title: female_naive_neurons_8

description:
S224_female
Supplementary Table 1 (cufflinks), Supplementary Table 3 (deseq2), Supplementary Table 6 (QC)


title: female_naive_neurons_7

status: Public on Nov 28 2017

type: SRA

city: London

first: Franziska

last: Denk

address: Guy's Campus

department: Wolfson SPaRC

country: United Kingdom

institute: King's College London

description:
S223_female
Supplementary Table 1 (cufflinks), Supplementary Table 3 (deseq2), Supplementary Table 6 (QC)

library_source: transcriptomic

data_processing:
alignment using STAR to mm10 (standard parameters, except with --outFilterMultimapNmax 1)
merging of bam files across lanes using samtools
cufflinks for expression level determination (including fragment bias, length and multi-read corrections)
cufflinks expression cut-off deemed mathematically and biologically appropriate: FPKM >= 1
Deseq2 for differential expression level analysis (standard parameters)
Genome_build: mm10
Supplementary_files_format_and_content: Supplementary Table 1: cufflinks-derived FPKM values for FACSed naïve DRG neurons with & without expression cut-off
Supplementary_files_format_and_content: Supplementary Table 2: cufflinks-derived FPKM values for MACSed DRG neurons after PSNL with and without expression cut-off
Supplementary_files_format_and_content: Supplementary Table 3: DESeq2 comparing male vs. female DRG neurons
Supplementary_files_format_and_content: Supplementary Table 6: RNA RIN values and alignment stats

molecule: total RNA

organism: Mus musculus

source_name: FACSed DRG neurons

extract_protocol:
RNA extractions were performed in a single, gender-matched batch using a Qiagen RNeasy micokit, following manufacturer’s protocols with some minor modifications. RNA quality was assessed using an Agilent Bioanalyser PicoChiP (for RIN values of sequencing samples see Supplementary Table 6).
RNA was sent for batch-controlled library preparation (SMARTer Ultra Low Input HV kit, 634820, Clontech) and sequencing at the High-Throughput Genomics Group at the Wellcome Trust Centre for Human Genetics at Oxford University. After amplification, samples were multiplexed in replicate flow cells on an Illumina HiSeq4500 platform to yield 75 bp reads of at least 25 M read depth.

tag: cell type

value: DRG neurons

tag: strain

tag: Sex

value: female

tag: injury condition

value: naïve

tag: estrus identification

value: no

tag: contaminant

value: satellite glial cells


title: 5#-0 RNA-seq

city: Shanghai

first: Hailiang

last: Liu

address: NO 1239, siping road

country: China

institute: Tongji university

description: 5#-0 RNA-seq

data_processing:
Illumina Casava1.7 software used for basecalling.
FastQC v0.10.1 used for quality control （default parameter）
Sequenced reads were trimmed  by Trimmomatic v0.32 (parameter: PE -threads 16 -phred33  HEADCROP:9 MINLEN:19)
Reads were mapped to JGI 221 G.raimondii D genome using TopHat v2.0.13(bowtie2 v2.2.4) (default parameter)
Alignment results converted from bam file to sam file and sorted by samtools(0.1.19)
Counting reads in genes using HTSeq( 0.6.0)(parameter: -t CDS -s no)
Calculating the adjusted p-value of genes(padj) of different samples by R package DESeq(1.18.0)
Genome_build: Gossypium raimondii (D5) genome JGI assembly v2.0 (annot v2.1)
Supplementary_files_format_and_content: Excel file contains DESeq output where all cotton color genotypes have been combined for a comparison between day 0 and day 12.

organism: Gossypium hirsutum

source_name: Transgenic Brown cotton fiber 0 day

extract_protocol:
Total RNA was extracted according to a guanidine thiocyanate method
The first step in the workflow involves purifying the poly-A containing mRNA molecules using poly-T oligo-attached magnetic beads. Following purification, the mRNA is fragmented into small pieces using divalent cations under elevated temperature. Then the cleaved RNA fragments are copied into first strand cDNA using reverse transcriptase and random primers. This is followed by second strand cDNA synthesis using DNA Polymerase I and RNaseH. These cDNA fragments then go through an end repair process, the addition of a single ‘A’ base, and then ligation of the adapters. These products are then purified and enriched with PCR to create the final cDNA library.  The cDNA library of each sample was constructed according to the instructions given in “Preparing Samples for Sequencing of mRNA Test Kits” (Illumina). Products were checked to verify the cDNA library quality and fragment length using an Agilent 2100 DNA 1000 Kit. Sequencing of the resulting cDNA library was carried out on an Illumina HiSeq 2000 paired-end 100-bp (PE 100) system.

