title: No Bacteria_plusP_100uM_exp2_rep3 [SC0351]

description: No Bacteria_plusP_100uM

value: TCTCGCG-CAGGACG


title: I2I3_minusP_100uM_exp1_rep1 [SC034]

status: Public on Aug 17 2017

type: SRA

city: Stanford

first: Sur

last: Herrera Paredes

state: CA

address: 371 Serra Mall,  Gilbert Building Room 109

department: Biology

country: USA

institute: Stanford University

library_source: transcriptomic

data_processing:
Illumina Casava version 1.8 was used for basecalling
Quality assessment of the sequencing reads was performed using the FASTX-Toolkit version 0.0.13.2
cutadapt version 1.3 was used to identify and discard reads containing Illumina adapter sequences
Quality-filtered reads were mapped to the Arabidopsis thaliana TAIR10 genome using Tophat version 2.1.1 with parameters set to allow only one mismatch (-N 1)
The Python package HTSeq version 0.6.1p1 was used to count reads that mapped to each one of the 27,206 nuclear protein-coding genes
Genome_build: TAIR10
Supplementary_files_format_and_content: tab-delimited text file including read counts per gene for each sample

molecule: total RNA

organism: Arabidopsis thaliana

growth_protocol: Arabidopsis wild-type Col-0 seeds were germinated in axenic condition on Johnson medium plus 0.5% sucrose with either 1 mM Pi or ~5 µM Pi [traces of Pi from the agar, (Difco)] in a vertical position for 7 days; then transferred (10 seedlings per plate) to 30 µM Pi and 100 µM Pi media (without sucrose) alone or with one of the bacterial synthetic communities at 10^5 c.f.u / mL of medium, for another 7 days. Plants were grown in a growth chamber in a 16-h light/8-h dark regime (24°C/21°C).

extract_protocol:
Total RNA was extracted from Arabidopsis seedlings according to Logemann, et al. (PMID: 2441623). Frozen seedlings were pulverized in liquid nitrogen. Samples were homogenized in 400 µl of Z6-buffer; 8 M guanidinium-HCl, 20 mM MES, 20 mM EDTA pH 7.0. Following the addition of 400 µl phenol:chloroform:isoamylalcohol; 25:24:1, samples were vortexed and centrifuged (20000 g, 10 min) for phase separation. The aqueous phase was transferred to a new 1.5 ml tube and 0.05 volumes of 1N acetic acid and 0.7 volumes 96 % ethanol were added. The RNA was precipitated at -20 °C overnight. Following centrifugation, (20,000 g, 10 min, 4 °C) the pellet was washed with 200 µl sodium-acetate (pH 5.2) and 70 % ethanol. The RNA was dried, and dissolved in 30 µl of ultrapure water and stored at -80 °C until use.
poly(A) RNA was isolated from 1000 ng total RNA and used to prepare non-stranded Illumina RNA-seq libraries. Following library preparation, quality control and quantification were performed using a 2100 Bioanalyzer instrument (Agilent) and the Quant-iT PicoGreen dsDNA Reagent (Invitrogen), respectively.

tag: ecotype background

tag: bacteria

tag: phosphate

tag: tissue

value: whole seedling

tag: replicate

tag: experiment

tag: extraction

tag: plate

tag: well

tag: index

value: ATTCAGA-ATAGAGG

tag: sample


title: No Bacteria_plusP_100uM_exp2_rep1 [SC0349]

description: No Bacteria_plusP_100uM

source_name: No Bacteria_plusP_100uM_exp2

value: No Bacteria

value: TCTCGCG-AGGCGAA


title: P1I1_plusP_100uM_exp2_rep2 [SC0353]

value: AGCGATA-TATAGCC


title: P2N3_minusP_30uM_exp1_rep1 [SC0364]

value: TAATGCG-GTACTGA


title: P1I1_plusP_30uM_exp2_rep1 [SC0340]

value: TCCGCGA-GGCTCTG


title: P1I1_plusP_30uM_exp2_rep2 [SC0341]

value: TCCGCGA-AGGCGAA


title: P1P3_plusP_100uM_exp2_rep2 [SC0359]

value: AGCGATA-CAGGACG


title: P1P3_plusP_30uM_exp2_rep1 [SC0346]

value: TCTCGCG-ATAGAGG


title: No Bacteria_plusP_30uM_exp2_rep2 [SC0338]

description: No Bacteria_plusP_30uM

source_name: No Bacteria_plusP_30uM_exp2

value: TCCGCGA-ATAGAGG


title: No Bacteria_plusP_30uM_exp2_rep1 [SC0337]

description: No Bacteria_plusP_30uM

value: TCCGCGA-TATAGCC


title: P2I1_plusP_30uM_exp2_rep2 [SC0344]

value: TCCGCGA-GTACTGA


title: P2I1_plusP_30uM_exp2_rep1 [SC0343]

value: TCCGCGA-CAGGACG


title: P2I1_plusP_30uM_exp2_rep3 [SC0345]

value: TCTCGCG-TATAGCC


title: No Bacteria_plusP_30uM_exp2_rep3 [SC0339]

description: No Bacteria_plusP_30uM

value: TCCGCGA-CCTATCC


title: P1I1_plusP_30uM_exp2_rep3 [SC0342]

value: TCCGCGA-TAATCTT


title: P1P3_plusP_30uM_exp2_rep3 [SC0348]

value: whole seedling

value: TCTCGCG-GGCTCTG


title: P1P3_plusP_30uM_exp2_rep2 [SC0347]

value: TCTCGCG-CCTATCC


title: P1P3_minusP_100uM_exp2_rep1 [SC0334]

value: CGGCTAT-TAATCTT


title: P2I1_minusP_100uM_exp2_rep3 [SC0333]

value: CGGCTAT-AGGCGAA


title: P1P3_minusP_100uM_exp2_rep3 [SC0336]

value: CGGCTAT-GTACTGA


title: I1I2_minusP_100uM_exp1_rep3 [SC033]

value: ATTCAGA-TATAGCC


title: P1P3_minusP_100uM_exp2_rep2 [SC0335]

value: CGGCTAT-CAGGACG


title: P2I1_minusP_100uM_exp2_rep1 [SC0331]

value: CGGCTAT-CCTATCC


title: P2I1_minusP_100uM_exp2_rep2 [SC0332]

value: CGGCTAT-GGCTCTG


title: P1I1_minusP_100uM_exp2_rep3 [SC0330]

value: CGGCTAT-ATAGAGG


title: I1I2_minusP_100uM_exp1_rep2 [SC032]

value: GAGATTC-GTACTGA


title: No Bacteria_minusP_100uM_exp2_rep1 [SC0325]

description: No Bacteria_minusP_100uM

source_name: No Bacteria_minusP_100uM_exp2

value: No Bacteria

value: TAATGCG-AGGCGAA


title: No Bacteria_minusP_100uM_exp2_rep2 [SC0326]

description: No Bacteria_minusP_100uM

value: TAATGCG-TAATCTT


title: P1I1_minusP_100uM_exp2_rep1 [SC0328]

value: TAATGCG-GTACTGA


title: No Bacteria_minusP_100uM_exp2_rep3 [SC0327]

description: No Bacteria_minusP_100uM

value: TAATGCG-CAGGACG


title: P1P3_minusP_30uM_exp2_rep1 [SC0322]

value: whole seedling

value: TAATGCG-ATAGAGG


title: P1P3_minusP_30uM_exp2_rep2 [SC0323]

value: TAATGCG-CCTATCC


title: P1P3_minusP_30uM_exp2_rep3 [SC0324]

value: TAATGCG-GGCTCTG


title: P1I1_minusP_30uM_exp2_rep3 [SC0318]

value: CTGAAGC-TAATCTT


title: P2I1_minusP_30uM_exp2_rep2 [SC0320]

value: CTGAAGC-GTACTGA


title: P2I1_minusP_30uM_exp2_rep3 [SC0321]

value: TAATGCG-TATAGCC


title: P2I1_minusP_30uM_exp2_rep1 [SC0319]

value: CTGAAGC-CAGGACG


title: No Bacteria_minusP_30uM_exp2_rep3 [SC0315]

description: No Bacteria_minusP_30uM

source_name: No Bacteria_minusP_30uM_exp2

value: CTGAAGC-CCTATCC


title: P1I1_minusP_30uM_exp2_rep1 [SC0316]

value: CTGAAGC-GGCTCTG


title: No Bacteria_minusP_30uM_exp2_rep2 [SC0314]

description: No Bacteria_minusP_30uM

value: CTGAAGC-ATAGAGG


title: I1I2_minusP_100uM_exp1_rep1 [SC031]

value: GAGATTC-CAGGACG


title: P1P3_plusP_100uM_exp1_rep2 [SC0311]

value: GAATTCG-CAGGACG


title: P1P3_plusP_100uM_exp1_rep1 [SC0310]

value: GAATTCG-TAATCTT


title: P1I1_minusP_100uM_exp2_rep2 [SC0329]

value: CGGCTAT-TATAGCC


title: No Bacteria_minusP_30uM_exp2_rep1 [SC0313]

description: No Bacteria_minusP_30uM

value: No Bacteria

value: CTGAAGC-TATAGCC


title: P2I1_plusP_100uM_exp1_rep3 [SC0309]

value: whole seedling

value: GAATTCG-AGGCGAA


title: P1I1_minusP_30uM_exp2_rep2 [SC0317]

value: CTGAAGC-AGGCGAA


title: P2I1_plusP_100uM_exp1_rep1 [SC0307]

value: GAATTCG-CCTATCC


title: P1I1_plusP_100uM_exp1_rep2 [SC0305]

value: GAATTCG-TATAGCC


title: P1I1_plusP_100uM_exp1_rep1 [SC0304]

value: ATTCAGA-GTACTGA


title: No Bacteria_plusP_100uM_exp1_rep3 [SC0303]

description: No Bacteria_plusP_100uM

source_name: No Bacteria_plusP_100uM_exp1

value: ATTCAGA-CAGGACG


title: P2I1_plusP_100uM_exp1_rep2 [SC0308]

value: GAATTCG-GGCTCTG


title: P1P3_plusP_100uM_exp1_rep3 [SC0312]

value: GAATTCG-GTACTGA


title: No Bacteria_plusP_100uM_exp1_rep2 [SC0302]

description: No Bacteria_plusP_100uM

value: ATTCAGA-TAATCTT


title: No Bacteria_plusP_100uM_exp1_rep1 [SC0301]

description: No Bacteria_plusP_100uM

value: ATTCAGA-AGGCGAA


title: P3I1_minusP_100uM_exp1_rep2 [SC029]

value: GAGATTC-AGGCGAA


title: P1P3_plusP_30uM_exp1_rep2 [SC0299]

value: ATTCAGA-CCTATCC


title: P1I1_plusP_100uM_exp1_rep3 [SC0306]

value: GAATTCG-ATAGAGG


title: P1P3_plusP_30uM_exp1_rep3 [SC0300]

value: ATTCAGA-GGCTCTG


title: P2I1_plusP_30uM_exp1_rep3 [SC0297]

value: ATTCAGA-TATAGCC


title: P1P3_plusP_30uM_exp1_rep1 [SC0298]

value: ATTCAGA-ATAGAGG


title: P2I1_plusP_30uM_exp1_rep2 [SC0296]

value: whole seedling

value: GAGATTC-GTACTGA


title: P3I1_minusP_100uM_exp1_rep3 [SC030]

value: GAGATTC-TAATCTT


title: P1I1_plusP_30uM_exp1_rep3 [SC0294]


title: P1I1_plusP_30uM_exp1_rep2 [SC0293]


title: No Bacteria_plusP_30uM_exp1_rep2 [SC0290]

description: No Bacteria_plusP_30uM

source_name: No Bacteria_plusP_30uM_exp1

value: No Bacteria

value: GAGATTC-ATAGAGG


title: No Bacteria_plusP_30uM_exp1_rep1 [SC0289]

description: No Bacteria_plusP_30uM

value: GAGATTC-TATAGCC


title: P3I1_minusP_100uM_exp1_rep1 [SC028]

value: GAGATTC-GGCTCTG


title: P1P3_minusP_100uM_exp1_rep3 [SC0288]

value: CGCTCAT-GTACTGA


title: P2I1_minusP_100uM_exp1_rep3 [SC0285]

value: CGCTCAT-AGGCGAA


title: P2I1_plusP_30uM_exp1_rep1 [SC0295]

value: GAGATTC-CAGGACG


title: P2I1_minusP_100uM_exp1_rep2 [SC0284]

value: CGCTCAT-GGCTCTG


title: P2I1_minusP_100uM_exp1_rep1 [SC0283]

value: CGCTCAT-CCTATCC


title: P1P3_minusP_100uM_exp1_rep1 [SC0286]

value: CGCTCAT-TAATCTT


title: P1P3_minusP_100uM_exp1_rep2 [SC0287]

value: CGCTCAT-CAGGACG


title: P1I1_minusP_100uM_exp1_rep2 [SC0281]

value: CGCTCAT-TATAGCC


title: P1I1_minusP_100uM_exp1_rep3 [SC0282]

value: CGCTCAT-ATAGAGG


title: No Bacteria_minusP_100uM_exp1_rep3 [SC0279]

description: No Bacteria_minusP_100uM

source_name: No Bacteria_minusP_100uM_exp1

value: TCCGGAG-CAGGACG


title: P1I1_minusP_100uM_exp1_rep1 [SC0280]

value: whole seedling

value: TCCGGAG-GTACTGA


title: P1P3_minusP_30uM_exp1_rep1 [SC0274]

value: TCCGGAG-ATAGAGG


title: No Bacteria_minusP_100uM_exp1_rep2 [SC0278]

description: No Bacteria_minusP_100uM

value: No Bacteria

value: TCCGGAG-TAATCTT


title: P2P3_minusP_100uM_exp1_rep3 [SC027]

value: GAGATTC-CCTATCC


title: No Bacteria_minusP_100uM_exp1_rep1 [SC0277]

description: No Bacteria_minusP_100uM

value: TCCGGAG-AGGCGAA


title: P1P3_minusP_30uM_exp1_rep2 [SC0275]

value: TCCGGAG-CCTATCC


title: No Bacteria_plusP_30uM_exp1_rep3 [SC0291]

