar ratios per pool. Sequencing was performed on an Illumina HiSeq 2500 machine (50 bp read length, single-end) according to Illumina standards.

tag: chip antibody

value: H3K4me2 (Millipore, Cat. # 07-030)

tag: library protocol

value: TrueSeq-Illumina, without gel-size selection


title: ChIPseq_E10.5-FNPMxMd-Ezh2cKO_H3K4me2_1

value: FNPMxMd


title: ChIPseq_Input-ChipSeqIllumina_1


title: ChIPseq_E10.5-FNP_H3K27me3_2

value: H3K27me3 (Millipore, Cat. # 07-449)


title: ChIPseq_E10.5-FNP_H3K27me3-H3K4me2_1

extract_protocol:
For sequential H3K27me3/H3K4me2 ChIP-Seq experiments, around 7 million of non-FACS isolated cells were used to perform the first ChIP as described above, up to the washing of beads with TE buffer, using several tubes. Denaturing conditions (DTT, high salt and detergent) and heat were used to release the chromatin from the antibody coated magnetic beads. Right after removal of the TE buffer, the DNA/H3K27me3-antibody/protein-G complexes were re-suspended in 25 ul of DTT 100 mM (5 minutes at RT). Then, 25 ul of Chromatin Release Buffer (500 mM NaCl, 2% SDS, 2% sodium Deoxycholate, PIC 2X) were added to each tube, samples were mixed thoroughly and incubated at 37°C for 30 min. After the release incubation, beads were removed and tubes were pooled, diluted 4 times in IP dilution buffer (10 mM Tris-HCl ph7.5, 140 mM NaCl, 0,1% Na-deoxycholate, 1% Triton X100, 1 mM EDTA, 1X PIC), and concentrated using a 50Kda cutoff Centricon (Amicon). The collected material was then diluted 16 times in IP dilution buffer and incubated over-night at 4°C with 5 μl of anti-H3K4me2 (Millipore). The next day, 50 μl of protein G coupled to magnetic beads (Dynabeads Protein G, Invitrogen) were added and the incubation continued for 2 hours. The beads were then washed 3 times with RIPA buffer and once with TE buffer. The protein-DNA complexes were eluted from the beads with 150 μl of Elution Buffer at 68 °C for 2 hours. DNA was purified using MinElute PCR purification kit (Qiagen).
ChIP libraries were prepared using bar-coded adapters according to standard Illumina library preparation protocols, following the manufactorer’s instructions for the NEBNext® Ultra™ DNA Library Prep Kit for Illumina®, or the TruSeq® ChIP sample preparation for Illumina with and without gel-size selection (for the protocol used, see “library protocol” annotation of each sample). The Quality of the libraries and size distribution was assessed on an Agilent 2100 Bioanalyzer (Agilent Technologies). One to four samples with different barcodes were mixed at equimolar ratios per pool. Sequencing was performed on an Illumina HiSeq 2500 machine (50 bp read length, single-end) according to Illumina standards.

value: H3K27me3 (Millipore, Cat. # 07-449) and H3K4me2 (Millipore, Cat. # 07-030)

value: NEBNextUltra-Illumina


title: ChIPseq_E10.5-Mx_H3K27me3-H3K4me2_1

value: Mx


title: 5FU_Day0_10


title: ChIPseq_E10.5-PA2_H3K27ac_1

value: H3K27ac (abcam, ab4729)


title: ChIPseq_E10.5-PA2_H3K27me3-H3K4me2_1


title: ChIPseq_E10.5-Md_H3K27me3-H3K4me2_1


title: ChIPseq_E10.5-Mx_H3K27ac_1


title: ChIPseq_E10.5-FNP_H3K27ac_1


title: ATACseq_NCCprogenitors_2


title: ChIPseq_E10.5-PA2_H3K27me3_1


title: ChIPseq_E10.5-Md_H3K27me3_1


title: ChIPseq_E10.5-Mx_H3K27me3_1


title: ChIPseq_E10.5-FNP_H3K27me3_1


title: ChIPseq_E10.5-Md-HoxA2OE_H3K27ac_0

value: TrueSeq-Illumina, with gel-size selection


title: ChIPseq_Input-NebNextUltra_1


title: ChIPseq_NCCprogenitors_H3K4me2_1


title: ChIPseq_E10.5-Mx_H3K4me2_1


title: ChIPseq_E10.5-FNP_H3K4me2_1


title: ChIPseq_NCCprogenitors_H3K27me3_1


title: ChIPseq_E10.5-Mx-Ezh2cKO_H3K27me3_0


title: RNAseq_E11.5-PA2-Hoxa2TempMut_3

data_processing:
Illumina RTA 1.18.64 and bcl2fastq2 v2.17 was used for basecalling and demultiplexing.
Reads were aligned to the GRCm38/mm10 reference genome using the QuasR Bioconductor package (PMID: 25417205, version 1.12.0) by the qAlign function with parameters maxHits=10 (allowing up to 10 hits per read, reporting one randomly) and splicedAlignment=TRUE (spiced alignment using SpliceMap PMID: 20371516).
For each gene, alignments overlapping any of its exons (obtained from Ensemble Release 71) were counted using the QuasR qCount function with default parameters.
Genome_build: GRCm38/mm10
Supplementary_files_format_and_content: Comma-seperated text file with gene annotation (Ensembl identifier, representative transcript, coordinates, strand, gene symbol and biotype, number of exonic bases) and raw alignment counts for each gene and sample.

extract_protocol:
Embryonic tissues were micro-dissected at E8.5, E10.5 or E11.5 and incubated in trypsin 0.5%/EDTA 1X at 37 °C for 10 minutes and rinsed in DMEM 1X/ FBS 10%. Cranial neural crest cells (CNCCs) were then dissociated with a pipette, filtered and collected by FACS. After sorting, CNCCs were pelleted by centrifugation for 5 minutes, 1000 rpm at 4°C. Total RNA was extracted using PicoPure® Kit (KIT0204 Life Technologies).
For sequencing of total RNA, unstranded RNA-Seq libraries were prepared using the ScriptSeq v2 RNA-Seq Library preparation Kit. Three samples with different barcodes were mixed at equimolar ratios per pool. Sequencing was performed on an Illumina HiSeq 2500 machine (50 bp read length, single-end reads) according to Illumina standards.


title: ChIPseq_E10.5-Mx_H3K27me3_0


title: ChIPseq_E10.5-FNPMxMd_H3K4me2_1


title: RNAseq_E11.5-PA2-Hoxa2TempMut_2


title: ChIPseq_E10.5-PA2_H3K4me2_1


title: ChIPseq_E10.5-Md_H3K4me2_1


title: RNAseq_E11.5-PA2-Control_2

value: Control


title: ChIPseq_Input-ChIPseqIlluminaWithGelCut_0


title: ChIPseq_E10.5-Md_H3K27ac_1


title: RNAseq_E11.5-PA2_1


title: RNAseq_E11.5-Md_3


title: RNAseq_E11.5-Md_1


title: RNAseq_E11.5-PA2-Hoxa2TempMut_1


title: RNAseq_E11.5-PA2_3


title: RNAseq_E11.5-PA2-Control_3


title: RNAseq_E11.5-Md_2


title: RNAseq_E11.5-PA2-Control_1


title: RNAseq_E11.5-Mx_3


title: RNAseq_E11.5-Mx_2


title: HCPro mutB sample 2

city: Madrid

first: Francisco

middle: Javier

last: del Toro

state: Madrid

address: Ramiro de Maeztu, 9

department: Enviromental Biology

country: Spain

institute: Centro de Investigaciones Biologicas (CSIC))

description:
small RNAs
RNA isolated from purified samples obtained from tissues infected

data_processing: Sequence reads from the two RNA libraries were computationally processed by first removing adaptor sequences from raw reads and discarding any insert reads of less than 18 nts in length using our own Perl scripts.  Reads were then mapped using Bowtie (for reads between 18 and 49 nts) (Langmead et al., 2009) or Bowtie2 (for reads of 50 nts and higher) (Langmead et al., 2012) considering only perfect matches.  Reads mapping to plant rRNA sequences, including mitochondrial and chloroplasts ones (from NCBI database) were removed from the analysis.  The remaining reads were then compared with a) the two viral construct genomes; b) the N. benthamiana genome, and c) known micro RNA (miRNA) sequences.  The genomic sequence of PVX was that found in the pgR107 binary construct (GenBank: AY297842.1) and that of PVX-P1-6x-HCPro was obtained by inserting the sequence of P1-6x-HCPro into the PVX vector polylinker, and confirmed by Sanger sequencing.  The genome of N. benthamiana was obtained from The University of Sydney (http://sydney.edu.au/), version 0.5 (Naim et al., 2012).  For the miRNAs, we used those in the Plant Non-coding RNA Database (PNRD; Yi et al., 2015), including mature as well as pre-miRNA sequences.  Sorting of sequences by length, counting of total and unique sequences, and of percentage of nucleotide identities at RNA 5’ ends were all achieved employing different Perl scripts developed in our laboratory.