tag: day post-anthesis

value: fiber development 0 day

tag: tissue

value: cotton fiber


title: 5#-12 RNA-seq

description: 5#-12 RNA-seq

source_name: Transgenic Brown cotton fiber 12 day

value: fiber development 12 day


title: female_naive_neurons_6

description:
S222_female
Supplementary Table 1 (cufflinks), Supplementary Table 3 (deseq2), Supplementary Table 6 (QC)


title: WCF-12 RNA-seq

description: WCF-12 RNA-seq

source_name: White cotton fiber 12 day


title: WCF-0 RNA-seq

description: WCF-0 RNA-seq

source_name: White cotton fiber 0 day


title: female_naive_neurons_4

description:
S216_female
Supplementary Table 1 (cufflinks), Supplementary Table 3 (deseq2), Supplementary Table 6 (QC)


title: female_naive_neurons_3

description:
S215_female
Supplementary Table 1 (cufflinks), Supplementary Table 3 (deseq2), Supplementary Table 6 (QC)


title: GCF-0 RNA-seq

description: GCF-0 RNA-seq

source_name: Green cotton fiber 0 day


title: female_naive_neurons_2

description:
S214_female
Supplementary Table 1 (cufflinks), Supplementary Table 3 (deseq2), Supplementary Table 6 (QC)


title: GCF-12 RNA-seq

description: GCF-12 RNA-seq

source_name: Green cotton fiber 12 day


title: female_naive_neurons_11

description:
S230_female
Supplementary Table 1 (cufflinks), Supplementary Table 3 (deseq2), Supplementary Table 6 (QC)


title: BCF-0 RNA-seq

description: BCF-0 RNA-seq

source_name: Brown cotton fiber 0 day


title: male_naive_neurons_10

description:
S226_male
Supplementary Table 1 (cufflinks), Supplementary Table 3 (deseq2), Supplementary Table 6 (QC)

value: male


title: 24#-0 RNA-seq

description: 24#-0 RNA-seq

source_name: Transgenic White cotton fiber 0 day


title: male_naive_neurons_11

description:
S231_male
Supplementary Table 1 (cufflinks), Supplementary Table 3 (deseq2), Supplementary Table 6 (QC)


title: BCF-12 RNA-seq

description: BCF-12 RNA-seq

source_name: Brown cotton fiber 12 day


title: 6#-0 RNA-seq

description: 6#-0 RNA-seq

source_name: Transgenic Green cotton fiber 0 day


title: male_naive_neurons_9

description:
S225_male
Supplementary Table 1 (cufflinks), Supplementary Table 3 (deseq2), Supplementary Table 6 (QC)


title: male_naive_neurons_8

description:
S220_male
Supplementary Table 1 (cufflinks), Supplementary Table 3 (deseq2), Supplementary Table 6 (QC)


title: RT25PCR7_1004J1ES_p8

city: Wako

first: Itoshi

last: NIKAIDO

state: Saitama

address: 2-1 Hirosawa

department: Center for Biosystems Dynamics Research

country: Japan

institute: RIKEN

description: We analyzed single-cells, which were sorted to 384-wells in total.

data_processing:
BCL files generated by Illumina NextSeq500 were converted to fastq files by bcl2fastq2 (v2.17.1.14) with demultiplexing pool barcodes.
Data for Quartz-Seq2 was processed by Quartz-Seq2 pipeline. Data for Drop-seq was processed by Drop-seq pipeline.
Genome_build: mm10
Supplementary_files_format_and_content: tab-delimited text files include UMI counts for each gene for each cell

source_name: J1 ES cells (2i/LIF)