description: No Bacteria_plusP_30uM


title: P2I1_minusP_30uM_exp1_rep3 [SC0273]

value: TCCGGAG-TATAGCC


title: P2I1_minusP_30uM_exp1_rep1 [SC0271]

value: ATTACTC-CAGGACG


title: P1I1_minusP_30uM_exp1_rep2 [SC0269]

value: ATTACTC-AGGCGAA


title: P2P3_minusP_100uM_exp1_rep2 [SC026]

value: GAGATTC-ATAGAGG


title: P1I1_minusP_30uM_exp1_rep1 [SC0268]

value: ATTACTC-GGCTCTG


title: No Bacteria_minusP_30uM_exp1_rep2 [SC0266]

description: No Bacteria_minusP_30uM

source_name: No Bacteria_minusP_30uM_exp1

value: ATTACTC-ATAGAGG


title: P1I1_plusP_30uM_exp1_rep1 [SC0292]

value: GAGATTC-GGCTCTG


title: N3P1_plusP_100uM_exp2_rep3 [SC0264]

value: TAATGCG-GGCTCTG


title: No Bacteria_minusP_30uM_exp1_rep3 [SC0267]

description: No Bacteria_minusP_30uM

value: ATTACTC-CCTATCC


title: N2N3_plusP_100uM_exp2_rep1 [SC0259]

value: whole seedling

value: CTGAAGC-CAGGACG


title: No Bacteria_minusP_30uM_exp1_rep1 [SC0265]

description: No Bacteria_minusP_30uM

value: ATTACTC-TATAGCC


title: P2P3_minusP_100uM_exp1_rep1 [SC025]

value: GAGATTC-TATAGCC


title: N1N2_plusP_100uM_exp2_rep2 [SC0257]

value: CTGAAGC-AGGCGAA


title: N3P1_plusP_100uM_exp2_rep2 [SC0263]

value: TAATGCG-CCTATCC


title: N1N2_plusP_100uM_exp2_rep1 [SC0256]

value: CTGAAGC-GGCTCTG


title: N1N2_plusP_100uM_exp2_rep3 [SC0258]

value: CTGAAGC-TAATCTT


title: N2N3_plusP_100uM_exp2_rep2 [SC0260]

value: CTGAAGC-GTACTGA


title: N2N3_plusP_100uM_exp2_rep3 [SC0261]

value: TAATGCG-TATAGCC


title: I3N1_plusP_100uM_exp2_rep2 [SC0254]

value: CTGAAGC-ATAGAGG


title: P1P3_minusP_30uM_exp1_rep3 [SC0276]

value: TCCGGAG-GGCTCTG


title: P2I1_minusP_30uM_exp1_rep2 [SC0272]

value: ATTACTC-GTACTGA


title: No Bacteria_plusP_100uM_exp2_rep3 [SC0252]

description: No Bacteria_plusP_100uM

value: No Bacteria

value: GAATTCG-GTACTGA


title: No Bacteria_plusP_100uM_exp2_rep2 [SC0251]

description: No Bacteria_plusP_100uM

value: GAATTCG-CAGGACG


title: P1P2_minusP_100uM_exp1_rep3 [SC024]

value: CGCTCAT-GTACTGA


title: N3P1_plusP_30uM_exp2_rep2 [SC0248]

value: whole seedling

value: GAATTCG-GGCTCTG


title: No Bacteria_plusP_100uM_exp2_rep1 [SC0250]

description: No Bacteria_plusP_100uM

value: GAATTCG-TAATCTT


title: P1I1_minusP_30uM_exp1_rep3 [SC0270]

value: ATTACTC-TAATCTT


title: N3P1_plusP_30uM_exp2_rep3 [SC0249]

value: GAATTCG-AGGCGAA


title: N3P1_plusP_100uM_exp2_rep1 [SC0262]

value: TAATGCG-ATAGAGG


title: N2N3_plusP_30uM_exp2_rep1 [SC0244]

value: ATTCAGA-GTACTGA


title: N1N2_plusP_30uM_exp2_rep3 [SC0243]

value: ATTCAGA-CAGGACG


title: N3P1_plusP_30uM_exp2_rep1 [SC0247]

value: GAATTCG-CCTATCC


title: N2N3_plusP_30uM_exp2_rep3 [SC0246]

value: GAATTCG-ATAGAGG


title: N1N2_plusP_30uM_exp2_rep2 [SC0242]

value: ATTCAGA-TAATCTT


title: I3N1_plusP_30uM_exp2_rep3 [SC0240]

value: ATTCAGA-GGCTCTG


title: I3N1_plusP_100uM_exp2_rep3 [SC0255]

value: CTGAAGC-CCTATCC


title: I3N1_plusP_30uM_exp2_rep1 [SC0238]

value: ATTCAGA-ATAGAGG


title: N1N2_plusP_30uM_exp2_rep1 [SC0241]

value: ATTCAGA-AGGCGAA


title: P1P2_minusP_100uM_exp1_rep2 [SC023]

value: CGCTCAT-CAGGACG


title: I3N1_plusP_30uM_exp2_rep2 [SC0239]

value: ATTCAGA-CCTATCC


title: No Bacteria_plusP_30uM_exp2_rep1 [SC0235]

description: No Bacteria_plusP_30uM

value: No Bacteria

value: whole seedling

value: GAGATTC-CAGGACG


title: I3N1_plusP_100uM_exp2_rep1 [SC0253]

value: CTGAAGC-TATAGCC


title: N3P1_minusP_100uM_exp2_rep1 [SC0232]

value: GAGATTC-GGCTCTG


title: N2N3_minusP_100uM_exp2_rep2 [SC0230]

value: GAGATTC-ATAGAGG


title: P1P2_minusP_100uM_exp1_rep1 [SC022]

value: CGCTCAT-TAATCTT


title: N3P1_minusP_100uM_exp2_rep2 [SC0233]

value: GAGATTC-AGGCGAA


title: N3P1_minusP_100uM_exp2_rep3 [SC0234]

value: GAGATTC-TAATCTT


title: No Bacteria_plusP_30uM_exp2_rep2 [SC0236]

description: No Bacteria_plusP_30uM

value: GAGATTC-GTACTGA


title: No Bacteria_plusP_30uM_exp2_rep3 [SC0237]

description: No Bacteria_plusP_30uM

value: ATTCAGA-TATAGCC


title: I3N1_minusP_100uM_exp2_rep2 [SC0224]

value: CGCTCAT-GGCTCTG


title: N2N3_minusP_100uM_exp2_rep1 [SC0229]

value: GAGATTC-TATAGCC


title: I3N1_minusP_100uM_exp2_rep3 [SC0225]

value: CGCTCAT-AGGCGAA


title: N2N3_plusP_30uM_exp2_rep2 [SC0245]

value: GAATTCG-TATAGCC


title: N1N2_minusP_100uM_exp2_rep1 [SC0226]


title: I3N1_minusP_100uM_exp2_rep1 [SC0223]

value: CGCTCAT-CCTATCC


title: N1N2_minusP_100uM_exp2_rep3 [SC0228]

value: CGCTCAT-GTACTGA


title: No Bacteria_minusP_100uM_exp2_rep3 [SC0222]

description: No Bacteria_minusP_100uM

value: No Bacteria

value: CGCTCAT-ATAGAGG


title: No Bacteria_minusP_100uM_exp1_rep3 [SC021]

description: No Bacteria_minusP_100uM

value: whole seedling


title: N3P1_minusP_30uM_exp2_rep2 [SC0218]

value: TCCGGAG-TAATCTT


title: N3P1_minusP_30uM_exp2_rep3 [SC0219]

value: TCCGGAG-CAGGACG


title: N2N3_minusP_30uM_exp2_rep2 [SC0215]

value: TCCGGAG-CCTATCC


title: N2N3_minusP_30uM_exp2_rep3 [SC0216]

value: TCCGGAG-GGCTCTG


title: N1N2_minusP_30uM_exp2_rep3 [SC0213]

value: TCCGGAG-TATAGCC


title: N2N3_minusP_30uM_exp2_rep1 [SC0214]

value: TCCGGAG-ATAGAGG


title: N2N3_minusP_100uM_exp2_rep3 [SC0231]

value: GAGATTC-CCTATCC


title: N1N2_minusP_100uM_exp2_rep2 [SC0227]

value: CGCTCAT-CAGGACG


title: N1N2_minusP_30uM_exp2_rep1 [SC0211]

value: ATTACTC-CAGGACG


title: I3N1_minusP_30uM_exp2_rep2 [SC0209]

value: ATTACTC-AGGCGAA


title: No Bacteria_minusP_100uM_exp1_rep2 [SC020]

description: No Bacteria_minusP_100uM

value: CGCTCAT-GGCTCTG


title: N1N2_minusP_30uM_exp2_rep2 [SC0212]

value: ATTACTC-GTACTGA


title: No Bacteria_minusP_30uM_exp2_rep3 [SC0207]

description: No Bacteria_minusP_30uM

value: ATTACTC-CCTATCC


title: No Bacteria_minusP_30uM_exp2_rep2 [SC0206]

description: No Bacteria_minusP_30uM

value: ATTACTC-ATAGAGG


title: I3N1_minusP_30uM_exp2_rep1 [SC0208]

value: ATTACTC-GGCTCTG


title: No Bacteria_minusP_30uM_exp2_rep1 [SC0205]

description: No Bacteria_minusP_30uM

value: No Bacteria

value: ATTACTC-TATAGCC


title: N3P1_plusP_100uM_exp1_rep3 [SC0204]

value: whole seedling

value: TAATGCG-GGCTCTG


title: No Bacteria_minusP_100uM_exp2_rep1 [SC0220]

description: No Bacteria_minusP_100uM

value: TCCGGAG-GTACTGA


title: N3P1_plusP_100uM_exp1_rep1 [SC0202]

value: TAATGCG-ATAGAGG


title: No Bacteria_minusP_100uM_exp2_rep2 [SC0221]

description: No Bacteria_minusP_100uM

value: CGCTCAT-TATAGCC


title: N3P1_plusP_100uM_exp1_rep2 [SC0203]

value: TAATGCG-CCTATCC


title: N2N3_plusP_100uM_exp1_rep2 [SC0200]

value: CTGAAGC-GTACTGA


title: N2N3_plusP_100uM_exp1_rep1 [SC0199]

value: CTGAAGC-CAGGACG


title: No Bacteria_minusP_100uM_exp1_rep1 [SC019]

description: No Bacteria_minusP_100uM

value: CGCTCAT-CCTATCC


title: I3N1_plusP_100uM_exp1_rep3 [SC0195]

value: CTGAAGC-CCTATCC


title: N1N2_plusP_100uM_exp1_rep3 [SC0198]

value: CTGAAGC-TAATCTT


title: N3P1_minusP_30uM_exp2_rep1 [SC0217]

value: TCCGGAG-AGGCGAA


title: No Bacteria_plusP_100uM_exp1_rep3 [SC0192]

description: No Bacteria_plusP_100uM

value: GAATTCG-GTACTGA


title: N1N2_plusP_100uM_exp1_rep2 [SC0197]

value: CTGAAGC-AGGCGAA


title: I3N1_plusP_100uM_exp1_rep2 [SC0194]

value: CTGAAGC-ATAGAGG


title: I3N1_minusP_30uM_exp2_rep3 [SC0210]

value: ATTACTC-TAATCTT


title: I3N1_plusP_100uM_exp1_rep1 [SC0193]

value: CTGAAGC-TATAGCC


title: N2N3_plusP_100uM_exp1_rep3 [SC0201]

value: whole seedling

value: TAATGCG-TATAGCC


title: N1N2_plusP_100uM_exp1_rep1 [SC0196]

value: CTGAAGC-GGCTCTG


title: I2I3_minusP_30uM_exp1_rep3 [SC018]

value: CGCTCAT-ATAGAGG


title: N3P1_plusP_30uM_exp1_rep2 [SC0188]

value: GAATTCG-GGCTCTG


title: N1N2_plusP_30uM_exp1_rep3 [SC0183]

value: ATTCAGA-CAGGACG


title: N2N3_plusP_30uM_exp1_rep1 [SC0184]

value: ATTCAGA-GTACTGA


title: N2N3_plusP_30uM_exp1_rep3 [SC0186]

value: GAATTCG-ATAGAGG


title: N3P1_plusP_30uM_exp1_rep3 [SC0189]

value: GAATTCG-AGGCGAA


title: N2N3_plusP_30uM_exp1_rep2 [SC0185]

value: GAATTCG-TATAGCC


title: N1N2_plusP_30uM_exp1_rep2 [SC0182]

value: ATTCAGA-TAATCTT


title: N1N2_plusP_30uM_exp1_rep1 [SC0181]

value: ATTCAGA-AGGCGAA


title: I2I3_minusP_30uM_exp1_rep2 [SC017]

value: CGCTCAT-TATAGCC


title: N3P1_plusP_30uM_exp1_rep1 [SC0187]

value: GAATTCG-CCTATCC


title: No Bacteria_plusP_30uM_exp1_rep3 [SC0177]

description: No Bacteria_plusP_30uM

value: No Bacteria

value: ATTCAGA-TATAGCC


title: No Bacteria_plusP_30uM_exp1_rep2 [SC0176]

description: No Bacteria_plusP_30uM

value: GAGATTC-GTACTGA


title: No Bacteria_plusP_100uM_exp1_rep1 [SC0190]

description: No Bacteria_plusP_100uM

value: whole seedling

value: GAATTCG-TAATCTT


title: I3N1_plusP_30uM_exp1_rep2 [SC0179]

value: ATTCAGA-CCTATCC


title: I3N1_plusP_30uM_exp1_rep1 [SC0178]

value: ATTCAGA-ATAGAGG


title: N3P1_minusP_100uM_exp1_rep1 [SC0172]

value: GAGATTC-GGCTCTG


title: No Bacteria_plusP_30uM_exp1_rep1 [SC0175]

description: No Bacteria_plusP_30uM

value: GAGATTC-CAGGACG


title: I3N1_plusP_30uM_exp1_rep3 [SC0180]

value: ATTCAGA-GGCTCTG


title: N3P1_minusP_100uM_exp1_rep2 [SC0173]

value: GAGATTC-AGGCGAA


title: N2N3_minusP_100uM_exp1_rep2 [SC0170]

value: GAGATTC-ATAGAGG


title: N2N3_minusP_100uM_exp1_rep3 [SC0171]