organism: Nicotiana benthamiana

source_name: Systemically infected leaves

growth_protocol: Nicotiana benthamiana plants were used in this study.  Plants were kept in controlled growth chambers at 25 °C day/20 °C night temperatures, with a 16/8 hours day/night photoperiod and ~2500 lux of daylight intensity. 4-5 week-old plants were agroinoculated with Agrobacterium tumefaciens containing the appropriate binary constructs.

extract_protocol:
Our protocol for the purification of HCPro tagged with six histidines (6x-HCPro or 6x-HCPro mutB) is similar to the one described by Tena et al. (2013) with minor modifications.  Briefly, fully developed leaves showing symptoms were collected at seven days post-agroinoculation of plants with either the PVX or with the PVX-P1-6x-HCPro vectors.  Around 100 g of systemic infected tissue were collected and kept frozen until the time of use.  Every step in the purification protocol was performed between 0º and 4º C degrees with pre-chilled buffers and laboratory equipment.  Leaves were homogenized in extraction buffer [3.3 ml of buffer/g of leaf tissue: 100 mM Tris-HCl pH 8.5, 20 mM MgSO4, 500 mM NaCl, 0.5 mM, ethyleneglycol-bis(2-aminoethyl ether)-N,N,N',N′-tetra acetic acid (EGTA), 20% sucrose, supplemented with 0.2% Na2SO3, 0.1% polyvinylpyrrolidone-40 and 5 mM 2-mercaptoethanol]. After filtration, centrifugation and ultracentrifugation, total soluble proteins were precipitated with 40% (NH4)2SO4. Proteins were resuspended in extraction buffer and incubated with 1ml of nickel-nitrilotriacetic acid agarose (Ni-NTA Agarose, Qiagen GmbH, Hilden, Germany).  Several rinses of the resin were performed with extraction buffer and after that two consecutive steps of elution were made with 4 ml of extraction buffer supplemented with 400 mM EGTA. In the first experiment ttotal RNA was extracted from 1.5 ml of the purified samples obtained from tissues infected with either PVX-P1-6x-HCPro (hereafter the HCPro sample 1) or with PVX (hereafter the control sample 1),whiler in the second experiment from 1.5 ml of the purified samples obtained from tissues infected with either PVX-P1-6x-HCPro (hereafter the HCPro sample 2) or with PVX-P1-6x-HCPro mutB (hereafter the HCPro mutB sample 2). In addition total RNAs were isolated from infected tissues from plants infected with PVX or PVX-P1-6x-HCPro to be analyzed as input (hereafter the input in PVX infected plants and the input in PVX-P1-6x-HCPro infected plants, respectively ). Total RNAs were isolated with TRIzol reagent (Invitrogen, Carlsbad, CA, USA) following the instruction of the manufacturers. Total RNAs from Input in PVX infected plant and,iInput in PVX-P1-6x-HCPro infected plants, HCPro sample 2nd experiment and HCPro mutB sample 2nd experiment were sent to Fasteris SA for deep-sequencing of the RNA population up to 50 nts in lengt . For total RNAs from purified proteins of the first experiment RNAs were size fractioned by electrophoresis in a 1.5% agarose gel.  For deep-sequencing of the RNA population of 500 nts in length or less, total RNA were fractioned by electrophoresis in 10%-PAGE containing 8M urea.  The gel area from ~12 nts to ~500 nts was sliced with a razor and RNA was eluted from the gel with 0.3 M NaCl, and precipitated with 1 ul of glycogen (20 ug/ul) (Roche, Basel, Switzerland) and 1 volume of isopropanol.
RNA sequencing was performed by Fasteris SA (Chemin du Pont-du-Centenaire, Genève, Switzerland). Briefly, RNA samples were subjected to 5' cap removing with tobacco acid pyrophosphatase (TAP) treatment and ends repair reactions, followed by single-stranded ligation of 3’ and 5’ indexed adapters.  Afterwards, reverse transcription and PCR amplification were performed to generate the two cDNA libraries (one from the control and another from the HCPro samples) that were multiplexed and sequenced in one lane of a 125 bp single-end run or in one lane of 50 bp single-end run, using Illumina HiSeq 2500 sequencer (Illumina, San Diego, California, USA).
RNA-seq 125 bp single-end run and RNA-seq 50 bp single-end run, using Illumina HiSeq 2500 sequencer (Illumina, San Diego, California, USA)

treatment_protocol: Three infectious Potato virus X (PVX) vectors were expressed from binary constructs: an “empty” PVX vector was expressed from binary construct pgR107, kindly provided by Prof. D. C. Baulcombe group (University of Cambridge, UK), which expresses an infectious PVX that contains an additional CP promoter and a polylinker for the insertion and expression of foreign genes (Lu et al., 2003).  Binary construct PVX-P1-6x-HCPro was derived from vector pgR107, modified to express the Potato virus Y (PVY) P1 protein and hexahistidine tagged HCPro cistrons, and was already described by Tena et al., (2013). Binary construct PVX-P1-6x-HCPro mutB was derived from binary construct PVX-P1-6x-HCPro, modified to contain the triplets for aa 131-132 of PVY HCPro (Asn-Leu, AAT-TTA) replaced by those of two Alanines (GCT-GCA).  This mutant is the equivalent of mutant HCL134H in Plum pox virus HCPro described by González-Jara et al., (2005)

tag: tissue

value: Systemically infected leaves

tag: virus

value: PVX-P1-6x-HCPro mutB

tag: collection day post inoculation

value: 7 dpi


title: HCPro sample 2


title: RNAseq_E11.5-Mx_1

value: Cranial neural crest cells


title: RNAseq_E11.5-FNP_3

value: WT


title: Input PVX

description:
small RNAs
Total RNA isolated from tissues infected

value: PVX


title: Input PVX-P1-6x-HCPro


title: RNAseq_E11.5-FNP_1

value: FNP


title: Control

description:
RNA of less than 500 nts
RNA isolated from purified samples obtained from tissues infected

data_processing:
Sequence reads from the two RNA libraries were computationally processed by first removing adaptor sequences from raw reads and discarding any insert reads of less than 18 nts in length using our own Perl scripts.  Reads were then mapped using Bowtie (for reads between 18 and 49 nts) (Langmead et al., 2009) or Bowtie2 (for reads of 50 nts and higher) (Langmead et al., 2012) considering only perfect matches.  Reads mapping to plant rRNA sequences, including mitochondrial and chloroplasts ones (from NCBI database) were removed from the analysis.  The remaining reads were then compared with a) the two viral construct genomes; b) the N. benthamiana genome, and c) known micro RNA (miRNA) sequences.  The genomic sequence of PVX was that found in the pgR107 binary construct (GenBank: AY297842.1) and that of PVX-P1-6x-HCPro was obtained by inserting the sequence of P1-6x-HCPro into the PVX vector polylinker, and confirmed by Sanger sequencing.  The genome of N. benthamiana was obtained from The University of Sydney (http://sydney.edu.au/), version 0.5 (Naim et al., 2012).  For the miRNAs, we used those in the Plant Non-coding RNA Database (PNRD; Yi et al., 2015), including mature as well as pre-miRNA sequences.  Sorting of sequences by length, counting of total and unique sequences, and of percentage of nucleotide identities at RNA 5’ ends were all achieved employing different Perl scripts developed in our laboratory.
Supplementary_files_format_and_content: sequence abundance


title: RNAseq_E10.5-Md-HoxA2OE_2

value: Md


title: DUX4CA_24hrs_rep1

city: Salt Lake City

first: Peter

middle: G

last: Hendrickson

state: UT

address: 2000 Circle of Hope Dr.

department: HHMI/Oncological Sciences

country: USA

institute: University of Utah

data_processing:
Raw RNA sequencing reads were aligned to a custom hg19 genome index using Novoalign (v2.8, Novocraft).
Splice junction alignments were converted to genomic coordinates and low quality and non-unique reads were removed using the SamTranscriptomeParser application (USeq package, v8.8.8).
Raw reads were counted across all individual exons and repeat elements and subsequently merged by corresponsing ensembl gene or repeat masker subfamily annotation. Differential gene and subfamily expression was called using DESeq2.
Genome_build: hg19
Supplementary_files_format_and_content: Excel file contains 2 tabs corresponding to genes and repeat elements (by subfamily annotation). Each tab contains read coverage and DESeq2 differential expression statistics at 14hrs and 24hrs post dox-induction.

organism: Homo sapiens

source_name: RNA-seq in hiPSCs

growth_protocol: hiPSCs were cultured on Matrigel in mTeSR1 supplemented with ROCK inhibitor.

extract_protocol:
RNA was extracted using TRIzol
RNA libraries were prepared using oligo-dT selection and sequenced on the Illumina HiSeq 2500 according to the manufacturer's instructions.

treatment_protocol: Cells were kept under constant drug selection (Puromycin) and induced with doxycycline for 14 or 24 hrs prior to RNA isolation.

value: human induced pluripotent stem cells

tag: transgene

tag: condition

value: 24hr dox-induction


title: Treatment


title: DUX4CA_24hrs_rep2


title: RNAseq_E11.5-FNP_2


title: RNAseq_E10.5-PA2-Ezh2cKO_2


title: RNAseq_E10.5-Md-Ezh2cKO_3


title: Luciferase_24hrs_rep1

value: Luciferase


title: RNAseq_E10.5-PA2-Ezh2cKO_3


title: RNAseq_E10.5-PA2-Ezh2cKO_1


title: Luciferase_24hrs_rep2


title: RNAseq_E10.5-Md-HoxA2OE_1


title: DUX4CA_14hrs_rep2

value: 14hr dox-induction


title: RNAseq_E10.5-Md-Ezh2cKO_1


title: DUX4CA-D4ChIP-rep2

description: Antibody: abcam124699

data_processing:
Raw read squences were trimmed to remove low quality bases and adapter sequences (Trim Galore!) before aligning to hg19 using Bowtie2 (v2.2.6) with the following parameters: (-t –q –N1 –L 25 –X 2000 –no-mixed –no-discordant) (*non-unique reads assigned to 1 location).
A read-normalized bedgraph file for each ChIP library was generated with MACS2 callpeak (-f BAMPE -B -SPMR).
Genome_build: hg19
Supplementary_files_format_and_content: The read normalized bedGraph files were  converted into bigwig format for genome browser visulization.

source_name: ChIP-seq in hiPSCs

extract_protocol:
Cells were crosslinked in 1% formaldehyde for 10 minutes. Cells were then washed in PBS (x2) and resuspended in lysis buffer (10mM Tris,100mM NaCl, 1mM EDTA, 0.5mM EGTA, 0.1% Na-Deoxycholate, 0.5% N-lauroylsarcosine) to extract chromatin. Lysates were then sonicated using a BioRuptor, centrifuged to pellet cellular debris, and immunoprecipitated.
After reversing crosslinks, libraries were prepped using the NEBnext DNA Library Prep Kit (NEB, E7370L). Adapter ligated DNA was size selected and purified using AMPure XP beads (Beckman Coulter, A63881) before sequencing.

treatment_protocol: Cells were kept under constant drug selection (Puromycin) and induced with doxycycline for 18hrs.

value: 18hr dox-induction


title: Luciferase_14hrs_rep1


title: RNAseq_E10.5-Mx-Ezh2cKO_3

value: Mx


title: RNAseq_E11.5-PA2_2


title: Luciferase_14hrs_rep2


title: DUX4CA-D4ChIP-rep1

description: Antibody: abcam124699


title: RNAseq_E10.5-Mx-Ezh2cKO_2


title: Luciferase-D4ChIP-rep2

description: Antibody: abcam124699


title: RNAseq_E10.5-Md-HoxA2OE_3


title: DUX4CA_14hrs_rep1


title: RNAseq_E10.5-FNP-Ezh2cKO_3


title: RNAseq_E10.5-Mx-Ezh2cKO_1


title: female164_FcRL4pos_Bcells

city: Stockholm

first: Daniel

last: Ramsköld

address: Nobels väg 3

country: Sweden

institute: Karolinska Institutet

description:
FcRL4+ B cells from synovial fluid from RA patient 164.
Processed data file: Expression.txt

data_processing:
Reads were aligned to the GRCh38 genome build with STAR 2.4.0g1 and gencode v21 exon junctions.
FPKM expression values were calculated using rpkmforgenes.py version 13 Mar 2015 with options -midread -readcount -rmnameoverlap.
microRNA and snoRNA genes were removed (grep -v MIR, grep -v SNOR).
Genome_build: hg38 (GRCh38)
Supplementary_files_format_and_content: Expression.txt: Tab-delimited text; FPKM values and read counts for genes.

molecule: polyA RNA

source_name: Synovial fluid,  FcRL4+ B cells

growth_protocol: PBS supplemented with 10% BSA/ 5 mM EDTA (maintenance).

extract_protocol:
PicoPure RNA extraction kit (ThermoFisher).
SMARTer amplification using olig(dT) primed reactions (Clontech cat# 635001) followed by Illumina TruSeq Stranded prep kit.

tag: disease status

value: rheumatoid arthritis

value: synovial fluid

value: CD19+/FcRL4+ B cells

tag: Sex

value: Female

tag: individual id


title: female167_FcRL4pos_Bcells

description:
FcRL4+ B cells from synovial fluid from RA patient 167.
Processed data file: Expression.txt


title: RNAseq_E10.5-FNP-Ezh2cKO_2


title: RNAseq_E10.5-Md-Ezh2cKO_2


title: female162_FcRL4pos_Bcells

description:
FcRL4+ B cells from synovial fluid from RA patient 162.
Processed data file: Expression.txt


title: RNAseq_E10.5-PA2_3


title: male118_FcRL4pos_Bcells

description:
FcRL4+ B cells from synovial fluid from RA patient 118.
Processed data file: Expression.txt

value: Male


title: female164_FcRL4neg_Bcells

description:
FcRL4- B cells from synovial fluid from RA patient 164.
Processed data file: Expression.txt

source_name: Synovial fluid,  FcRL4- B cells

value: CD19+/FcRL4- B cells


title: RNAseq_E10.5-FNP-Ezh2cKO_1

value: Cranial neural crest cells


title: female162_FcRL4neg_Bcells

description:
FcRL4- B cells from synovial fluid from RA patient 162.
Processed data file: Expression.txt


title: female167_FcRL4neg_Bcells

description:
FcRL4- B cells from synovial fluid from RA patient 167.
Processed data file: Expression.txt