value: GAGATTC-CCTATCC


title: I2I3_minusP_30uM_exp1_rep1 [SC016]

value: TCCGGAG-GTACTGA


title: N1N2_minusP_100uM_exp1_rep1 [SC0166]

value: CGCTCAT-TAATCTT


title: I3N1_minusP_100uM_exp1_rep3 [SC0165]

value: CGCTCAT-AGGCGAA


title: N2N3_minusP_100uM_exp1_rep1 [SC0169]

value: GAGATTC-TATAGCC


title: N1N2_minusP_100uM_exp1_rep2 [SC0167]

value: CGCTCAT-CAGGACG


title: N1N2_minusP_100uM_exp1_rep3 [SC0168]

value: CGCTCAT-GTACTGA


title: N3P1_minusP_100uM_exp1_rep3 [SC0174]

value: GAGATTC-TAATCTT


title: No Bacteria_minusP_100uM_exp1_rep2 [SC0161]

description: No Bacteria_minusP_100uM

value: No Bacteria

value: whole seedling

value: CGCTCAT-TATAGCC


title: No Bacteria_minusP_100uM_exp1_rep3 [SC0162]

description: No Bacteria_minusP_100uM

value: CGCTCAT-ATAGAGG


title: I3N1_minusP_100uM_exp1_rep2 [SC0164]

value: CGCTCAT-GGCTCTG


title: N3P1_minusP_30uM_exp1_rep2 [SC0158]

value: TCCGGAG-TAATCTT


title: I1I2_minusP_30uM_exp1_rep3 [SC015]

value: TCCGGAG-CAGGACG


title: N3P1_minusP_30uM_exp1_rep3 [SC0159]


title: No Bacteria_minusP_100uM_exp1_rep1 [SC0160]

description: No Bacteria_minusP_100uM


title: N2N3_minusP_30uM_exp1_rep2 [SC0155]

value: TCCGGAG-CCTATCC


title: N2N3_minusP_30uM_exp1_rep3 [SC0156]

value: TCCGGAG-GGCTCTG


title: N2N3_minusP_30uM_exp1_rep1 [SC0154]

value: TCCGGAG-ATAGAGG


title: N1N2_minusP_30uM_exp1_rep3 [SC0153]

value: TCCGGAG-TATAGCC


title: No Bacteria_minusP_30uM_exp1_rep2 [SC0146]

description: No Bacteria_minusP_30uM

value: ATTACTC-ATAGAGG


title: I1I2_minusP_30uM_exp1_rep2 [SC014]


title: I3N1_minusP_30uM_exp1_rep1 [SC0148]

value: ATTACTC-GGCTCTG


title: No Bacteria_minusP_30uM_exp1_rep3 [SC0147]

description: No Bacteria_minusP_30uM

value: ATTACTC-CCTATCC


title: N1N2_minusP_30uM_exp1_rep2 [SC0152]

value: ATTACTC-GTACTGA


title: I3N1_minusP_30uM_exp1_rep2 [SC0149]

value: ATTACTC-AGGCGAA


title: I1I2_minusP_30uM_exp1_rep1 [SC013]

value: TCCGGAG-AGGCGAA


title: I3N1_minusP_30uM_exp1_rep3 [SC0150]

value: ATTACTC-TAATCTT


title: I3N1_minusP_100uM_exp1_rep1 [SC0163]

value: whole seedling

value: CGCTCAT-CCTATCC


title: P3I1_minusP_30uM_exp1_rep3 [SC012]


title: N1N2_minusP_30uM_exp1_rep1 [SC0151]

value: ATTACTC-CAGGACG


title: No Bacteria_minusP_30uM_exp1_rep1 [SC0145]

description: No Bacteria_minusP_30uM

value: No Bacteria

value: ATTACTC-TATAGCC


title: P2P3_minusP_30uM_exp1_rep2 [SC008]


title: P2P3_minusP_30uM_exp1_rep3 [SC009]


title: P2P3_minusP_30uM_exp1_rep1 [SC007]


title: P1P2_minusP_30uM_exp1_rep1 [SC004]


title: N3P1_minusP_30uM_exp1_rep1 [SC0157]


title: P3I1_minusP_30uM_exp1_rep1 [SC010]


title: No Bacteria_minusP_30uM_exp1_rep3 [SC003]

description: No Bacteria_minusP_30uM


title: P1P2_minusP_30uM_exp1_rep2 [SC005]


title: edm2-4_mRNA-Seq_rep2

city: West Lafayette

first: Jian-Kang

last: Zhu

state: IN

address: 625 Agriculture Mall Dr.

department: Department of Horticulture and Landscape Architecture

institute: Purdue University

data_processing:
MethylC-Seq:Reads were mapped to TAIR10 using BRAT-BW v2.0.1 with parameters -m 2 -i 0 -a 1000. Copy-duplicates were remove using remove-dupl command of BRAT-BW. acgt-count command of BRAT-BW was used to count mapped Cs and Ts at each cytosine of forward and reverse strands of the reference
mRNA-Seq: Clean reads were mapped to TAIR10 by TopHat2.
Genome_build: TAIR10
Supplementary_files_format_and_content: For MethylC-Seq: The wig files give methylation level for all Cs(three contexts) with depth >= 4. Methylation level of each cytosine base was calculated according to number of converted and unconverted C mapped at that position (unconverted_C / ( converted_C+ unconverted_C ) ).
Supplementary_files_format_and_content: For sRNA-Seq and mRNA-Seq: The bedGraph files were generated using genomeCoverageBed from bedtools with parameter  -bga -split

source_name: seedlings

extract_protocol: Total RNA were extracted from 2-week-old seedlings using Trizol reagent (Ambion, 15596) and treated with DNase (Turbo) to remove any genomic DNA contaminants.  Library construction and deep sequencing were performed by the Core Facility of Genomics in Shanghai Plant Stress Biology Center, China.

tag: genotype/variation

tag: ecotype


title: No Bacteria_minusP_30uM_exp1_rep1 [SC001]

description: No Bacteria_minusP_30uM


title: No Bacteria_minusP_30uM_exp1_rep2 [SC002]

description: No Bacteria_minusP_30uM


title: Col-0_mRNA-Seq_rep2

value: wt


title: edm2-4_mRNA-Seq_rep1


title: Col-0_mRNA-Seq_rep1


title: asi1-2_mRNA-Seq_rep2


title: P3I1_minusP_30uM_exp1_rep2 [SC011]

value: TCCGGAG-CCTATCC


title: Col-0_MethylC-Seq_rep2

description: treated with bisulfite

library_source: genomic

molecule: genomic DNA

extract_protocol: Genomic DNA was extracted from 2-week-old seedlings and sent to Genomics Core Facility of Shanghai Center for Plant Stress Biology for bisulfite treatment, library preparation, and sequencing. Bisulfite sequencing libraries were prepared using standard Illumina protocols.


title: aipp1-2_MethylC-Seq_rep2

description: treated with bisulfite


title: P1P2_minusP_30uM_exp1_rep3 [SC006]


title: aipp1-1_MethylC-Seq_rep1

description: treated with bisulfite


title: Col-0_MethylC-Seq_rep1

description: treated with bisulfite


title: aipp1-2_MethylC-Seq_rep1

description: treated with bisulfite


title: asi1-2_mRNA-Seq_rep1


title: aipp1-1_mRNA-Seq_rep2


title: aipp1-1_mRNA-Seq_rep1


title: aipp1-1_MethylC-Seq_rep2

description: treated with bisulfite


title: RNA NMP Single Cell CS95

city: Berlin

first: Frederic

last: Koch

address: Ihnestraße 63-73

department: Developmental Genetics

country: Germany

institute: Max Planck Institute for Molecular Genetics

description:
embryos obtained from tatraploid complementation of embryonic stem cells
FPKM_SingleCell.txt
Fluidigm Single-Cell mRNA-Seq

data_processing:
For single-cell RNA-Seq data, those libraries with very few reads (2 samples with less than 1x10^6 reads) were excluded from further processing. Reads were mapped to the mm10 genome as previously described for the bulk RNA-Seq analysis with TopHat2 (version 2.1.0) (Kim et al., 2013) using bowtie (Langmead et al., 2009), providing refSeq annotations in gtf format (UCSC) and the options  --no-mixed --no-discordant -g1 --mate-inner-dist 200 --mate-std-dev 100, without initial trimming of reads. We then calculated FPKM values using Cuffquant and Cuffnorm from the Cufflinks (version 2.2.1) (Trapnell et al., 2012) package. Downstream analysis was carried out using the R Monocle package (version 2.4.0) (Qiu, et al., 2017; Trapnell, et al., 2014), by reading in the FPKM, sample and gene attributes tables from the Cufflinks output.
Genome_build: mm10
Supplementary_files_format_and_content: tab-delimited text file includes FPKM values for every gene across all single-cell RNA-Seq samples

molecule: polyA RNA

organism: Mus musculus

source_name: T+ / Sox2+ FACS sorted cells from caudal ends of WT embryos

growth_protocol: ES cell lines were cultured according to standard conditions. Cells were maintained on gelatinized plates and mitotically inactive primary mouse embryo fibroblasts with standard ESC medium, refreshed daily, containing 15% FCS and 1000 U/ml leukemia inhibitory factor (LIF, Chemicon ESG1107). ES cells were used to generate embryos via the tetraploid complementation assay.

extract_protocol:
The IFC was processed according to the Fluidigm mRNA-Seq protocol. A 1:4000 dilution of ERCC spike-ins (Ambion) were added to the lysis buffer. Upon visual inspection, 55 capture sites were called to contain cells. The amplified cDNA was measured using the Qubit DNA HS assay and a selection of samples was verified on a Bioanalyzer DNA HS chip (Agilent).
The final libraries for those 55 capture sites were prepared with the Nextera XT (Illumina) kit according to the protocol, pooled and sequenced on a NextSeq (Illumina) sequencer using 2x75bp read lengths.

treatment_protocol: In order to obtain single cell suspensions from embryo tissue for FACS sorting, 10ul of 5% BSA in PBS was added to the pooled samples and 100ul of ice-cold 0.05% trypsin was added. A homogenous suspension was achieved by slow pipetting and the reaction was stopped with the addition of 200ul of 5% BSA in PBS. Before cell sorting on a FACS Aria II (Becton Dickinson), the cell suspension was filtered through a 35µm mesh. the sorted cells were collected in Eppendorf tubes containing filtered 5% BSA in PBS at 4C. Cells were pelleted by centrifugation and subsequently resuspended in 10ul PBS. The density and viability (>90%) were checked on a Luna cell counter (Logos Biosystems). The suspension was adjusted to a cell density of 250 cells/µl, resuspension buffer added (4.5:1 ratio of cells:buffer) and approximately 5µl were loaded onto a 5-10µm C1 IFC (Fluidigm).

tag: strain

tag: developmental stage


title: RNA NMP Single Cell CS94


title: RNA NMP Single Cell CS47


title: RNA NMP Single Cell CS90


title: RNA NMP Single Cell CS45


title: RNA NMP Single Cell CS46


title: RNA NMP Single Cell CS91


title: RNA NMP Single Cell CS93


title: RNA NMP Single Cell CS44


title: RNA NMP Single Cell CS92


title: RNA NMP Single Cell CS39


title: RNA NMP Single Cell CS41


title: RNA NMP Single Cell CS40


title: RNA NMP Single Cell CS38


title: No Bacteria_plusP_100uM_exp1_rep2 [SC0191]

description: No Bacteria_plusP_100uM

value: GAATTCG-CAGGACG


title: RNA NMP Single Cell CS37


title: RNA NMP Single Cell CS36


title: RNA NMP Single Cell CS35


title: RNA NMP Single Cell CS42


title: RNA NMP Single Cell CS43


title: RNA NMP Single Cell CS33


title: RNA NMP Single Cell CS31


title: RNA NMP Single Cell CS29


title: RNA NMP Single Cell CS28


title: RNA NMP Single Cell CS32


title: RNA NMP Single Cell CS27


title: RNA NMP Single Cell CS23


title: RNA NMP Single Cell CS30


title: RNA NMP Single Cell CS25


title: RNA NMP Single Cell CS20


title: RNA NMP Single Cell CS26


title: RNA NMP Single Cell CS14


title: RNA NMP Single Cell CS24


title: RNA NMP Single Cell CS22


title: RNA NMP Single Cell CS12


title: RNA NMP Single Cell CS11


title: RNA NMP Single Cell CS10


title: RNA NMP Single Cell CS17


title: RNA NMP Single Cell CS19


title: RNA NMP Single Cell CS15


title: RNA NMP Single Cell CS16


title: RNA NMP Single Cell CS13


title: RNA NMP Single Cell CS34


title: RNA NMP Single Cell CS8


title: RNA NMP Single Cell CS7


title: RNA NMP Single Cell CS9


title: RNA NMP Single Cell CS6


title: RNA NMP Single Cell CS5


title: RNA NMP Single Cell CS4


title: RNA NMP Single Cell CS18


title: RNA NMP Single Cell CS21


title: RNA NMP Single Cell CS3


title: RNA NMP Single Cell CS2


title: ChIP Input

description: in vitro differentiated neuromesodermal progenitors

data_processing:
ChIP-Seq data was using bowtie. For Tbx6, we estimated the fragment size distribution of the library and determined the maximum length cutoff by performing an initial mapping of a limited number of reads using the options -y -m 1 -S -I 100 -X 500. The final mapping was performed using the same options and the determined maximum length parameter (-X). For the NMP ChIP’s and input, we used -X 1000 for the maximum length parameter.  Paired-end reads from the .sam file were combined in a bed file by generating the original fragment using a custom perl script, duplicates were removed and .wig files were generated using BEDtools. For NMP’s, we made use of the input file and removed all reads overlapping with regions displaying enrichment in the input from downstream analysis.
Genome_build: mm10
Supplementary_files_format_and_content: bigWig files representing read density for ChIP-Seq

source_name: in vitro differentiated ES cells

growth_protocol: For NMP differentiation, we used a previously described protocol (Gouti, et al., 2014) with slight modifications. ESCs were removed from feeders by dissociation using 0.05% trypsin and then plated onto tissue culture plates for four short successive periods (25’, 20’, 15’, 10’) to remove feeders. We then cultivated the cells for at least two passages in 2i + Lif medium (1:1 DMEM/F12:Neurobasal medium, 1x Pen/Strep, 1x Glutamax, 1x non-essential amino-acids, 1x Na-Pyruvate, 0,11mM β-mercaptoethanol, 1x N2, 1x B-27 without vitamin A, 1µM PD0325901 (Stemgent), 3µM CHIR99021 (Stemgent), 1x Lif). Cells were then counted and plated on CellBINDSurface dishes (Corning) precoated with Synthemax II-SC (Corning) at a density of 1x104 cells per cm2 in a medium consisting of ‘N2B27’ medium  (Gouti et al., Plos Biol 2014), supplemented with 10ng/ml bFGF (Peprotech) for 3 days (d1-d3). From d2-d3 5µM CHIR99021 (CHI, Calbiochem) was added to the medium.