title: RNAseq_E10.5-PA2_2

value: WT


title: RNAseq_E10.5-Md_3


title: RNAseq_E10.5-Md_2


title: RNAseq_E10.5-PA2_1


title: RNAseq_E10.5-Md_1


title: male118_FcRL4neg_Bcells

description:
FcRL4- B cells from synovial fluid from RA patient 118.
Processed data file: Expression.txt


title: RNAseq_E10.5-Mx_2


title: RNAseq_E10.5-Mx_1


title: RNAseq_E10.5-FNP_3


title: RNAseq_E10.5-FNP_2


title: Luciferase-D4ChIP-rep1

description: Antibody: abcam124699


title: differentiation_RNASeq.32.1

city: Bethesda

first: Bruce

middle: Hamilton

last: Howard

state: MD

address: 9000 Rockville Pike

department: Intramural Program

institute: NIH/NICHD

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 116
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 150625_ROSIE_C75APANXX.7.10316646.bam

data_processing:
ChIP alignment: sequence reads were mapped to the human grch37 genome using Bowtie or Bowtie2 with default settings.
ChIP genome-wide coverage annotation: all mapped reads were written to znt2 annotation files by incrementing the file position value by 1 over the length of the read.
RNA-seq alignment: sequence reads were mapped to the human grch37 genome using STAR with default settings.
RNA-seq quantification was done using htseq and edgeR with default settings. FDR values were calculated with the Benjamini-Hochberg option.  Key is provided by the Sample "Title" field, where first index denotes merged data used in htseq/edgeR calculations, and second index denotes raw file replicate number (1-2 for monocytes and 1-4 for dendritic cells).
Abundance measurements (htseq) are reported in htseq*.txt files.
Differential expression (edgeR) are reported in de*.txt files.
For 'average insert size' field in paired-end experiments section median values from the bam TLEN field are provided to minimize bias from outliers related to splicing.
Genome_build: human grch37
Supplementary_files_format_and_content: ChIP-seq annotation znt2 files are binary files initialized (initially written) as short int (little endian) zeros with the length of the file in zeros equal to the number of bases in the relevant chromosome.
Supplementary_files_format_and_content: RNA-seq differentiation: differential expression files de.com.diff_cb.mono_dc.df and htseq_count.diff_cb.mono_dc.txt are derived from samples 20-31 (newborn monocyes) and samples 32-59 (newborn dendritic cells). RNA-seq differential expression files de.com.diff_ad.mono_dc.df and htseq_count.diff_ad.mono_dc.txt are derived from samples 60-75 (adult monocytes) and samples 76-119 (adult dendritic cells).
Supplementary_files_format_and_content: RNA-seq development: differential expression files de.com.dev_mono.cb_ad.df and htseq_count.dev_mono.cb_ad.txt are derived from samples 20-31 (newborn monocyes) and samples 60-71 (adult monocytes). RNA-seq differential expression files de.com.dev_dc.cb_ad.df and htseq_count.dev_dc.cb_ad.txt are derived from samples 32-59 (newborn dendritic cells) and samples 76-119 (adult dendritic cells).

source_name: peripheral blood

growth_protocol: Cord blood and adult peripheral blood samples were enriched by Ficoll-paque gradient centrifugation (density 1.077) and pelleted twice to reduce platelet contamination. After a second lower density Ficoll-paque gradient step, monocytes were further purified by subtractive magnetic cell sorting using a monocyte isolation kit (Miltenyi, Auburn, CA) following the manufacturer’s protocol.

extract_protocol:
For ChIP-seq lysates were clarified from non-fixed Dnase I-treated nuclei and histone-DNA complexes were isolated with anti-trimethyl-Histone H3 (Lys27) antibody (Millipore catalog no. 07-449). For RNA-seq total RNA was extracted from purified monocytes or dendritic cells using RNeasy Micro kit (Qiagen). RNA integrity and concentration were measured using RNA 6000 Nano kits (Agilent) on a Agilent 2100 Bioanalyzer.
ChIP-seq libraries were prepared according to Illumina's instructions.  RNA-seq libraries were prepared following the Illumina TruSeq RNA low-throughput protocol.

treatment_protocol: Dendritic cells (DC) obtained by plating in RPMI 1640, 10% FCS, 50 ng/ml GM-CSF, 20 ng/ml IL-4. Cultures were refed every other day and DC were harvested by day 6 or 7. Purities of monocyte and DC preparations were assessed by flow cytometry (FACSCalibur;BD Biosciences, San Jose, CA) with the following antibodies: anti-CD14 FITC, anti-biotin PE (Miltenyi); anti-HLA-DR, and anti-CD86 (BD Biosciences).

value: dendritic cells

tag: developmental stage

value: adult


title: differentiation_RNASeq.32.4

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 119
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 150626_EDISON_C75D4ANXX.4.10316646.bam

tag: cell type


title: differentiation_RNASeq.32.3

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 118
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 150626_EDISON_C75D4ANXX.5.10316646.bam


title: differentiation_RNASeq.32.2

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 117
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 150626_EDISON_C75D4ANXX.6.10316646.bam


title: RNAseq_E10.5-FNP_1

value: FNP


title: differentiation_RNASeq.31.4

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 115
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 150626_EDISON_C75D4ANXX.4.10316634.bam


title: differentiation_RNASeq.31.2

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 113
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 150626_EDISON_C75D4ANXX.5.10316634.bam


title: RNAseq_NCCprogenitors_3

value: NCCprogenitors


title: differentiation_RNASeq.31.3

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 114
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 150626_EDISON_C75D4ANXX.6.10316634.bam


title: RNAseq_NCCprogenitors_2


title: RNAseq_NCCprogenitors_1


title: differentiation_RNASeq.30.3

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 110
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 150626_EDISON_C75D4ANXX.5.10316640.bam


title: differentiation_RNASeq.30.1

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 108
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 150626_EDISON_C75D4ANXX.6.10316640.bam


title: differentiation_RNASeq.30.2

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 109
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 150625_ROSIE_C75APANXX.7.10316640.bam


title: differentiation_RNASeq.29.4

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 107
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 150626_EDISON_C75D4ANXX.5.10316644.bam


title: differentiation_RNASeq.29.3

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 106
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 150626_EDISON_C75D4ANXX.4.10316644.bam


title: Gcgr mAb-treated Gcgr+/+ 3

city: Nashville

first: Danielle

last: Dean

state: TN

address: 7465 MRBIV, 2215 Garland Avenue

department: Division of Diabetes, Endocrinology, and Metabolism

institute: Vanderbilt University Medical Center

description:
processed_data_column_header: 2552-ACP-0009
Normalized expression values_all_2552ACP

data_processing:
Post-processing of the sequencing reads from RNA-seq experiments for each sample was performed using HudsonAlpha’s unique in-house RNA-seq data analysis pipeline. Briefly, quality control checks on raw sequence data for each sample were performed using FastQC (Babraham Bioinformatics, Cambridge, UK). Raw reads were mapped to the reference mm9 using TopHat v2.0. The alignment metrics of the mapped reads were estimated using SAMtools. Aligned reads were imported to the commercial data analysis platform AvadisNGS (Strand Scientifics, CA, USA). After quality inspection, the aligned reads were filtered on the basis of read quality metrics; reads with a base quality score of less than 30, alignment score of less than 95, and mapping quality of less than 40 were removed. Remaining reads were then filtered on the basis of their read statistics; missing mates, translocated, unaligned, and flipped reads were removed. The reads list was then filtered to remove duplicates. Samples were grouped and transcript abundance was quantified for this final read list using Trimmed Means of M-values (TMM) as the normalization method. Output data utilized for all subsequent comparisons were summarized as normalized signal values generated by AvadisNGS. Differential expression of genes was calculated on the basis of fold changes (using the default cut-off ≥ ±2.0) observed in comparisons between defined conditions, and the p-value of the differentially expressed gene list was estimated by ANOVA using Benjamini Hochberg corrections of 0.05 for false-discovery rate.
Genome_build: mm9
Supplementary_files_format_and_content: Excel files include RPKM values for each Sample

source_name: liver

growth_protocol: Adult (16-20 weeks) male mice after 6 hr fast

extract_protocol:
Qiagen Rneasy Mini
The concentration and integrity of the extracted total RNA were estimated using the Qubit® 2.0 Fluorometer (Invitrogen)and Agilent 2100 Bioanalyzer (Applied Biosystems, Carlsbad, CA, USA), respectively. Five hundred nanograms of total RNA was required for downstream RNA-seq applications. Polyadenylated RNAs were isolated using NEBNext Magnetic Oligo d(T)25 Beads. The NEBNext mRNA Library Prep Reagent Set for Illumina (New England BioLabs Inc., Ipswich, MA, USA) was then used to prepare individually bar-coded next-generation sequencing expression libraries as per manufacturer's recommended protocol. Library quality was assessed using the Qubit 2.0 Fluorometer, and the library concentration was estimated by utilizing a DNA 1000 Chip on an Agilent 2100 Bioanalyzer. Accurate quantification for sequencing applications was determined using the qPCR-based KAPA Biosystems Library Quantification Kit. Each library was diluted to a final concentration of 12.5 nM and pooled in an equimolar ratio prior to clustering. Paired-end sequencing (50 million, 50-bp, paired-end reads) was performed using a 200 Cycle TruSeq SBS HS v4 Kit on an Illumina HiSeq2500 sequencer (Illumina, Inc., San Diego, CA, USA).

treatment_protocol: Treated intraperitoneally once a week for 10 days with either PBS or 10mg/kg of a humanized monoclonal antibody against the Glucagon receptor (Amgen, Inc)

tag: strain

value: liver

value: 16-20 weeks

value: Gcgr+/+

tag: treatment

value: Gcgr mAb


title: Gcgr mAb-treated Gcgr+/+ 2

description:
processed_data_column_header: 2552-ACP-0008
Normalized expression values_all_2552ACP


title: PBS-treated Gcgr-/- 2

description:
processed_data_column_header: 2552-ACP-0005
Normalized expression values_all_2552ACP

value: Gcgr-/-

value: PBS


title: Gcgr mAb-treated Gcgr+/+ 1

description:
processed_data_column_header: 2552-ACP-0007
Normalized expression values_all_2552ACP


title: PBS-treated Gcgr-/- 3

description:
processed_data_column_header: 2552-ACP-0006
Normalized expression values_all_2552ACP


title: PBS-treated Gcgr-/- 1

description:
processed_data_column_header: 2552-ACP-0004
Normalized expression values_all_2552ACP


title: PBS-treated Gcgr+/+ 3

description:
processed_data_column_header: 2552-ACP-0003
Normalized expression values_all_2552ACP


title: PBS-treated Gcgr+/+ 2

description:
processed_data_column_header: 2552-ACP-0002
Normalized expression values_all_2552ACP


title: differentiation_RNASeq.29.1

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 104
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 150625_ROSIE_C75APANXX.7.10316644.bam


title: differentiation_RNASeq.29.2

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 105
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 150626_EDISON_C75D4ANXX.6.10316644.bam


title: differentiation_RNASeq.28.4

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 103
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 150626_EDISON_C75D4ANXX.5.10316637.bam


title: PBS-treated Gcgr+/+ 1

description:
processed_data_column_header: 2552-ACP-0001
Normalized expression values_all_2552ACP


title: differentiation_RNASeq.28.2

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 101
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 150626_EDISON_C75D4ANXX.6.10316637.bam


title: differentiation_RNASeq.28.3

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 102
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 150626_EDISON_C75D4ANXX.4.10316637.bam


title: differentiation_RNASeq.31.1

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 112
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 150625_ROSIE_C75APANXX.7.10316634.bam


title: differentiation_RNASeq.30.4

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 111
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 150626_EDISON_C75D4ANXX.4.10316640.bam


title: differentiation_RNASeq.28.1

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 100
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 150625_ROSIE_C75APANXX.7.10316637.bam


title: differentiation_RNASeq.27.2

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 97
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 150626_EDISON_C75D4ANXX.4.10316642.bam


title: Sorted-GFPpos-rep1

data_processing:
Raw RNA sequencing reads were aligned to a custom genome index using Novoalign (v2.8, Novocraft).
Splice junction alignments were converted to genomic coordinates and low quality and non-unique reads were removed using the SamTranscriptomeParser application (USeq package, v8.8.8).
Raw reads were counted across all individual exons and repeat elements and subsequently merged by corresponsing ensembl gene or repeat masker subfamily annotation. Differential gene and subfamily expression was called using DESeq2.
Genome_build: mm10
Supplementary_files_format_and_content: Excel file contains 3 tabs corresponding to each RNA-seq experiment (Transient, unsorted, and sorted).  Each tab contains read coverage and DESeq2 differential expression statistics for all Ensemble genes (mm10) and repetitive elements (by subfamily annotation).

source_name: mESCs

growth_protocol: mESCs were cultured on gelatin in ES-DMEM supplemented with serum, leukemia Inhibitory Factor, and 2i (CHIR99021, PD0325901).

extract_protocol:
RNA was extracted using TRIzol and purified using the Direct-zol RNA MiniPrep Kit by Zymo Research
Intact poly(A) RNA was purified from total RNA samples (100-500 ng) with oligo(dT) magnetic beads and stranded mRNA sequencing libraries were prepared as described using the Illumina TruSeq Stranded mRNA Library Preparation Kit (RS-122-2101, RS-122-2102).  Purified libraries were qualified on an Agilent Technologies 2200 TapeStation using a D1000 ScreenTape assay (cat# 5067-5582 and 5067-5583).  The molarity of adapter-modified molecules was defined by quantitative PCR using the Kapa Biosystems Kapa Library Quant Kit (cat#KK4824).  Individual libraries were normalized to 10 nM and equal volumes were pooled in preparation for Illumina sequence analysis. Sequencing libraries (25 pM) were chemically denatured and applied to an Illumina HiSeq v4 single- or paired-end flow cell using an Illumina cBot.  Hybridized molecules were clonally amplified and annealed to sequencing primers with reagents from an Illumina HiSeq SR Cluster Kit v4-cBot (GD-401-4001) or PE Cluster Kit v4-cBot (PE-401-4001).  Following transfer of the flowcell to an Illumina HiSeq 2500 instrument (HCS v2.2.38 and RTA v1.18.61), a 50 cycle single-read or a 125 cycle paired-end sequence run was performed using HiSeq SBS Kit v4 sequencing reagents (FC-401-4003).

treatment_protocol: Stable cells were kept under constant drug selection (Puromycin, G418) and induced with doxycycline for 24 hrs prior to nucleic acid isolation or FACS.