extract_protocol:
For sonication, complete protease inhibitors without EDTA (Roche) at 1x final concentration was added to all lysis buffers (LB) prior to use. Each pellet was resuspended in LB1 (50mM Hepes pH7.5, 140mM NaCl, 1mM EDTA pH8, 10% glycerol, 0.75% NP-40, 0.25% triton X-100) and rotated at 4°C for 20 minutes. The cell suspension was homogenized using a douncer with a tight pestle with 3x 15 strokes, with short resting times on ice in between. The chromatin was pelleted by centrifugation at 1400g and 4°C for 5 minutes and resuspended in 2.5ml LB2 (200mM NaCl, 1mM EDTA pH8, 0.5mM EGTA pH8, 10mM Tris pH 8) per cell pellet. After 10 minutes rotation at 4°C, the centrifugation step was repeated and each pellet was resuspended in 1.5ml LB3 (1mM EDTA pH8, 0.5mM EGTA, 10mM Tris pH8, 100mM NaCl, 0.1% Na-deoxycholate, 0.5% N-lauroylsarcosine), transferred to 15ml Falcon tubes and sonicated using a W-450D Digital Sonifier (Branson) sonicator for 14 cycles of 10s on/50s off in a 4°C coldroom with tubes chilled in ice water. After sonication, 150ul of triton X-100 was added per tube, transferred to two 1.5ml Eppendorf tubes and debris was pelleted by centrifugation at 20,000g and 4°C for 10 minutes. The solubilized chromatin was then pooled and mixed thoroughly. After taking 50ul as an input control, the chromatin was distributed into 1.5ml aliquots, snap frozen and stored at -80C until use.  The input was reverse-crosslinked with the addition of 50ul 2x elution buffer (100mM Tris pH8, 20mM EDTA pH8, 2% SDS) and overnight (13h-15h) incubation at 65°C. The next day, 100ul of TE buffer were added to dilute the SDS of the elution buffer and RNA was digested using RNase A at a 0.2ug/ml final concentration at 37°C for 2 hours. Proteins were digested with the addition of 0.2ug/ml proteinase K and incubation at 55°C for 2 hours. The DNA was purified with two subsequent phenol:chloroform:isoamylalcohol (25:24:1, pH8) isolations and subsequent MinElute (Qiagen) purification. Purification was performed according to manufacturer’s recommendations, except that the columns were washed 2x with PE and air dried for at least 10 minutes before elution with EB. The input DNA was quantified using a NanoPhotometer (Implen) and the shearing was verified by running 600ng of DNA on a 1.5% agarose gel. For sonication to be considered successful and sonicated chromatin to be usable for ChIP, the bulk DNA of the input had to be located between 100bp-500bp.  For ChIP, 50µl or 100ul (for β-catenin) of protein-coated Dynal beads (Life Technologies) were washed 3x using 1ml of blocking solution (PBS containing 0.5% BSA), resuspended in 500µl of blocking solution together with approximately 5µg or 10 µg (for β-catenin) antibody and rotated overnight at 4°C. The antibodies used were a goat polyclonal anti-T (Santa Cruz, sc-17743), a goat polyclonal anti-Sox2 (R&D Systems, AF2018), a rabbit polyclonal anti-β-catenin (Invitrogen, 71-2700) and a rabbit polyclonal anti-Tbx6 (this paper). The next day, three further washes using 1ml blocking buffer were performed and the beads were resuspended in 100µl blocking buffer. Chromatin equivalent to approximately 25x106 or 50x106 (for β-catenin) cells was added and rotated overnight at 4°C. The following day, washing steps (6, 8 and 9 washes for Tbx6, Sox2 / β-catenin and T respectively) were performed using 1ml RIPA buffer (50mM Hepes pH7.6, 500mM LiCl, 1mM EDTA pH8, 1% NP-40, 0.7% Na-Deoxycholate) containing 1x complete protease inhibitors without EDTA and finally with 1ml of TEN (10mM Tris pH8, 1mM EDTA pH8, 50mM NaCl). The immunoprecipates were eluted in two subsequent steps using 100µl of 1x elution buffer (50mM Tris pH8, 10mM EDTA pH8, 1% SDS) and incubation at 65°C while shaking for 10 minutes each. The eluates were combined and incubated overnight (13h-15h) at 65°C under agitation. The next day, 200µl of TE were added to dilute the SDS of the elution buffer and the ChIPs were purified as described for the input above. ChIP DNA was quantified using the Qubit (Life Technologies) DNA HS assay.
The ChIP-Seq libraries were generated using the TruSeq ChIP library kit (Illumina) with the following modifications. After adapter ligation, 0.95x of AMPure XP beads (Beckman Coulter) were used for a single purification and the DNA was eluted using 15µl of resuspension buffer (RSB, Illumina). In the case of Tbx6, 14µl of the eluate were used for PCR pre-amplification in order to convert adapters into double-stranded DNA. After the addition of 1µl primer mix (25mM each, Primer 1: 5’- AATGATACGGCGACCACCGA*G-3’; Primer2: 5’- CAAGCAGAAGACGGCATACGA*G-3’) and 15µl 2x Kapa HiFi HotStart Ready Mix (Kapa Biosystems), amplification was performed for 45 seconds at 98°C, 5 cycles of [15 seconds at 98°C, 30 seconds at 63°C and 30 seconds at 72°C] and a final 1 minute incubation at 72°C. The PCR products were purified using 0.95x of beads and eluted using 21µl of RSB. Libraries were directly amplified for 18 cycles. The libraries were quantified using the Qubit DNA HS assay and the library size was validated using DNA HS bioanalyzer chips.

treatment_protocol: Crosslinking was performed directly on differentiating cells in growth medium with the addition of 1/10th volume of crosslinking solution (11% formaldehyde, 100mM NaCl, 1mM EDTA pH8, 0.5mM EGTA pH8, 50mM Hepes pH7.8) for 10 minutes at room temperature, with the dishes placed on a shaker. The crosslinking reaction was quenched with the addition of 1/10th volume of 2.5M glycine and 5 minutes incubation. Dishes were placed on ice or in a 4°C cold room and washed twice with cold PBS. Cells were scraped and washed off using cold PBS, containing 0.05% Triton X-100 to break the surface tension. Cells were pooled in aliquots of ~30-50x106 in Eppendorf tubes, snap frozen and stored at -80°C until sonication.


title: RNA NMP Single Cell CS1


title: Hypothalamus_WT3 RNA-Seq

city: Copenhagen

first: Aditya

last: Sankar

email: cnj376@ku.dk, aditya.sankar@embl.de

address: Ole Maaloes Vej 5

department: Biotech Research and Innovation Centre

country: Denmark

institute: University of Copenhagen

description:
2.5dpc (Timed mating)
DeSeq2-Hypothalamus_WT3

data_processing:
Galaxy: Fastq sequencing files were concatenated,and quality checked using FastQC.
Galaxy: Adaptor reads were trimmed using Trimmomatic prior to mapping
Galaxy: Files were aligned to mouse mm10 reference genome using RNA-STAR tool for Illumina single end sequencing. The individual BAM files were counted using the htseq-count tool using Union mode and unstranded to generate individual count tables.
Galaxy: Counting was followed by computing differential gene expression using DESeq2 for count tables comparing three knockout samples versus three wildtype samples in a batch mode.
Output file were converted into text format and then opened in excel and sorted in decending order of most downregulated gene to most upregulated with log2 fold change of 0.75 and corrected p-value less than 0.1 .
Genome_build: mm10
Supplementary_files_format_and_content: DESeq2 generates a counts file.
Supplementary_files_format_and_content: DESeq2 generates a tabular KO_vs_WT file containing the following columns:
Column Descriptions:
1 Gene Identifiers
2 mean normalised counts, averaged over all samples from both conditions
3 the logarithm (to basis 2) of the fold change (See the note in inputs section)
4 standard error estimate for the log2 fold change estimate
5 Wald statistic
6 p value for the statistical significance of this change
7 p value adjusted for multiple testing with the Benjamini-Hochberg procedure which controls false discovery rate (FDR)

source_name: Whole Hypothalamus at 2.5dpc, WT

growth_protocol: Virgin mature female mice between 8-10 weeks of age of appropriate genotype were housed together.

extract_protocol:
At 2.5 dpc (days post coitus), females were sacrificed and checked for presence of preimplantation embryos by flushing the oviduct. Whole uterus, hypothalamus, pituitary and ovaries were dissected and immediately cleaned of fat and flash frozen in liquid nitrogen. RNA was extracted using Qiagen RNAeasyPlus mini kit following homogenization in RLTplus buffer., tissue RNA quality was measured using an Agilent 2100 Bioanalyzer system (Agilent, #G2940CA) and RNA 6000 Nano reagents (Agilent, #5067-1511). RIN scores were at least 8 for samples used in the RNA Seq with three biological replicates for each tissue
RNASeq libraries were generated using the TruSeq RNA Sample Prep Kit v2 (Illumina, # RS-122-2001) using 12 indexes each from  Set A and B. 0.5 μg of total RNA was enriched for the Poly-A mRNA and reverse transcribed to double-stranded cDNA. The cDNA was end repaired, adenylated, and ligated with Illumina indexes prior to 13 cycles of PCR and quantified using Qubit (Thermo Scientific, #Q32851) and the Agilent 2100 Bioanalyzer system (Agilent, #G2940CA).

treatment_protocol: Females were exposed to dirty bedding of stud male briefly and then set up for timed mating with a stud male (2 females per male)

value: Hypothalamus

tag: age

value: 8-10 week old

tag: genotype

value: Wildtype

tag: Stage


title: Hypothalamus_KO2 RNA-Seq

description:
2.5dpc (Timed mating)
DeSeq2-Hypothalamus_KO2

source_name: Whole Hypothalamus at 2.5dpc, KO

value: Knockout


title: Pituitary_KO2 RNA-Seq

description:
2.5dpc (Timed mating)
DeSeq2-Pituitary_KO2

source_name: Whole Pituitary at 2.5dpc, KO

value: Pituitary


title: Pituitary_WT2 RNA-Seq

description:
2.5dpc (Timed mating)
DeSeq2-Pituitary_WT2

source_name: Whole Pituitary at 2.5dpc, WT


title: Hypothalamus_WT1 RNA-Seq

description:
2.5dpc (Timed mating)
DeSeq2-Hypothalamus_WT1


title: Pituitary_KO1 RNA-Seq

description:
2.5dpc (Timed mating)
DeSeq2-Pituitary_KO1


title: Hypothalamus_KO1 RNA-Seq

description:
2.5dpc (Timed mating)
DeSeq2-Hypothalamus_KO1


title: Uterus_KO2 RNA-Seq

description:
2.5dpc (Timed mating)
DeSeq2-Uterus_KO2

source_name: Whole Uterus at 2.5dpc, KO

value: Uterus


title: Uterus_KO1 RNA-Seq

description:
2.5dpc (Timed mating)
DeSeq2-Uterus_KO1


title: Pituitary_WT1 RNA-Seq

description:
2.5dpc (Timed mating)
DeSeq2-Pituitary_WT1


title: Uterus_KO3 RNA-Seq

description:
2.5dpc (Timed mating)
DeSeq2-Uterus_KO3


title: Uterus_WT2 RNA-Seq

description:
2.5dpc (Timed mating)
DeSeq2-Uterus_WT2

source_name: Whole Uterus at 2.5dpc, WT


title: Ovary_KO1 RNA-Seq

description:
2.5dpc (Timed mating)
DeSeq2-Ovary_KO1

source_name: Whole Ovary at 2.5dpc, KO

value: Ovary


title: Pituitary_WT3 RNA-Seq

description:
2.5dpc (Timed mating)
DeSeq2-Pituitary_WT3


title: Ovary_KO3 RNA-Seq

description:
2.5dpc (Timed mating)
DeSeq2-Ovary_KO3


title: Ovary_WT3 RNA-Seq

description:
2.5dpc (Timed mating)
DeSeq2-Ovary_WT3

source_name: Whole Ovary at 2.5dpc, WT


title: Uterus_WT1 RNA-Seq

description:
2.5dpc (Timed mating)
DeSeq2-Uterus_WT1


title: Ovary_WT2 RNA-Seq

description:
2.5dpc (Timed mating)
DeSeq2-Ovary_WT2


title: Ovary_WT1 RNA-Seq

description:
2.5dpc (Timed mating)
DeSeq2-Ovary_WT1


title: Input WT

description: Histone input_WT

data_processing:
Galaxy: Fastq sequencing files were concatenated, subjected to FASTQ grooming
Galaxy: Cutadapt tool was used to remove adaptor sequences
Galaxy: Files were aligned to mouse mm9 reference genome using Bowtie for Illumina single end sequencing, the BAM files were then converted to BED files, and peaks were called using MACS peak calling tool including an option to generate "wig" files (attached processed data). For peak calling, Kdm4a KO was used as negative control for Kdm4a Chip, and input chromatin of appropriate genotype was used as negative control for histone ChiP
EaSeq: Galaxy generated BED files were used as input into EaSeq software. Unique reads from duplicates were pooled and shrunk randomly equally for wildtype and knockout samples to allow for direct comparison.
EaSeq: Entire data processing to generate heatmaps, TSS regionsets and filled tracks were followed as instructed in this tutorial. (https://www.youtube.com/watch?v=o3Waz8xwZXU)
Genome_build: mm9
Supplementary_files_format_and_content: wig