value: mouse embryonic stem cells (in 2i media)

value: mDuxCA

value: 24hr dox-induction, FACS-sorted


title: differentiation_RNASeq.27.4

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 99
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 150625_ROSIE_C75APANXX.7.10316642.bam


title: differentiation_RNASeq.27.3

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 98
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 150626_EDISON_C75D4ANXX.6.10316642.bam


title: Sorted-GFPpos-rep2


title: Unsorted-NoDox-rep2

value: No dox


title: Sorted-GFPneg-rep1


title: Sorted-GFPneg-rep2


title: differentiation_RNASeq.27.1

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 96
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 150626_EDISON_C75D4ANXX.5.10316642.bam


title: Unsorted-plusDox-rep2

value: 24hr dox-induction


title: Unsorted-NoDox-rep1


title: Unsorted-plusDox-rep1


title: differentiation_RNASeq.26.4

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 95
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 150626_EDISON_C75D4ANXX.5.10316632.bam


title: differentiation_RNASeq.26.3

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 94
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 150625_ROSIE_C75APANXX.7.10316632.bam


title: Transient-control-rep2

value: Transient expression


title: Transient-control-rep1


title: differentiation_RNASeq.26.2

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 93
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 150626_EDISON_C75D4ANXX.6.10316632.bam


title: Transient-mDUX-rep2


title: differentiation_RNASeq.26.1

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 92
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 150626_EDISON_C75D4ANXX.4.10316632.bam


title: differentiation_RNASeq.25.4

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 91
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 150626_EDISON_C75D4ANXX.5.10316641.bam


title: Transient-mDUX-rep1


title: Sorted-GFPneg-ATAC-rep2

description:
30k cells
mDUX-GFPneg_ATAC.bw

data_processing:
Library strategy: ATAC-Seq
Raw read squences were trimmed to remove low quality bases and adapter sequences (Trim Galore!) before aligning to mm10 using Bowtie2 (v2.2.6) with the following parameters: (-t –q –N1 –L 25 –X 2000 –no-mixed –no-discordant) (*non-unique reads assigned to 1 location).
Replicates were merged and a single, read-normalized bedgraph file was generated with MACS2 callpeak (-B -SPMR --nomodel --nolambda --shift -100 --extsize 200).
Genome_build: mm10
Supplementary_files_format_and_content: The read normalize bedGraph file was then converted into bigwig format for genome browser visulization.

source_name: mouse embryonic stem cells

growth_protocol: mESCs containing two stably integrated transgenes (TRE::3xHA-mDux; MERVL::EGFP) were cultured on gelatin in ES-DMEM supplemented with serum, leukemia Inhibitory Factor, and 2i (CHIR99021, PD0325901).

extract_protocol:
Immediately following FACS, the cells were lysed in cold lysis buffer (10 mM Tris-HCl, pH 7.4, 10 mM NaCl, 3 mM MgCl2 and 0.1% IGEPAL CA-630) and the nuclei were pelleted.
The Tn5 transposition reaction was carried out for 30 minutes at 37°C. Following DNA purification, the Nextera libraries were amplified for 12 cycles using the NEBnext PCR master mix and purified using the Qiagen PCR cleanup kit.

treatment_protocol: Cells were kept under constant drug selection (Puromycin, G418) and induced with doxycycline for 24 hrs prior to FACS.

value: 24hr dox-induction; FACS-sorted


title: differentiation_RNASeq.25.3

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 90
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 150626_EDISON_C75D4ANXX.4.10316641.bam

value: dendritic cells


title: Sorted-GFPneg-ATAC-rep1

description:
30k cells
mDUX-GFPneg_ATAC.bw


title: differentiation_RNASeq.25.2

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 89
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 150625_ROSIE_C75APANXX.7.10316641.bam

value: adult


title: differentiation_RNASeq.25.1

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 88
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 150626_EDISON_C75D4ANXX.6.10316641.bam


title: Sorted-GFPpos-ATAC-rep2

description:
30k cells
mDUX-GFPpos_ATAC.bw


title: differentiation_RNASeq.24.4

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 87
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 150626_EDISON_C75D4ANXX.4.10316633.bam


title: Sorted-GFPpos-ATAC-rep1

description:
30k cells
mDUX-GFPpos_ATAC.bw


title: differentiation_RNASeq.24.3

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 86
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 150626_EDISON_C75D4ANXX.5.10316633.bam


title: differentiation_RNASeq.24.1

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 84
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 150625_ROSIE_C75APANXX.7.10316633.bam


title: differentiation_RNASeq.23.4

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 83
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 150626_EDISON_C75D4ANXX.4.10316639.bam


title: differentiation_RNASeq.23.2

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 81
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 150625_ROSIE_C75APANXX.7.10316639.bam


title: differentiation_RNASeq.23.3

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 82
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 150626_EDISON_C75D4ANXX.5.10316639.bam


title: RP 1h_rep1

city: München

first: Caroline

last: Friedel

address: Amalienstr. 17

department: Institut für Informatik

institute: Ludwig-Maximilians-Universität München

data_processing:
Reads were mapped against the mouse genome (GRCm38) and rRNA sequences using ContextMap v2.5.2. For 4sU- and total RNA, reads were mapped in paired-end mode. For ribosome-associated mRNA, the sequence of the ~30nt ribosome-protected mRNA sequence was determined first as the consensus sequence from both 101nt read pairs and then mapped in single-end mode.
To calculate FPKM (Fragments Per Kilobase Of Exon Per Million Fragments Mapped) values, fragment counts per gene were calculated using the featureCounts program from the subread package (v1.4.6-p3). For  4sU- and total RNA, fragment counts were determined for all exons. For ribosome-associated mRNA, only fragments mapped to the coding sequence was were counted.
Genome_build: GRCm38
Supplementary_files_format_and_content: [.txt] FPKM values

source_name: RP 1h

growth_protocol: Peripheral CD4+ T cells were isolated with CD4 Dynabeads (Invitrogen) and kept in RPMI medium (supplemented with 10% (vol/vol) FCS, β-mercaptoethanol (0.05 mM, Gibco), penicillin-streptomycin (100 U/ml, Gibco), Na-Pyrovat (1 mM, Lonza), Non-Essential Amino Acids (1x, Gibco), MEM Vitamin Solution (1x, Gibco), Glutamax (1x, Gibco) and HEPES pH 7.2 (10 mM, Gibco)).

extract_protocol:
RNA was isolated using QIAzol Lysis Reagent. Sample preparation for 4sU-RNA and total RNA was performed as described before (Rädle et al., 2013) with minor changes. In step 5 we eluted the newly transcribed RNA into 400 µl Agencourt RNAClean XP Beads (Beckman Coulter) and continued with the recovery following the manufacturer’s instructions. RNA was eluted in 11 µl nuclease-free H2O and quantified using Qubit 2.0 Fluorometer. Samples for ribosome profiling were prepared according to manufacturer’s instructions (Illumina, TrueSeq Ribo Profile, Mammalian). At least 60 x 10e6 Th1 cells were used for each time point.
For both total and 4sU-RNA samples, library preparation and rRNA depletion was performed using the TruSeq® Stranded Total RNA Library Prep Kit (Illumina) starting with 400 ng RNA as input for each sample. Only 11 cycles were used for PCR amplification to minimize PCR bias. Libraries for ribosome profiling were prepared according to manufacturer’s instructions (Illumina, TrueSeq Ribo Profile, Mammalian) except that we used the whole amount of RNA for both gel purifications steps. Overnight gel extraction was used at every stage of the procedure. Depletion of rRNA was performed with Ribo-Zero Gold rRNA Removal Kit (Illumina, Human/Mouse/Rat) and RNA purification was done with RNA clean and concentrator kit according to the manufacurer’s advice (Zymo Research).

treatment_protocol: Th1 cells were restimulated with plate-bound goat anti–hamster immunoglobulin G and soluble anti-CD3 (1 μg/ml) and anti-CD28 (2.5 μg/ml). Metabolic labeling of newly transcribed RNA with 4sU (Carbosynth, NT0618690) was performed as described before (Rädle et al., 2013) with minor changes. 500 µM 4sU was used for 0.5h of labeling and 200µM 4sU was used for 1h labeling. Samples were collected at the indicated time points.