source_name: Uterus

extract_protocol:
At 2.5 dpc (days post coitus), females were sacrificed and checked for presence of preimplantation embryos by flushing the oviduct. Whole uterus were immediately cleaned of fat and cut into halves and pieces from each female were cross linked with 1% paraformaldehyde in PBS at room temperature for 10 minutes. Then, they were placed in 0.125 M glycine for 5 minutes to stop cross-linking. Subsequently, samples were washed in cold PBS twice and then homogenized to single cells using mechanical shearing of tissue in lysis buffer without detergent (SDS). Cells were pellleted at 1200 rpm for 5 minutes and then supernatant aspirated. Pellet was resuspended in lysis buffer with detergent (SDS) to release chromatin for 10 minutes at room temperature. Loose chromatin was pelleted at 1500 rpm for 5 minutes and supernatant discarded. Pellet was resuspended in ice-cold sonication buffer and subjected to sonication using Diagenode Picoruptor. Sonication efficiency is checked on 1% agarose gel upon decrosslinking and sheared chromatin is obtained by spinning at 20000G/4C/15 mins to pellet down non-chromatin fragments. Pure chromatin is measured for protein concentration and stored at -80 C till immunoprecipitation.
Protocol followed manufacturers instructions for ChiPSeq libraries using the NEBNext Ultra II DNA library prep kit (Illumina, #E7645)

value: WT

value: 2.5 dpc

tag: antibody

tag: antibody amount

tag: input amount

value: Chromatin ( 1 ug = 1% input))


title: IgG WT ChIP-Seq

value: Anti-Rabbit IgG (Sigma Aldrich; Catalog nr #I5006)

tag: tissue

value: Chromatin (100 ug)


title: Pituitary_KO3 RNA-Seq

description:
2.5dpc (Timed mating)
DeSeq2-Pituitary_KO3


title: Hypothalamus_WT2 RNA-Seq

description:
2.5dpc (Timed mating)
DeSeq2-Hypothalamus_WT2


title: ChIP Beta-Catenin


title: Input KO

description: Histone input_KO

value: KO


title: Uterus_WT3 RNA-Seq

description:
2.5dpc (Timed mating)
DeSeq2-Uterus_WT3


title: Hypothalamus_KO3 RNA-Seq

description:
2.5dpc (Timed mating)
DeSeq2-Hypothalamus_KO3


title: H3K9me3_KO2 ChIP-Seq

value: Anti-H3K9me3 (Abcam; Catalog Nr ab8898)


title: H3K9me3_WT2 ChIP-Seq


title: H3K9me3_KO1 ChIP-Seq


title: H3K9me3_WT1 ChIP-Seq


title: Ovary_KO2 RNA-Seq

description:
2.5dpc (Timed mating)
DeSeq2-Ovary_KO2


title: H3K4me3_WT2 ChIP-Seq

value: Anti-H3K4me3 (Cell Signaling; Catalog Nr #9751)


title: H3K4me3_KO1 ChIP-Seq


title: H3K4me3_KO2 ChIP-Seq


title: H3K4me3_WT1 ChIP-Seq


title: Kdm4a_KO ChIP-Seq

value: Anti-Jmjd2a (Cell Signaling; Catalog Nr #5328)

value: Chromatin (1mg)


title: Kdm4a_WT2 ChIP-Seq


title: Kdm4a_WT1 ChIP-Seq


title: ATAC T+ WT TS13

description:
ATAC
embryos obtained from tatraploid complementation of embryonic stem cells

data_processing:
For RNA-Seq, reads were mapped with TopHat2 (version 2.1.0) (Kim et al., 2013) using bowtie (version 1.1.2) (Langmead et al., 2009), providing refSeq annotations in gtf format (UCSC) and the options --no-coverage-search --no-mixed --no-discordant -g1 --library-type (“fr-firststrand” or “fr-secondstrand” for libraries generated using the TotalScript and Scriptseq kits respectively). Reads that were called to be spliced and containing ambiguous strand information (e.g. containing both XS:A:+ and XS:A:- tags) were removed together with their respective mates. For visualization, wiggle tracks were generated with BEDTools (version 2.23.0) (Quinlan and Hall, 2010), normalized towards the sample with the lowest read count, converted into bigwig format and loaded into the Integrated Genome Browser (Freese et al., 2016). For calculation of FPKM’s, we used Cuffdiff, part of Cufflinks (version 2.2.1) (Trapnell et al., 2012), with the options -u --no-effective-length-correction -b.
ChIP-Seq data was using bowtie version 1.1.2. To estimate the fragment size distribution of the libraries and determine the maximum length cutoff, we performed an initial mapping of a limited number of reads using the options -y -m 1 -S -I 100 -X 500. The final mapping was performed using the same options and the determined maximum length parameter (-X). Paired-end reads were combined in a bed file by generating the original fragment using a custom perl script, duplicates were removed and .wig files were generated using BEDtools.
For ATAC-Seq, reads were trimmed to 50bp length and adapters were detected and removed using fastq-mcf of ea-utils  (https://expressionanalysis.github.io/ea-utils, version 1.04.738). Those reads in which the adapter sequences were not found, were further trimmed by 5bp (to account for potentially undetected but present short adapter sequences) and paired-end mapped using bowtie ((Langmead, et al., 2009) version 1.1.2), allowing a maximum fragment size of 2000bp. Possible PCR duplicates were removed using Picard (https://broadinstitute.github.io/picard, version 1.103). Reads for which the adapter sequences were found and removed were independently mapped from their mates to the mm10 genome (containing chromosomes 1-19, X, Y and M) using bowtie and supplying the options -y -m 1 –S.. After merging the resulting output files, the samtools ((Li, et al., 2009), version 1.2) fixmate function was used to combine corresponding reads into mates. Only those properly mapped mates with a maximum fragment length of 2000bp were filtered and potential PCR duplicates were removed using Picard. The resulting pairs were combined with the previously mapped reads, resulting in the final alignment file. Inspection of the data revealed 35 regions in the genome, mostly repetitive and on average 18kb in size, displaying mapping artefacts (Buenrostro, et al., 2015). We used samtools to remove reads, together with their corresponding mates, falling into these regions. Due to the repetitive nature of the Y chromosome and non-informative mitochondrial genome, reads mapped to either of them were filtered out. Paired-end reads were combined in a bed file by generating the original fragment using a custom perl script, duplicates were removed and .wig files were generated using BEDtools.
Genome_build: mm10
Supplementary_files_format_and_content: bigWig files representing read density for ATAC-Seq and ChIP-Seq
Supplementary_files_format_and_content: tab-delimited text files include FPKM values for each RNA-Seq sample for refseq genes

source_name: T+ FACS sorted cells from caudal ends of WT embryos

extract_protocol:
In total, 1,000-5,000 cells were used per ATAC-Seq experiment. After spinning down the FACS sorted cells, the pellets were resuspended in 50ul of lysis buffer (10mM Tris pH7.4, 10mM NaCl, 3mM MgCl2, 0.1% NP-40) and immediately spun down at 500g and 4°C for 10 min. The supernatant was discarded and each pellet was resuspended in the transposition reaction mix (25ul 2x TD buffer, 2.5ul Tn5 transposase, 22ul H2O) and incubated at 37°C for 30 minutes. After incubation, the reaction was stopped with the addition of PB buffer (Qiagen) and the tagmented DNA was purified using the MinElute kit (Qiagen).
The DNA was combined with the Nextera PCR primers and 2x Kapa HiFi Hotstart Readymix and pre-amplified (98°C 30 seconds, 8x [98°C 10 seconds, 63°C 30 seconds, 72°C 1 minute] in a 50ul volume. To determine the remaining cycles to avoid potential overamplification, 5ul of the preamplification mix was combined with the primers, 1x Evagreen Sybr green (Jena Biosciences) and 2x Kapa HiFi Hotstart Readymix in a 15ul total volume and run for 30 cycles on a StepOne Plus. The remaining 45ul pre-amplified samples were amplified for a further 9 cycles, resulting in a total of 17 cycles. The libraries were purified using MiElute column (Qiagen) and the concentration was measured using the DNA HS Qubit assay (life). Approximately 4ng of each library were run on a DNA HS Bioanalyzer ChIP to verify library size and calculate molarities.

treatment_protocol: In order to obtain single cell suspensions from embryo tissue for FACS sorting, 10ul of 5% BSA in PBS was added to the pooled samples and 100ul of ice-cold 0.05% trypsin was added. A homogenous suspension was achieved by slow pipetting and the reaction was stopped with the addition of 200ul of 5% BSA in PBS. Before cell sorting on a FACS Aria II (Becton Dickinson), the cell suspension was filtered through a 35µm mesh. For downstream RNA-Seq experiments cells were sorted directly into Eppendorf tubes containing 350µl RLT buffer (Qiagen) and 3.5ul b-mercaptoethanol. For ATAC-Seq, cells were sorted into FACS-tubes containing 5% BSA in PBS, which had previously been rotating for several hours to pre-coat the tube walls with BSA. Sorted cells were centrifuged at 1000rpm and 4°C for 10 minutes, transferred to Eppendorf tubes containing PBS with 0.05% Triton X-100 and centrifuged at 1000rpm and 4°C for 10 minutes. The supernatant was removed and cells were directly processed for ATAC-Seq.


title: ChIP Sox2

description:
ChIP-Seq
in vitro differentiated neuromesodermal progenitors

extract_protocol:
For sonication, complete protease inhibitors without EDTA (Roche) at 1x final concentration was added to all lysis buffers (LB) prior to use. Each pellet was resuspended in LB1 (50mM Hepes pH7.5, 140mM NaCl, 1mM EDTA pH8, 10% glycerol, 0.75% NP-40, 0.25% triton X-100) and rotated at 4°C for 20 minutes. The cell suspension was homogenized using a douncer with a tight pestle with 3x 15 strokes, with short resting times on ice in between. The chromatin was pelleted by centrifugation at 1400g and 4°C for 5 minutes and resuspended in 2.5ml LB2 (200mM NaCl, 1mM EDTA pH8, 0.5mM EGTA pH8, 10mM Tris pH 8) per cell pellet. After 10 minutes rotation at 4°C, the centrifugation step was repeated and each pellet was resuspended in 1.5ml LB3 (1mM EDTA pH8, 0.5mM EGTA, 10mM Tris pH8, 100mM NaCl, 0.1% Na-deoxycholate, 0.5% N-lauroylsarcosine), transferred to 15ml Falcon tubes and sonicated using a W-450D Digital Sonifier (Branson) sonicator for 14 cycles of 10s on/50s off in a 4°C coldroom with tubes chilled in ice water. After sonication, 150ul of triton X-100 was added per tube, transferred to two 1.5ml Eppendorf tubes and debris was pelleted by centrifugation at 20,000g and 4°C for 10 minutes. The solubilized chromatin was then pooled and mixed thoroughly. After taking 50ul as an input control, the chromatin was distributed into 1.5ml aliquots, snap frozen and stored at -80C until use.  The input was reverse-crosslinked with the addition of 50ul 2x elution buffer (100mM Tris pH8, 20mM EDTA pH8, 2% SDS) and overnight (13h-15h) incubation at 65°C. The next day, 100ul of TE buffer were added to dilute the SDS of the elution buffer and RNA was digested using RNase A at a 0.2ug/ml final concentration at 37°C for 2 hours. Proteins were digested with the addition of 0.2ug/ml proteinase K and incubation at 55°C for 2 hours. The DNA was purified with two subsequent phenol:chloroform:isoamylalcohol (25:24:1, pH8) isolations and subsequent MinElute (Qiagen) purification. Purification was performed according to manufacturer’s recommendations, except that the columns were washed 2x with PE and air dried for at least 10 minutes before elution with EB. The input DNA was quantified using a NanoPhotometer (Implen) and the shearing was verified by running 600ng of DNA on a 1.5% agarose gel. For sonication to be considered successful and sonicated chromatin to be usable for ChIP, the bulk DNA of the input had to be located between 100bp-500bp.
For ChIP, 50µl or 100ul (for β-catenin) of protein-coated Dynal beads (Life Technologies) were washed 3x using 1ml of blocking solution (PBS containing 0.5% BSA), resuspended in 500µl of blocking solution together with approximately 5µg or 10 µg (for β-catenin) antibody and rotated overnight at 4°C. The antibodies used were a goat polyclonal anti-T (Santa Cruz, sc-17743), a goat polyclonal anti-Sox2 (R&D Systems, AF2018), a rabbit polyclonal anti-β-catenin (Invitrogen, 71-2700) and a rabbit polyclonal anti-Tbx6 (this paper). The next day, three further washes using 1ml blocking buffer were performed and the beads were resuspended in 100µl blocking buffer. Chromatin equivalent to approximately 25x106 or 50x106 (for β-catenin) cells was added and rotated overnight at 4°C. The following day, washing steps (6, 8 and 9 washes for Tbx6, Sox2 / β-catenin and T respectively) were performed using 1ml RIPA buffer (50mM Hepes pH7.6, 500mM LiCl, 1mM EDTA pH8, 1% NP-40, 0.7% Na-Deoxycholate) containing 1x complete protease inhibitors without EDTA and finally with 1ml of TEN (10mM Tris pH8, 1mM EDTA pH8, 50mM NaCl). The immunoprecipates were eluted in two subsequent steps using 100µl of 1x elution buffer (50mM Tris pH8, 10mM EDTA pH8, 1% SDS) and incubation at 65°C while shaking for 10 minutes each. The eluates were combined and incubated overnight (13h-15h) at 65°C under agitation. The next day, 200µl of TE were added to dilute the SDS of the elution buffer and the ChIPs were purified as described for the input above. ChIP DNA was quantified using the Qubit (Life Technologies) DNA HS assay.
The ChIP-Seq libraries were generated using the TruSeq ChIP library kit (Illumina) with the following modifications. After adapter ligation, 0.95x of AMPure XP beads (Beckman Coulter) were used for a single purification and the DNA was eluted using 15µl of resuspension buffer (RSB, Illumina). In the case of Tbx6, 14µl of the eluate were used for PCR pre-amplification in order to convert adapters into double-stranded DNA. After the addition of 1µl primer mix (25mM each, Primer 1: 5’- AATGATACGGCGACCACCGA*G-3’; Primer2: 5’- CAAGCAGAAGACGGCATACGA*G-3’) and 15µl 2x Kapa HiFi HotStart Ready Mix (Kapa Biosystems), amplification was performed for 45 seconds at 98°C, 5 cycles of [15 seconds at 98°C, 30 seconds at 63°C and 30 seconds at 72°C] and a final 1 minute incubation at 72°C. The PCR products were purified using 0.95x of beads and eluted using 21µl of RSB. Libraries were directly amplified for 18 cycles. The libraries were quantified using the Qubit DNA HS assay and the library size was validated using DNA HS bioanalyzer chips.


title: ChIP T


title: ATAC T+ S+ WT TS12


title: ATAC T+ WT TS12


title: ATAC T+ S+ WT TS13


title: ATAC T+ S+ KO TS13

source_name: T+ / Sox2+ FACS sorted cells from caudal ends of T2J/2J embryos


title: ATAC T+ S+ KO TS12


title: ZR1186_10: Calf born to a cow with no supplementation of methyldonors

city: Caldes de Montbui

first: Alex

last: Bach

state: Barcelona

address: Torre Marimon

department: Ruminant Production

country: Spain

institute: IRTA

description: library stategy: Methyl-miniSeq

data_processing:
Libraries were prepared from 200-500 ng of genomic DNA digested with 60 units of TaqαI and 30 units of MspI (NEB) sequentially and then extracted with Zymo Research (ZR) DNA Clean & ConcentratorTM-5 kit (Cat#: D4003)
Adaptor-ligated fragments of 150–250 bp and 250–350 bp in size were recovered from a 2.5% NuSieve 1:1 agarose gel
Sequence reads from bisulfite-treated EpiQuest libraries were identified using standard Illumina base-calling software and Bismark
The --non_directional parameter was applied while running Bismark. All other parameters were set to default. Filled-in nucleotides were trimmed off when doing methylation calling.
Only CpGs with a sequencing depth of >10x and a methylation difference >5% in all samples were considered for further analysis
Genome_build: Bovine assembly UMD3.1.1/bosTau8

organism: Bos taurus

source_name: Peripheral blood mononuclear cells

extract_protocol:
DNA was extracted from isolated PBMC with DNeasy Blood & Tissue Kit (Qiagen, Madrid, Spain) and submitted to a quality control test using a Nanodrop-1000 (JH Bio, USA) and 0.8% agarose gel to determine the purity and quantity.
Libraries were prepared from 200-500 ng of genomic DNA digested with 60 units of TaqαI and 30 units of MspI (NEB) sequentially and then extracted with Zymo Research (ZR) DNA Clean & ConcentratorTM-5 kit (Cat#: D4003). Fragments were ligated to pre-annealed adapters containing 5’-methyl-cytosine instead of cytosine according to Illumina’s specified guidelines (www.illumina.com). Adaptor-ligated fragments of 150–250 bp and 250–350 bp in size were recovered from a 2.5% NuSieve 1:1 agarose gel (ZymocleanTM Gel DNA Recovery Kit, ZR Cat#: D4001). The fragments were then bisulfite-treated using the EZ DNA Methylation-LightningTM Kit (ZR, Cat#: D5020). Preparative-scale PCR was performed and the resulting products were purified (DNA Clean & ConcentratorTM - ZR, Cat#D4005) for sequencing on an Illumina HiSeq. Reads were quality-controlled with FastQC (Babraham informatics, Cambridge, UK)
and trimmed to remove adapter sequences with Trim Galore (Babraham informatics, Cambridge, UK). Then, processed reads
were mapped to the in silico bisulfite-converted reference genome (Bovine assembly UMD3.1.1/bosTau8)

tag: development stage

value: Newborn calves

value: Peripheral blood mononuclear cells


title: ZR1186_9: Calf born to a cow with supplementation of methyldonors

description: library stategy: Methyl-miniSeq


title: ATAC S+ WT TS13

source_name: Sox2+ FACS sorted cells from caudal ends of WT embryos


title: ATAC T+ KO TS12

source_name: T+ FACS sorted cells from caudal ends of T2J/2J embryos


title: ATAC S+ WT TS12


title: ATAC S+ KO TS13

source_name: Sox2+ FACS sorted cells from caudal ends of T2J/2J embryos


title: ATAC S+ KO TS12


title: ATAC T+ KO TS13


title: ZR1186_6: Calf born to a cow with no supplementation of methyldonors

description: library stategy: Methyl-miniSeq


title: ZR1186_8: Calf born to a cow with supplementation of methyldonors

description: library stategy: Methyl-miniSeq


title: 2ndOrderSpared_L4_4

city: Nijmegen

first: Koen

last: Kole

address: Heyendaalsweg 135

department: Neurophysiology

country: Netherlands

institute: Donders Institute for Brain Cognition and Behaviour

data_processing:
Basecalls were performed using Illumina RTA version 2.4.6.
Reads were trimmed to 75 bp using Trimmomatic version 0.36, with the configurations: PE (Paired End) and CROP:75
Reads were mapped to the mm10 genome using STAR version 2.5.2a; output for all samples can be found in Genecounts_Colourcoded.xlsx
Genome_build: mm10
Supplementary_files_format_and_content: Genecounts_Colourcoded.xlsx contains read counts for all samples after STAR mapping; colours indicate experimental groups (n=4 for all)

source_name: Barrel Cortex A/E Columns Layer 4

extract_protocol:
Tissue samples originating from the same animal, columns and layer were pooled. Tissue was quickly dissolved in Qiazol (Qiagen #79306) and RNA was isolated using the miRNeasy Mini kit (Qiagen #217004). The extracted RNA was DNAse treated (Thermo Scientific, #EN0521) and finally cleaned up using RNeasy MinElute kit (Qiagen #74204).
The cDNA library was generated using the non-stranded NEBNext Ultra RNA Library Preparation  Kit for Illumina (New England Biolabs, catalogue #E7530). For the library enrichment, 13-14 PCR cycles were performed

tag: group

value: Spared, second order

value: Postnatal day 23-24


title: ZR1186_7: Calf born to a heifer with no supplementation of methyldonors

description: library stategy: Methyl-miniSeq


title: 2ndOrderSpared_L4_2


title: ZR1186_5: Calf born to a heifer with supplementation of methyldonors

description: library stategy: Methyl-miniSeq


title: ZR1186_4: Calf born to a heifer with no supplementation of methyldonors

description: library stategy: Methyl-miniSeq


title: ZR1186_3: Calf born to a cow with supplementation of methyldonors

description: library stategy: Methyl-miniSeq


title: 2ndOrderSpared_L4_1


title: 1stOrderSpared_L4_3

source_name: Barrel Cortex B/D Columns Layer 4

value: Spared, first order


title: 1stOrderSpared_L4_4


title: ZR1186_2: Calf born to a heifer with no supplementation of methyldonors

description: library stategy: Methyl-miniSeq


title: 1stOrderSpared_L4_2


title: ZR1186_1: Calf born to a cow with supplementation of methyldonors

description: library stategy: Methyl-miniSeq


title: Deprived_L4_4

source_name: Barrel Cortex C Column Layer 4

value: Deprived


title: 1stOrderSpared_L4_1


title: Deprived_L4_2


title: Deprived_L4_3


title: Deprived_L4_1


title: 2ndOrderSpared_L4_3


title: Control_L4_2

value: Control


title: Control_L4_4


title: Control_L4_3


title: Control_L4_1


title: 2ndOrderSpared_L2/3_2

source_name: Barrel Cortex A/E Columns Layer 2/3


title: 2ndOrderSpared_L2/3_3


title: 2ndOrderSpared_L2/3_4


title: 2ndOrderSpared_L2/3_1


title: 1stOrderSpared_L2/3_4

source_name: Barrel Cortex B/D Columns Layer 2/3


title: Deprived_L2/3_2

source_name: Barrel Cortex C Column Layer 2/3


title: Deprived_L2/3_1


title: 1stOrderSpared_L2/3_1


title: 1stOrderSpared_L2/3_2


title: Deprived_L2/3_3


title: Control_L2/3_2


title: Control_L2/3_4


title: Control_L2/3_1


title: Deprived_L2/3_4


title: Control_L2/3_3


title: Control 38

city: Baltimore

first: Carrie

last: Wright

address: 855 N Wolfe St

institute: Lieber Institute for Brain Development

description:
sample51
RNAseq of Total RNA from 100mg of homogenate tissue.
Processed data file: log2RPKMcounts.xlsx
Processed data file: log2RPKMcounts.rda

data_processing:
Basecalls performed using CASAVA version 1.8.2.
Alignment performed using TopHat (v2.0.4) to enforce strand specificity and to align the sequencing reads to known transcripts of the Ensembl Build GRCh37.67.
FeatureCounts (v1.4.4) to count the total number of reads overlapping each gene using the default settings, with paired-end and reverse-stranded counting specified using the Ensembl Build GRCh37.67 gtf file.
Counts were merged from both reads of the paired-end sequencing and normalized by library size and coding gene length to form Reads Per Kilobase of Gene per Million mapped reads (RPKM) values.
RPKM values were transformed using log2 after applying an offset of 1 to each count.
Normalized expression values were then filtered for genes with mean normalized expression values less than 0.5.
Genome_build: GRCh37.67/hg19
Supplementary_files_format_and_content: log2RPKMcounts.xlsx: Excel file, normalized log2(RPKM +1) counts.
Supplementary_files_format_and_content: log2RPKMcounts.rda: .rda format created in R, normalized log2(RPKM +1) counts.

organism: Homo sapiens

source_name: Postmortem Brain, DLPFC

extract_protocol:
RNeasy kit (Qiagen).
TruSeq RNA Sample Preparation v2 kit from Illumina.

tag: disease status

value: Healthy control

value: Postmortem brain, dorsolateral prefrontal cortex (DLPFC)

tag: brainid

tag: rin

tag: Sex

value: Female

tag: race

value: CAUC

tag: totalnummappedreads

tag: totalnumreads

tag: ph

tag: percentexonicmapping


title: Control 39

description:
sample52
RNAseq of Total RNA from 100mg of homogenate tissue.
Processed data file: log2RPKMcounts.xlsx
Processed data file: log2RPKMcounts.rda

tag: age

value: Male


title: ASD 12

description:
sample49
RNAseq of Total RNA from 100mg of homogenate tissue.
Processed data file: log2RPKMcounts.xlsx
Processed data file: log2RPKMcounts.rda

value: Autism spectrum disorder (ASD)


title: ASD 11

description:
sample48
RNAseq of Total RNA from 100mg of homogenate tissue.
Processed data file: log2RPKMcounts.xlsx
Processed data file: log2RPKMcounts.rda

value: AA


title: ASD 10

description:
sample47
RNAseq of Total RNA from 100mg of homogenate tissue.
Processed data file: log2RPKMcounts.xlsx
Processed data file: log2RPKMcounts.rda


title: ASD 9

description:
sample46
RNAseq of Total RNA from 100mg of homogenate tissue.
Processed data file: log2RPKMcounts.xlsx
Processed data file: log2RPKMcounts.rda


title: frozen fixed HEK293T and NIH/3T3 cell mixture (16 x 80 sci-RNA-seq)

city: Seattle

first: Junyue

last: Cao

state: WA

address: Foege Building S-210, 3720 15th Ave NE

department: Department of Genome Sciences

institute: University of Washington

description: sciRNAseq with HEK293T and NIH/3T3 cells (cells are methanol fixed and frozen in -80 degree for 4 days)

data_processing:
Base calls were converted to fastq format and demultiplexed using Illumina’s bcl2fastq/ 2.16.0.10 tolerating one mismatched base in barcodes (edit distance (ED) < 2)
RT barcode (10bp) and UMI (8bp) sequence in Read1 is attached to the read name of Read2. Reads not matching to the expected RT barcodes were discarded. Read2 were then adaptor clipped using trim_galore/0.4.1 with default settings.
Trimmed reads were mapped to the human reference genome (hg19), mouse reference genome (mm10), C.elegans reference genome (PRJNA13758) or a chimeric reference genome of hg19, mm10 and PRJNA13758, using STAR/v 2.5.2b (38) with default settings and gene annotations (GENCODE V19 for human; GENCODE VM11 for mouse, WormBase PRJNA13758.WS253.canonical_gene set for C.elegans).
Uniquely mapping reads were extracted, and duplicates were removed using the unique molecular identifier (UMI) sequence (ED < 2, including insertions and deletions), reverse transcription (RT) index, and read 2 end-coordinate.
Mapped reads were split into constituent cellular indices by further demultiplexing reads using the RT index (ED < 2, including insertions and deletions).
To generate digital expression matrices, we calculated the number of strand-specific UMIs mapping to the exonic and intronic regions of each gene, for each cell; generally, fewer than 3% of total UMIs strand-specifically mapped to multiple genes. For multi-mapped reads, reads were assigned to the closest gene, except in cases where another intersected gene fell within 100 bp to the end of the closest gene, in which case the read was discarded.
Genome_build: human reference genome (hg19), mouse reference genome (mm10), C.elegans reference genome (PRJNA13758)
Supplementary_files_format_and_content: Processed data file is a sparsematrix in R: each row corresponds to genes, and each column corresponds to single cell, number in the matrix is the UMI count data.

organism:
Homo sapiens
Mus musculus

source_name: cell line

growth_protocol: All cultured cell lines were cultured at 37°C with 5% CO2, and were maintained in high glucose DMEM (Gibco cat. no. 11965) supplemented with 10% FBS and 1X Pen/Strep (Gibco cat. no. 15140122; 100U/ml penicillin, 100µg/ml streptomycin).

extract_protocol:
All cell lines were trypsinized, spun down at 300xg for 5 min (4°C). and washed once in 1X PBS. L2 stage worms were disscociated with SDS-DTT solution and pronase to generate single cell suspension. Detailed protocol was described in "J. Cao et al., Comprehensive single cell transcriptional profiling of a multicellular organism by combinatorial indexing. bioRxiv, 104844 (2017)."
library construction protocol was described in "J. Cao et al., Comprehensive single cell transcriptional profiling of a multicellular organism by combinatorial indexing. bioRxiv, 104844 (2017)."

treatment_protocol: For c.elegans, a synchronized L2 population was obtained by two cycles of bleaching gravid adults to isolate fertilized eggs allowing the eggs to hatch in the absence of food to generate a population of starved L1 animals.