value: Th1 effector cell (CD4+ T cell)

tag: protocol

value: ribosome profiling

tag: time point

tag: molecule subtype

value: ribosome-associated mRNA


title: RP 4h_rep1

source_name: RP 4h


title: RP 0h_rep2

source_name: RP 0h


title: RP 1h_rep2


title: RP 4h_rep2


title: RP 0.5h_rep1

source_name: RP 0.5h


title: RP 0h_rep1


title: differentiation_RNASeq.22.4

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 79
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 150626_EDISON_C75D4ANXX.6.10316638.bam

tag: chip antibody


title: differentiation_RNASeq.22.1

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 76
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 150626_EDISON_C75D4ANXX.5.10316638.bam


title: differentiation_RNASeq.22.3

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 78
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 150625_ROSIE_C75APANXX.7.10316638.bam


title: RP 2h_rep1

source_name: RP 2h


title: differentiation_RNASeq.21.2

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 75
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 140909_OPTIMUS_C5LCPANXX.7.7589007.bam

value: monocytes


title: Total 4h_rep2

source_name: Total 4h

value: total RNA


title: differentiation_RNASeq.22.2

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 77
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 150626_EDISON_C75D4ANXX.4.10316638.bam


title: NT 0.5h_30min labeling

source_name: NT 0.5h_30min labeling

value: metabolic labeling of newly transcribed RNA with 4sU (0.5h of labeling)

value: 4sU RNA


title: Total 1h_rep2

source_name: Total 1h


title: NT 0h_30min labeling

source_name: NT 0h_30min labeling


title: Total 0h_rep2

source_name: Total 0h


title: differentiation_RNASeq.20.1

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 72
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 140909_OPTIMUS_C5LCPANXX.7.7588991.bam


title: differentiation_RNASeq.24.2

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 85
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 150626_EDISON_C75D4ANXX.6.10316633.bam


title: differentiation_RNASeq.21.1

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 74
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 140916_OPTIMUS_C5L92ANXX.1.7589007.bam


title: NT 1h_rep2

source_name: NT 1h

value: metabolic labeling of newly transcribed RNA with 4sU


title: differentiation_RNASeq.19.1

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 70
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 140909_OPTIMUS_C5LCPANXX.8.7588999.bam


title: differentiation_RNASeq.23.1

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 80
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 150626_EDISON_C75D4ANXX.6.10316639.bam


title: NT 0h_rep2

source_name: NT 0h


title: Total 4h_rep1


title: differentiation_RNASeq.19.2

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 71
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 140916_OPTIMUS_C5L92ANXX.2.7588999.bam


title: differentiation_RNASeq.18.2

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 69
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 140916_OPTIMUS_C5L92ANXX.1.7589002.bam


title: differentiation_RNASeq.18.1

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 68
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 140909_OPTIMUS_C5LCPANXX.7.7589002.bam


title: Total 2h_rep1

source_name: Total 2h


title: NT 4h_rep1

source_name: NT 4h


title: differentiation_RNASeq.17.2

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 67
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 140909_OPTIMUS_C5LCPANXX.7.7589000.bam

source_name: peripheral blood


title: Total 0h_rep1


title: differentiation_RNASeq.17.1

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 66
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 140916_OPTIMUS_C5L92ANXX.1.7589000.bam


title: NT 2h_rep1

source_name: NT 2h


title: NT 4h_rep2


title: NT 0h_rep1


title: NT 3h_rep1

source_name: NT 3h


title: NT 1h_rep1


title: differentiation_RNASeq.15.2

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 63
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 140909_OPTIMUS_C5LCPANXX.8.7588995.bam


title: differentiation_RNASeq.16.1

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 64
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 140916_OPTIMUS_C5L92ANXX.1.7588992.bam


title: differentiation_RNASeq.20.2

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 73
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 140916_OPTIMUS_C5L92ANXX.1.7588991.bam


title: differentiation_RNASeq.13.4

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 59
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 150626_EDISON_C75D4ANXX.5.10316648.bam

source_name: cord blood

value: newborn


title: differentiation_RNASeq.13.3

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 58
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 150626_EDISON_C75D4ANXX.6.10316648.bam


title: differentiation_RNASeq.15.1

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 62
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 140916_OPTIMUS_C5L92ANXX.2.7588995.bam


title: differentiation_RNASeq.14.1

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 60
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 140916_OPTIMUS_C5L92ANXX.2.7589004.bam


title: differentiation_RNASeq.13.1

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 56
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 150626_EDISON_C75D4ANXX.4.10316648.bam


title: differentiation_RNASeq.13.2

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 57
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 150625_ROSIE_C75APANXX.7.10316648.bam


title: differentiation_RNASeq.12.4

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 55
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 150626_EDISON_C75D4ANXX.5.10316636.bam


title: differentiation_RNASeq.12.3

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 54
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 150625_ROSIE_C75APANXX.7.10316636.bam


title: differentiation_RNASeq.12.1

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 52
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 150626_EDISON_C75D4ANXX.4.10316636.bam


title: differentiation_RNASeq.11.4

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 51
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 150626_EDISON_C75D4ANXX.5.10316631.bam


title: differentiation_RNASeq.11.2

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 49
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 150625_ROSIE_C75APANXX.7.10316631.bam


title: differentiation_RNASeq.11.1

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 48
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 150626_EDISON_C75D4ANXX.6.10316631.bam


title: differentiation_RNASeq.16.2

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 65
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 140909_OPTIMUS_C5LCPANXX.7.7588992.bam


title: differentiation_RNASeq.14.2

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 61
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 140909_OPTIMUS_C5LCPANXX.8.7589004.bam


title: differentiation_RNASeq.10.4

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 47
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 150626_EDISON_C75D4ANXX.5.10316643.bam


title: differentiation_RNASeq.10.3

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 46
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 150626_EDISON_C75D4ANXX.6.10316643.bam


title: differentiation_RNASeq.8.1

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 36
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 150626_EDISON_C75D4ANXX.4.10316645.bam


title: differentiation_RNASeq.9.3

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 42
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 150625_ROSIE_C75APANXX.7.10316647.bam


title: differentiation_RNASeq.10.1

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 44
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 150626_EDISON_C75D4ANXX.4.10316643.bam


title: differentiation_RNASeq.10.2

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 45
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 150625_ROSIE_C75APANXX.7.10316643.bam


title: differentiation_RNASeq.7.3

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 34
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 150626_EDISON_C75D4ANXX.4.10316635.bam


title: differentiation_RNASeq.7.2

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 33
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 150625_ROSIE_C75APANXX.7.10316635.bam


title: differentiation_RNASeq.8.4

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 39
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 150625_ROSIE_C75APANXX.7.10316645.bam


title: differentiation_RNASeq.9.4

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 43
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 150626_EDISON_C75D4ANXX.5.10316647.bam


title: differentiation_RNASeq.9.1

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 40
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 150626_EDISON_C75D4ANXX.4.10316647.bam


title: differentiation_RNASeq.8.2

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 37
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 150626_EDISON_C75D4ANXX.6.10316645.bam


title: differentiation_RNASeq.6.1

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 30
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 140916_OPTIMUS_C5L92ANXX.2.7589006.bam


title: differentiation_RNASeq.4.2

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 27
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 140916_OPTIMUS_C5L92ANXX.2.7588994.bam


title: differentiation_RNASeq.5.1

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 28
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 140916_OPTIMUS_C5L92ANXX.2.7588997.bam


title: differentiation_RNASeq.5.2

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 29
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 140909_OPTIMUS_C5LCPANXX.8.7588997.bam


title: differentiation_RNASeq.8.3

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 38
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 150626_EDISON_C75D4ANXX.5.10316645.bam


title: differentiation_RNASeq.4.1

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 26
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 140909_OPTIMUS_C5LCPANXX.8.7588994.bam


title: differentiation_RNASeq.12.2

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 53
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 150626_EDISON_C75D4ANXX.6.10316636.bam


title: cattle-heart rep3

city: Wen’jiang,Chengu

first: long

middle: ke

last: ren

state: Sichuan

address: Huimin road, 211#

department: 1	Institute of Animal Genetics & Breeding

country: China

institute: sichuan agriculture university

description: small RNA

data_processing:
Illumina Casava1.7 software used for basecalling.
Sequenced reads were trimmed for adaptor sequence, and masked for low-complexity or low-quality sequence, then mapped to mm8 whole genome using bowtie v0.12.2 with parameters -q -p 4 -e 100 -y -a -m 10 --best --strata
Reads Per Megabase of library size  were calculated using a protocol from Chepelev et al., Nucleic Acids Research, 2009.
Supplementary_files_format_and_content: raw counts