tag: cell line

value: HEK293T and NIH/3T3


title: ASD 8

description:
sample45
RNAseq of Total RNA from 100mg of homogenate tissue.
Processed data file: log2RPKMcounts.xlsx
Processed data file: log2RPKMcounts.rda


title: ASD 7

description:
sample44
RNAseq of Total RNA from 100mg of homogenate tissue.
Processed data file: log2RPKMcounts.xlsx
Processed data file: log2RPKMcounts.rda


title: HEK293T, HeLa S3 and NIH/3T3 cell and nuclei mixture (96 x 96 sci-RNA-seq)

description: sciRNAseq with HEK293T, Hela S3 and NIH/3T3 cells

value: HEK293T, HeLa S3 and NIH/3T3


title: 1stOrderSpared_L2/3_3

tag: strain


title: HEK293T, HeLa S3 and NIH/3T3 cell mixture (384 x 384 sci-RNA-seq)


title: ASD 6

description:
sample43
RNAseq of Total RNA from 100mg of homogenate tissue.
Processed data file: log2RPKMcounts.xlsx
Processed data file: log2RPKMcounts.rda


title: ASD 5

description:
sample42
RNAseq of Total RNA from 100mg of homogenate tissue.
Processed data file: log2RPKMcounts.xlsx
Processed data file: log2RPKMcounts.rda


title: ASD 4

description:
sample41
RNAseq of Total RNA from 100mg of homogenate tissue.
Processed data file: log2RPKMcounts.xlsx
Processed data file: log2RPKMcounts.rda


title: TS1

city: Beijing

first: Zijing

last: Liu

address: Fengtai District, #20 East St.

country: China

institute: Beijing Institute of Biotechnology

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 mm10 whole genome using STAR v2.5.2a
Reads Per Kilobase of exon per Megabase of library size (RPKM) were calculated using a protocol from Chepelev et al., Nucleic Acids Research, 2009. In short, exons from all isoforms of a gene were merged to create one meta-transcript. The number of reads falling in the exons of this meta-transcript were counted and normalized by the size of the meta-transcript and by the size of the library.
Genome_build: mm10
Supplementary_files_format_and_content: tab-delimited text files include RPKM values for each Sample ...

source_name: dorsal root ganglion

extract_protocol:
Total RNA of DRGs were extracted with RNeasy Mini Kit (Qiagen) under the manufacturer’s protocol
RNA libraries were prepared for sequencing using standard Illumina protocols

tag: genotype

value: about 2 months old


title: TS3


title: ASD 3

description:
sample40
RNAseq of Total RNA from 100mg of homogenate tissue.
Processed data file: log2RPKMcounts.xlsx
Processed data file: log2RPKMcounts.rda


title: ASD 2

description:
sample39
RNAseq of Total RNA from 100mg of homogenate tissue.
Processed data file: log2RPKMcounts.xlsx
Processed data file: log2RPKMcounts.rda


title: ASD 13

description:
sample50
RNAseq of Total RNA from 100mg of homogenate tissue.
Processed data file: log2RPKMcounts.xlsx
Processed data file: log2RPKMcounts.rda


title: WT3

value: wild type


title: ASD 1

description:
sample38
RNAseq of Total RNA from 100mg of homogenate tissue.
Processed data file: log2RPKMcounts.xlsx
Processed data file: log2RPKMcounts.rda


title: Control 35

description:
sample35
RNAseq of Total RNA from 100mg of homogenate tissue.
Processed data file: log2RPKMcounts.xlsx
Processed data file: log2RPKMcounts.rda


title: Control 36

description:
sample36
RNAseq of Total RNA from 100mg of homogenate tissue.
Processed data file: log2RPKMcounts.xlsx
Processed data file: log2RPKMcounts.rda


title: Control 34

description:
sample34
RNAseq of Total RNA from 100mg of homogenate tissue.
Processed data file: log2RPKMcounts.xlsx
Processed data file: log2RPKMcounts.rda


title: Control 33

description:
sample33
RNAseq of Total RNA from 100mg of homogenate tissue.
Processed data file: log2RPKMcounts.xlsx
Processed data file: log2RPKMcounts.rda


title: HEK293T and NIH/3T3 cell mixture (16 x 84 sci-RNA-seq)

description: sciRNAseq with HEK293T and NIH/3T3 cells


title: Control 32

description:
sample32
RNAseq of Total RNA from 100mg of homogenate tissue.
Processed data file: log2RPKMcounts.xlsx
Processed data file: log2RPKMcounts.rda


title: Control 31

description:
sample31
RNAseq of Total RNA from 100mg of homogenate tissue.
Processed data file: log2RPKMcounts.xlsx
Processed data file: log2RPKMcounts.rda


title: Control 30

description:
sample30
RNAseq of Total RNA from 100mg of homogenate tissue.
Processed data file: log2RPKMcounts.xlsx
Processed data file: log2RPKMcounts.rda


title: Control 29

description:
sample29
RNAseq of Total RNA from 100mg of homogenate tissue.
Processed data file: log2RPKMcounts.xlsx
Processed data file: log2RPKMcounts.rda


title: Control 28

description:
sample28
RNAseq of Total RNA from 100mg of homogenate tissue.
Processed data file: log2RPKMcounts.xlsx
Processed data file: log2RPKMcounts.rda


title: Control 27

description:
sample27
RNAseq of Total RNA from 100mg of homogenate tissue.
Processed data file: log2RPKMcounts.xlsx
Processed data file: log2RPKMcounts.rda


title: TS2


title: Control 25

description:
sample25
RNAseq of Total RNA from 100mg of homogenate tissue.
Processed data file: log2RPKMcounts.xlsx
Processed data file: log2RPKMcounts.rda


title: YH_MB-seq

first: Yueyi

last: Shi

address: No.28, Fuxing Road

department: Department of Stomatology

institute: Chinese PLA General Hospital

data_processing:
library strategy: MB-seq
Basecalls performed using CASAVA version 1.8
The low quality reads and adapter sequences were firstly removed from the raw reads. The quality of resulted clean reads was checked by FastQC
MB-seq/MRB-seq reads were aligned to the hg18 genome assembly using SOAP2.21 with two mismatches allowed in the seed (40 bp)
BSMAP v2.72 was used to process the alignment results to identify the methylcytosines where at least three reads were covered
Genome_build: UCSC hg18
Supplementary_files_format_and_content: Tab-delimited text files include methylation information of each chromosone for each Sample.Scores represent chromosome,position,strand,context ratio,eff_CT_count,C_count CT_count,rev_G_count,rev_GA_count,CI_lower,CI_upper

source_name: YH cells

extract_protocol:
DNA from YH’s peripheral blood mononuclear cells was extracted by QIAamp DNA Blood Mini Kit (QIAGEN).
DNA from MeDIP and MeDIP repeat-removal captures were treated with sodium bisulfite using the EZ DNA Methylation-Gold Kit (Zymo Research), respectively.

value: YH

tag: cell type

value: peripheral blood mononuclear cells (PBMCs)


title: Control 24

description:
sample24
RNAseq of Total RNA from 100mg of homogenate tissue.
Processed data file: log2RPKMcounts.xlsx
Processed data file: log2RPKMcounts.rda

tag: tissue


title: YH_MRB-seq

data_processing:
library strategy: MRB-seq
Basecalls performed using CASAVA version 1.8
The low quality reads and adapter sequences were firstly removed from the raw reads. The quality of resulted clean reads was checked by FastQC
MB-seq/MRB-seq reads were aligned to the hg18 genome assembly using SOAP2.21 with two mismatches allowed in the seed (40 bp)
BSMAP v2.72 was used to process the alignment results to identify the methylcytosines where at least three reads were covered
Genome_build: UCSC hg18
Supplementary_files_format_and_content: Tab-delimited text files include methylation information of each chromosone for each Sample.Scores represent chromosome,position,strand,context ratio,eff_CT_count,C_count CT_count,rev_G_count,rev_GA_count,CI_lower,CI_upper


title: WT2


title: Control 26

description:
sample26
RNAseq of Total RNA from 100mg of homogenate tissue.
Processed data file: log2RPKMcounts.xlsx
Processed data file: log2RPKMcounts.rda


title: Control 23

description:
sample23
RNAseq of Total RNA from 100mg of homogenate tissue.
Processed data file: log2RPKMcounts.xlsx
Processed data file: log2RPKMcounts.rda


title: Control 37

description:
sample37
RNAseq of Total RNA from 100mg of homogenate tissue.
Processed data file: log2RPKMcounts.xlsx
Processed data file: log2RPKMcounts.rda


title: WT Input, rep2

city: New Haven

first: Huabing

last: Li

state: CONNECTICUT

address: 1 Gilbert Street, S570

department: Immunobiology

institute: Yale University School of Medicine

description:
mRNA
Processed data file: control_norm.wig

data_processing:
Alignment (Tophat v2.0.14).
Generation of normalized abundance measurements (Cufflinks v2.2.1).
Genome_build: mm10 (GRCm38)
Supplementary_files_format_and_content: control_norm.wig: Wig file; Output after Tophat and Cuffdiff contains all the gene reads for input.
Supplementary_files_format_and_content: RIP_norm.wig: Wig file; Output after Tophat and Cuffdiff contains all the gene reads enriched for m6A marker.

source_name: CD4+ T cells, WT

extract_protocol:
Total RNAs were isolated from the pure CD4+ T cells, subjected to standard m6A RIP protocol.
10 ng RNA from input and enriched IP RNA samples were used for library preparation with the SMARTer Stranded Total RNA-Seq Kit – Pico Input Mammalian (Clontech) according to the manufacturer’s instructions.

treatment_protocol: CD4+ T cells were isolated by StemCell CD4+ T cell isolation kit of spleen and lymph nodes WT mice.

tag: strain/background

tag: genotype/variation

value: wildtype

value: CD4+ T cells

tag: antibody


title: WT RIP, rep1

description:
mRNA
Processed data file: RIP_norm.wig


title: KO Ribosome Profiling, rep1

description:
mRNA
Sample_6-KO1_006
KO1_FPKM
Processed data file: Ribosome_Profiling_March152017.xlsx

data_processing:
The Tuxedo suite.
GeneraHTseq was used to calculate translation efficiency.
Genome_build: mm10 (GRCm38)
Supplementary_files_format_and_content: Input_Mett3gene_exp.diff: Tab-delimited text file; output of the input after Tophat and Cuffdiff including all the gene reads.
Supplementary_files_format_and_content: Ribosome_Profiling_March152017.xlsx: Excel file; output of the ribosome profiling after Tuxedo suite including all the gene reads.

source_name: CD4+ T cells, Mettl3 KO

extract_protocol:
1/10 of the cell lysate was used to isolate RNA for preparing input RNA library, and the remaining was used to isolate ribosome protected fragments and library preparation.
Libraries were generated with ribosome protected fragments and Input RNAs using Illumina TruSeq Ribo Profile (Mammalian) Kit. Briefly, after cleaning up ribosome protected fragments, the ends were repaired using PNK, then ligated 3' adapters, reverse transcribed the libraries. The cDNA libraries were PAGE purified for the 70-100 nt fragments, then circularized, and PCR amplified for 9 cycles. The amplified libraries were purified again by PAGE for the 140~160bp products, and then sent for sequencing.

treatment_protocol: 50 million naïve T cells were isolated from WT and Mettl3 KO mice, and washed and treated with 0.1 mg/ml cycloheximide for 1min.

value: Mettl3 knockout


title: KO Ribosome Profiling, rep2

description:
mRNA
Sample_7-KO2_007
KO2_FPKM
Processed data file: Ribosome_Profiling_March152017.xlsx


title: Control 22

description:
sample22
RNAseq of Total RNA from 100mg of homogenate tissue.
Processed data file: log2RPKMcounts.xlsx
Processed data file: log2RPKMcounts.rda

tag: disease status

tag: brainid


title: WT Input, rep1


title: WT Ribosome Profiling, rep2

description:
mRNA
Sample_5-WT2_005
WT4_FPKM
Processed data file: Ribosome_Profiling_March152017.xlsx


title: Control 20

description:
sample20
RNAseq of Total RNA from 100mg of homogenate tissue.
Processed data file: log2RPKMcounts.xlsx
Processed data file: log2RPKMcounts.rda

tag: rin

tag: Sex


title: Control 19

description:
sample19
RNAseq of Total RNA from 100mg of homogenate tissue.
Processed data file: log2RPKMcounts.xlsx
Processed data file: log2RPKMcounts.rda

tag: race

tag: totalnummappedreads


title: Ribosome Profiling Input for WT, rep2

description:
mRNA
Sample_Total_WT2_002
Processed data file: Input_Mett3gene_exp.diff


title: WT Ribosome Profiling, rep1

description:
mRNA
Sample_4-WT1_004
WT3_FPKM
Processed data file: Ribosome_Profiling_March152017.xlsx


title: Ribosome Profiling Input for KO, rep2

description:
mRNA
Sample_Total_KO2_011
Processed data file: Input_Mett3gene_exp.diff


title: Control 21

description:
sample21
RNAseq of Total RNA from 100mg of homogenate tissue.
Processed data file: log2RPKMcounts.xlsx
Processed data file: log2RPKMcounts.rda

tag: totalnumreads

tag: ph


title: Mett3_Naive_WT3_006

description: Processed data file: Mett3_gene_exp.diff

data_processing:
Alignment (Tophat v2.0.14).
Generation of normalized abundance measurements (Cufflinks v2.2.1).
Genome_build: mm10 (GRCm38)
Supplementary_files_format_and_content: Mett3_gene_exp.diff: Tab-delimited text file; output after Tophat & Cuffdiff including all the gene reads.

source_name: Naïve T cells, wildtype

extract_protocol:
Total RNA was purified with RiboZero kit.
RNA libraries were prepared for sequencing using standard Illumina protocols.

treatment_protocol: Naïve T cells were obtained by StemCell CD4+ T cell isolation kit of spleen from Mettl3 KO and WT mice followed by FACS sorting (CD4+CD25-CD45RB-Hi).

value: Wildtype

value: Naive T cells


title: Control 18

description:
sample18
RNAseq of Total RNA from 100mg of homogenate tissue.
Processed data file: log2RPKMcounts.xlsx
Processed data file: log2RPKMcounts.rda

value: Healthy control

value: Postmortem brain, dorsolateral prefrontal cortex (DLPFC)

tag: age

tag: percentexonicmapping


title: WT1

tag: strain


title: WT RIP, rep2

description:
mRNA
Processed data file: RIP_norm.wig


title: Mett3_Naive_WT4_012


title: Mett3_Naive_KO2_005

source_name: Naïve T cells, Mettl3 KO

value: Mettl3 KO


title: Mett3_Naive_KO1_004


title: Control 17

description:
sample17
RNAseq of Total RNA from 100mg of homogenate tissue.
Processed data file: log2RPKMcounts.xlsx
Processed data file: log2RPKMcounts.rda

value: Male

value: CAUC


title: Control 16

description:
sample16
RNAseq of Total RNA from 100mg of homogenate tissue.
Processed data file: log2RPKMcounts.xlsx
Processed data file: log2RPKMcounts.rda

value: AA


title: Control 15

description:
sample15
RNAseq of Total RNA from 100mg of homogenate tissue.
Processed data file: log2RPKMcounts.xlsx
Processed data file: log2RPKMcounts.rda


title: Wild-type sample 2

city: Seoul

first: Kihoon

last: Han

address: 73, Inchon-ro, Seongbuk-gu

department: Neuroscience

country: South Korea

institute: Korea University College of Medicine

data_processing:
Pre-processing of raw reads was carried out using Trimmomatic (version 0.35, options: LEADING:3 TRAILING:3 MAXINFO:80:0.4 MINLEN:36)
The trimmed reads were mapped to the Mus musculus genome (GRCm38) using TopHat2 (version 2.1.0, default options).
The gene-level read counts were created from the aligned reads by HTSeq then normalized on library size factors by using DEseq2 package.
Differential gene expression analysis was carried out using DEseq2 and the P-values were adjusted for multiple testing with the Benjamini-Hochberg correction.
Genome_build: GRCm38
Supplementary_files_format_and_content: tab-delimited text file includes normalized read count matrix calculated by using HTSeq and DESeq2

source_name: Wild-type brain

extract_protocol:
RNA samples were prepared for sequencing using a TruSeq RNA Sample Prep Kit v2 (Illumina) according to the manufacturer’s instructions.
RNA libraries were prepared for sequencing using standard Illumina protocols

value: striatum

value: 12 week

value: WT


title: Transgenic sample 3

source_name: Transgenic brain

value: TG


title: Wild-type sample 3


title: Transgenic sample 1


title: Wild-type sample 1


title: Transgenic sample 2


title: Control 13

description:
sample13
RNAseq of Total RNA from 100mg of homogenate tissue.
Processed data file: log2RPKMcounts.xlsx
Processed data file: log2RPKMcounts.rda


title: Ribosome Profiling Input for KO, rep1

description:
mRNA
Sample_Total_KO1_003
Processed data file: Input_Mett3gene_exp.diff


title: Control 14

description:
sample14
RNAseq of Total RNA from 100mg of homogenate tissue.
Processed data file: log2RPKMcounts.xlsx
Processed data file: log2RPKMcounts.rda


title: Ribosome Profiling Input for WT, rep1

description:
mRNA
Sample_Total_WT1_001
Processed data file: Input_Mett3gene_exp.diff


title: RNA Tbx6+ KO TS14

description:
total RNA
embryos obtained from tatraploid complementation of embryonic stem cells
FPKM_T_Tbx6_KO.txt

source_name: Tbx6+ FACS sorted cells from caudal ends of WT embryos

extract_protocol:
Total RNA from approximately 20,000 cells was isolated using the RNeasy Micro kit (Qiagen) and genomic DNA was digested on column with the addition of an extra 1ul of RNase-free DNase I (Roche) to ensure complete digestion.
The RNA from approximately 20,000 cellswas quantified using the Qubit RNA HS Assay and the integrity was verified using Bioanalyzer RNA Pico chips. Approximately 80-100ng of total RNA was used for the generation of strand-specific RNA-Seq libraries using the ScriptSeq v2 (Epicentre) low input libarary preparation kit according to manufacturer’s instructions. During library preparation, the purification after rRNA-depletion was performed using RNeasy Micro columns and the cDNA was purified using the MinElute PCR Purification Kit (Qiagen). The library was finally amplified with 15 PCR cycles.

tag: developmental stage


title: RNA Tbx6+ WT TS14

source_name: Tbx6+ FACS sorted cells from caudal ends of Tbx6-/- embryos


title: RNA T+ WT TS13

source_name: T+ FACS sorted cells from caudal ends of WT embryos


title: RNA T+ KO TS12

source_name: T+ FACS sorted cells from caudal ends of T2J/2J embryos


title: Control 11

description:
sample11
RNAseq of Total RNA from 100mg of homogenate tissue.
Processed data file: log2RPKMcounts.xlsx
Processed data file: log2RPKMcounts.rda


title: RNA T+ KO TS13


title: RNA T+ WT TS12


title: Control 10

description:
sample10
RNAseq of Total RNA from 100mg of homogenate tissue.
Processed data file: log2RPKMcounts.xlsx
Processed data file: log2RPKMcounts.rda


title: RNA NMP T+

description:
total RNA
embryos obtained from tatraploid complementation of embryonic stem cells
FPKM_NMP.txt

extract_protocol:
Total RNA from approximately 2,500 cells was isolated using the RNeasy Micro kit (Qiagen) and genomic DNA was digested on column with the addition of an extra 1ul of RNase-free DNase I (Roche) to ensure complete digestion.
RNA-Seq libraries with the RNA equivalent of approximately 1,000 cells were generated using the TotalScript RNA-Seq Kit (Epicentre) with random hexamers according to manufacturer’s instructions with slight modifications. Briefly, all reagents during the cDNA synthesis steps were scaled down in order to achieve a total volume of 39µl, allowing all of the material to be used in the subsequent tagmentation procedure. To further improve strand-specificity of the libraries, Actinomycin D was added to the first strand cDNA synthesis reaction at a 5ng/µl final concentration and the libraries were finally amplified using 14 cycles.


title: Control 7

description:
sample7
RNAseq of Total RNA from 100mg of homogenate tissue.
Processed data file: log2RPKMcounts.xlsx
Processed data file: log2RPKMcounts.rda

value: Female


title: Control 9

description:
sample9
RNAseq of Total RNA from 100mg of homogenate tissue.
Processed data file: log2RPKMcounts.xlsx
Processed data file: log2RPKMcounts.rda


title: c.elegans cells (L2 stage) 2

description: sciRNAseq with c.elegans cells (cell name with "cele-all": during sorting, c.elegans cells were gated to include normal c.elegans cells and polyploid cells; cell name with "cele-high": during sorting, c.elegans cells were gated to inlcude polyploid cells only; cell name with "cele-extra-high": during sorting, c.elegans cells were gated to include only polyploid c.elegans cells and human cells

organism: Caenorhabditis elegans

source_name: whole c.elegans (L2 stage)

value: A C. elegans strain (RW12139 stIs11435(unc-120::H1-Wcherry;unc-119(+));unc-119(tm4063)) carrying an integrated Punc-120::mCherry gene in a wild type background was used


title: RNA NMP S+

source_name: Sox2+ FACS sorted cells from caudal ends of WT embryos


title: Control 6

description:
sample6
RNAseq of Total RNA from 100mg of homogenate tissue.
Processed data file: log2RPKMcounts.xlsx
Processed data file: log2RPKMcounts.rda


title: Control 2

description:
sample2
RNAseq of Total RNA from 100mg of homogenate tissue.
Processed data file: log2RPKMcounts.xlsx
Processed data file: log2RPKMcounts.rda


title: Control 5

description:
sample5
RNAseq of Total RNA from 100mg of homogenate tissue.
Processed data file: log2RPKMcounts.xlsx
Processed data file: log2RPKMcounts.rda


title: Control 1

description:
sample1
RNAseq of Total RNA from 100mg of homogenate tissue.
Processed data file: log2RPKMcounts.xlsx
Processed data file: log2RPKMcounts.rda


title: Control 4

description:
sample4
RNAseq of Total RNA from 100mg of homogenate tissue.
Processed data file: log2RPKMcounts.xlsx
Processed data file: log2RPKMcounts.rda


title: Control 3

description:
sample3
RNAseq of Total RNA from 100mg of homogenate tissue.
Processed data file: log2RPKMcounts.xlsx
Processed data file: log2RPKMcounts.rda


title: HOXB13.sample1.unbound

city: Palo Alto

first: Zheng

last: Zuo

address: Stanford

department: Genetics

institute: Stanford university

description: In vitro protein-DNA binding reactions

data_processing:
Methylation property identification: For any particular DNA variant inside library, it carries particular barcode to indicate its methylation property, i.e., unmethylated, top hemimethylated, bottom hemimethylated, duplex methylated. Thus acording to the unique methylation barcode information, each read was identified and sorted to its methylation type.
Variant reads counting and sorting: For each randomized binding site, its number in Bound and Unbound portions were counted separately. The Bound/Unbound ratio was supposedly porportional to its relative binding affinity, and the negative logarithm of this ratio indicates the relative binding energy in kT unit.
Energy logo visualization: Based on the binding energy for particular reference and all its single variants, an energy logo can be generated to indicate the specificity of particular protein-DNA interactions. Futhermore, besides conventional G-T-A-C bases, we used two extra “M" and ”W" to represent methylated cytosine on upper and lower strand of DNA respectively, in which the height of M or W indicate the extra binding energy contribution between methylated and unmethylated base.
Supplementary_files_format_and_content: All processed data were saved in .excel files, including the methylation property, bound reads, unbound reads, Bound/Unbound ratio, relative binding energy for each variant.

organism: synthetic construct

source_name: mouse HOXB13

growth_protocol: No

extract_protocol:
After EMSA, DNA fragments were cut and extracted by gel extraction buffer at 50C for 30mins.
For each portion of DNA fragment, PE1 and PE2 PCR primers were used to PCR amplify extracted DNA by at least 8 cycles (95C 15s, 52C 15s, 72C 30s)
For each particular sample, there are two raw data files, i.e., bound reads and unbound reads with different Illumina Index. The processed data file was based on analysis of Bound and Unbound data together.

treatment_protocol: All EMSA reactions were done in 12% Tris-glycine gel, 200V, 30mins

tag: dna library composition

value: Including unmethylated dsDNA library HOXB13-UN, top hemimethylated HOXB13-tH, bottom hemimethylated HOXB13-bH, and duplexmethylated HOXB14-dM

tag: recombinant protein used

value: mouse HOXB13 DNA binding domain (In vitro expreesed by NEB Purexpress system)


title: UMCG-AB-005 Leucocyte

city: Ghent

first: Jeroen

last: Galle

address: Coupure Links 653

department: Dept. Of Mathematical Modelling, Statistics and Bioinformatics

country: Belgium

institute: Ghent University

description: Processed data file: MBD2 R vs NR promoter -2000 - +500bp exon_Count Data.xlsx

data_processing:
Data were mapped using BOWTIE.
For each nucleotide, the amount of mapped fragments was considered (= coverage).
Data files were compressed by merging neighbouring nucleotides with equal coverages into single rows, with indication of start and end position of nucleotides with that specific coverage.
Genome_build: hg19 (GRCh37)
Supplementary_files_format_and_content: MBD2_R_vs_NR_promoter_-2000_-_+500bp_exon_Count_Data.xlsx: Excel file with columns containing resp. region id, version, Ensembl gene id, Entrez Gene id, gene label, gene description, chromosome, begin position, end position (nucleotide), location, location info, middle difference and amount of mapped fragments perl sample for the promoter region (-2000 bps and +500 bps of TSS).

source_name: Leucocytes, healthy

extract_protocol:
DNA extracted using Qiagen Blood and Tissue Kit according to manufacturer's instructions.
Libraries were prepared using standard multiplexed paired-end ChIP-seq protocol (Illumina). In summary, 500 ng of DNA per sample was fragmented by ultrasound mediated DNA shearing (Covaris S2, Covaris, Woburn, Massachusetts, USA) set to obtain fragments with an average length of 200bp. We employed the MethylCap kit (Diagenode AF-100-0048, Liège, Belgium) to obtain methylated DNA fragments using the highest salt concentration single elution step option. This was followed by the library preparation for the enriched DNA, which was performed with a modified 'multiplexed paired-end ChIP protocol' (Illumina, San Diego, California, USA). Therefore, the NEBNext® DNA Sample Prep Master Mix Set 1 (New England BioLabs (NEB) E6040, Ipswich, Massachusetts, USA) was combined with the Multiplexing Sample Preparation Oligo Kit (Illumina PE-400-1001). Per sample, 22µl of the prepared sample was subjected to PCR (21 cycles) following the Illumina Library Amplification Index Protocol. After purification (QIAquick PCR Purification columns, Qiagen), elution and concentration (evaporation), the libraries were pooled and denatured (NaOH), followed by Illumina HiSeq 2000 sequencing (multiplexed paired-end run, 2 times 45 cycles) of 10pM/pool.

tag: disease

value: Healthy

tag: tissue

value: Leucocytes


title: HOXB13.sample1.bound

description: In vitro protein-DNA binding reactions

value: Including unmethylated dsDNA library HOXB13-UN, top hemimethylated HOXB13-tH, bottom hemimethylated HOXB13-bH, and duplexmethylated HOXB13-dM


title: Control 12

description:
sample12
RNAseq of Total RNA from 100mg of homogenate tissue.
Processed data file: log2RPKMcounts.xlsx
Processed data file: log2RPKMcounts.rda

source_name: Postmortem Brain, DLPFC

value: Healthy control


title: UMCG-AB-045 Leucocyte


title: UMCG-TT-20 Non-responder

source_name: Ovarian tumor, non-responder

value: High-grade serous ovarian cancer (HGSOC)

value: Ovarian tumor

tag: response to platinum-based chemotherapy

value: Non-responder


title: RNA NMP T+ S+

source_name: T+ / Sox2+ FACS sorted cells from caudal ends of WT embryos


title: UMCG-TT-19 Non-responder


title: Control 8

description:
sample8
RNAseq of Total RNA from 100mg of homogenate tissue.
Processed data file: log2RPKMcounts.xlsx
Processed data file: log2RPKMcounts.rda

tag: disease status

value: Postmortem brain, dorsolateral prefrontal cortex (DLPFC)

value: Male

value: CAUC


title: UMCG-TT-18 Responder

source_name: Ovarian tumor, responder

value: Extreme responder