organism: Bos taurus

source_name: heart

extract_protocol:
Three unrelated 2-year old adult females for both of yaks and cattle were used in this study. Two of significant hypoxia-responsive tissues (heart and lung) were rapidly separated from each carcass, washed three times with physiological saline, immediately frozen in liquid nitrogen, and stored at − 80 °C until RNA extraction.
small RNA libraries were prepared for sequencing using standard Illumina protocols

value: heart

tag: age


title: cattle-heart rep2

description: small RNA

tag: tissue


title: differentiation_RNASeq.3.1

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 24
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 140909_OPTIMUS_C5LCPANXX.8.7589003.bam


title: cattle-heart rep1

description: small RNA


title: differentiation_RNASeq.3.2

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 25
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 140916_OPTIMUS_C5L92ANXX.2.7589003.bam


title: cattle-lung rep3

description: small RNA

source_name: lung

value: lung


title: differentiation_RNASeq.2.2

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 23
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 140909_OPTIMUS_C5LCPANXX.8.7588993.bam


title: differentiation_RNASeq.6.2

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 31
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 140909_OPTIMUS_C5LCPANXX.8.7589006.bam


title: differentiation_RNASeq.2.1

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 22
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 140916_OPTIMUS_C5L92ANXX.2.7588993.bam


title: cattle-lung rep2

description: small RNA


title: differentiation_RNASeq.1.1

description:
abundance measurements (htseq) htseq*.txt files
differential expression (edgeR) de*.txt files
sample 20
See Keys_samples20-119.xls
See Keys_samples20-119.xls, raw file: 140909_OPTIMUS_C5LCPANXX.8.7589005.bam


title: Endoderm_H3K4me3_Input_rep1 (Group7)

city: Manchester

first: Angela

last: Simeone

address: 200 Hathersage Rd

institute: Genomic Services QIAGEN

description: Processed data file: Eva_HiSeq1500_2.Index15_vs_Eva_HiSeq1500_2.Index13_broad_peaks.bed

data_processing:
Fastq files processed by filtering out low-quality reads (<Q20) and low-quality bases (<Q20) with sickle. Adaptors were removed with cutadapt 1.0.
RNA-seq data have been aligned to X. laevis genome 6.1 using TopHat 2.0.6.
ChIP-seq data have been aligned to X. laevis genome 6.1 using BWA 0.6.2 with default options. Duplicate reads were then removed.
Peaks were called using MACS2 2.0.9 with --broad options and q-value<0.01.
Differential analysis was conducted using EdgeR package.
The 553,960 assembled transcripts were provided by the International Xenopus Genome Project (http://www.marcottelab.org/index.php/Xenopus_Genome_Project) in October 2012. This assembly was augmented with Xenopus laevis sequences from the NCBI RefSeq database downloaded in February 2012 (30,611 sequences). The combined transcript sequences were filtered with cd-hit-est 4.5.7 (Li and Godzik, 2006) with a similarity score of 95% to remove redundant sequences. This resulted in a final set of 39,384 transcripts. To provide gene names, orthologs were found against the M. musculus proteome (downloaded in January 2013 - NCBI RefSeq) using Inparanoid 4 (Alexeyenko et al. 2006) on predicted ORFs from the Trinity Suite (Grabherr et al. 2011). The sequences were further annotated using InterProScan 4.8 (Zdobnov and Apweiler, 2001) to provide both InterPro Domains (Release 35) and Panther 7.2 ontology terms (Thomas et al. 2003). Xenopus laevis NCBI Descriptions were provided for transcripts that originated at the NCBI. The gtf file containing this information is reported as supplementary material of the previous study: PMID: 27034506, DOI: 10.1101/gr.201541.115.
Genome_build: X. laevis genome 6.1 (ftp://ftp.xenbase.org/pub/Genomics/JGI/Xenla6.1/)
Supplementary_files_format_and_content: *.xls: Excel files contain raw counts.
Supplementary_files_format_and_content: *.bed: BED files contain coordinates of broad peaks called with MACS2 (2.0.9).

organism: Xenopus laevis

source_name: Endoderm, input

extract_protocol:
ChIP and ChIP-seq preparation: Chromatin Immunoprecipitation (ChIP) was performed as described. Neurula (stage 18) embryos were generated by in vitro fertilization. For each ChIP experiment, 50 embryos were dissected in 1xMBS to obtain the endoderm tissue. Samples were fixed in 2 ml of 1% Formaldehyde in 0.1x MMR for 25 min at room temperature, followed by 4 washes with 1 mL 0.1xMMR and equilibration in 500 μl HEG solution (50mM Hepes-KOH pH 7.5, 1 mM EDTA, 20% Glycerol) at 4°C, then excess buffer was removed and samples were frozen at -80°C. To extract chromatin, the samples were homogenized in 2 ml buffer E1 (50mM Hepes-KOH pH 7.5, 140mM NaCl, 1mM EDTA pH 8.0, 10% Glycerol, 0.5% Igepal CA-630, 0.25% Triton X-100, 1mM DTT, complete protease inhibitors (Roche)). Chromatin was collected by centrifugation for 2 min at 3500 rmp, 4°C and then washed two times with 2 ml E1, three times with 2ml buffer E2 (10mM Tris pH 8.0, 200mM NaCl, 1mM EDTA, 0.5mM EGTA, complete protease inhibitors (Roche)) and three times with 500 μl Buffer E3 (10mM Tris pH 8.0, 200mM NaCl, 1mM EDTA, 0.5mM EGTA, 0.1% Na-deoxycholate, 0.5% N-lauroylsarcosine, complete protease inhibitors (Roche)). Chromatin was fragmented by sonication for 20 cycles (30s on and 30s off) using a Bioruptor (Diagenode). The samples were centrifuged at 15min, 4°C at full speed, the supernatant was collected and Triton X-100 was added to 1%. 25 μl of the solution were put aside to serve as Input for later analysis. Before ChIP, primary anti-H3K4me3 (Abcam ab8580, 0.5 μg per 50 embryos) antibodies were bound to PBS washed magnetic beads conjugated with secondary antibody (Invitrogen 11204D, 25 μl per 50 embryos) in 500 μl 1xPBS 0.1% BSA overnight at 4°C on a rotating wheel. Beads were washed 3 times with 1xPBS 0.1% BSA, added to the fragmented chromatin solution and incubated overnight at 4°C on a rotating wheel. Beads were then washed 6 times with RIPA buffer (50mM Hepes-KOH pH 7.5, 500mM LiCl, 1mM EDTA, 1% Igepal CA-630, 0.7% Na-deoxycholate, complete protease inhibitors (Roche)) and twice with TEN buffer (10mM Tris pH 8.0, 1mM EDTA, 150mM NaCl, complete protease inhibitors (Roche)) for each 10 min. For crosslink reversal, the beads were resuspended in 150 μl Stop buffer (40mM Tris pH 8.0, 10mM EDTA, 1% SDS) and 125 μl Stop buffer was added to the input fraction. The samples were supplemented with Proteinase K (0.3 μg/μl), NaCl (250 mM) and incubated at 65°C overnight. RNase A (DNase free) was added to a final concentration of 200 μg/μl and DNA was Phenol/Chloroform extracted. 150 μg/μl Glycogen was added and DNA was recovered by Ethanol precipitation. The pellet was resuspended in 30 μl H2O, and half was subjected for ChIP-seq library preparation with the TruSeq DNA kit (Illumina, FC-121-2001). Two independent biological replicates were generated for each H3K4me3 ChIP experiment.
Library construction: ChIP-seq library preparation was performed using the TruSeq DNA kit (Illumina, FC-121-2001) according to the manufacturer's protocol.

tag: starting material

value: 100 embryos

tag: developmental stage/condition

value: stage 18-21 embryo

value: endoderm

tag: pairing of samples