extract_protocol:
We performed two single-cell RNA-seq methods (Drop-seq and Quartz-Seq2). Drop-seq was performed as reported previously [Cell. 2015;161:1202–14]. Moreover, we established Quartz-Seq2 (a high-throughput single-cell RNA-sequencing method that effectively uses limited sequence reads). We used two sets of RT primer for Quartz-Seq2. The set of v3.1 RT primers has 384 kinds of unique cell barcodes, with a length of 14 nucleotides (OPC purification, FASMAC). The set of v3.2 RT primers has 1,536 kinds of unique cell barcodes, with a length of 15 nucleotides (OPC purification, Sigma). The set of v3.1 RT primers corresponds to one set of the 384-well PCR plate with lysis buffer, the wells of which have unique barcodes. The set of v3.2 RT primers corresponds to four sets of the 384-well PCR plate with lysis buffer. Single cells were isolated in the 384-well PCR plate with 1 µL of lysis buffer (0.1111 µM respective RT primers, 0.12 mM dNTP mix, 0.3% NP-40, 1 unit/µL RNasin plus) containing ERCC spike mix I.
For Quartz-Seq2, cryopreserved 384-well plates with single-cell lysate were centrifuged at 10,000 g and 4ºC for 1 min. Subsequently, we denatured total RNA in each 384 plate at 70ºC for 90 s and hybridized the RT primer to poly-adenylated RNA at 35ºC for 15 s using the C1000/S1000 thermal cycler. The resulting plates were again centrifuged at 10,000 g and 4ºC for 1 min. Next, the plates were placed on the 384 aluminum plate at 0ºC. We peeled away the seal and added 1 µL of RT premix (2x Thermopol buffer, 5 units/µL SuperScript III, 0.55 units/µL RNasin plus) to 1 µL of lysis buffer for each well using a Mantis microfluidic dispensing system (Formulatrix) or a 384 Transfer Plate system (1859-384S, Watson). The above RT solution was used for the RT25 condition. For the RT100 condition, we used the following RT solution: 2x Thermopol buffer, 20 units/µL SuperScript III, and 2.2 units/µL RNasin plus. We sealed the plates again and agitated them at 2,600 rpm and 4ºC for 1 min. The plates were then centrifuged at 10,000 g and 4ºC for 1 min. We then performed reverse transcription at 35ºC for 5 min and 50ºC for 50 min. The reverse transcription was stopped at 70ºC for 15 min. Then, the plates were placed on a prechilled aluminum block, after which we peeled off their seals. Subsequently, we turned the plates upside down on the assembled collector type A or type B (supplemental figure). We mainly used type A. We centrifuged the plates with an assemble collector at 3,010 g and 4ºC for 3 min with swing-bucket rotors. Subsequently, we collected the cDNA solution into a disposable reservoir. Typically, we obtained 650–700 µL of cDNA solution from one 384-well PCR plate. We purified and concentrated the cDNA solution using the DNA Clean & Concentrator™-5 kit (Zymo Research). We used three purification columns for one 384-well PCR plate in the case of the v3.1 RT primer system (384-cell barcode). Purified cDNA was extracted into 20 µL of nuclease-free water from one column purification and transferred into an eight-linked PCR tube (TaKaRa). The PCR tubes were placed on an aluminum PCR stand at 0ºC. We added 25 µL of TdT solution [1x Thermopol buffer, 2.4 mM dATP, 0.0384 units/µL RNase H (Invitrogen), 26.88 units/µL terminal transferase (Roche)] into 20 µL of extracted cDNA using a pipette at 0ºC. The resulting 45 µL of TdT solution was mixed with a pipette at 0ºC or ThermoMixer at 2,000 g and 0ºC for 1 min. Immediately thereafter, the PCR tubes were centrifuged at 10,000 g and 0ºC for 1 min. We used a C1000/S1000 thermal cycler equipped with the 96-Deep Well Reaction Module for the following steps. The PCR tubes were placed on the block of the thermal cycler, which had been prechilled to 0ºC. We then performed a poly-A tailing reaction at 37ºC for 75 s. The solution was inactivated at 65ºC for 10 min. The PCR tubes were placed on an aluminum PCR stand at 0ºC. We then dispensed approximately 11 µL of solution into four wells from 45 µL of TdT solution. We added 46.16 µL of PCR I premix (1.08492x MightyAmp Buffer version 2, 0.06932 µM Tagging primer, 0.05415 units/µL MightyAmp DNA polymerase) to 11 µL of TdT solution for the respective wells of the PCR tube. We performed gentle inversion mixing on the resulting solution in the PCR tube. The tubes were then centrifuged at 10,000 g and 4ºC for 1 min. Subsequently, the solution was mixed with ThermoMixer at 2,000 rpm and 4ºC for 2 min. Then, we spun down the tube again. Next, we denatured the solution at 98ºC for 130 s and hybridized Tagging primer to poly-A-tailed cDNA at 40ºC for 1 min. After that, we performed the “Increment step” by heating to 68ºC at 0.2ºC every second and performed second-strand synthesis at 68ºC for 5 min. The tubes were placed on an aluminum PCR stand at 0ºC. We added 50.232 µL of PCR II premix (0.99697x MightyAmp Buffer version.2, 1.8952 µM gM primer) to 56.16 µL of PCR I solution. We performed gentle inversion mixing on the resulting solution in the PCR tube. The tubes were then centrifuged at 10,000 g and 4ºC for 1 min. Subsequently, the solution was mixed with ThermoMixer at 2,000 rpm and 4ºC for 2 min, after which we spun down the tube again. We then placed it on the block of the thermal cycler at 68ºC. Subsequently, we amplified the cDNA for 11 cycles under the following conditions: 98ºC for 10 s, 65ºC for 15 s, and 68ºC 5 min. We then incubated the tube at 68ºC for an additional 5 min. Finally, we transferred all of the PCR solution, derived from one 384-well PCR plate, to a 50-mL Polypropylene Centrifuge Tube (Watson). Typically, we obtained approximately 1.2 mL of PCR solution per 384-well PCR plate. We added 32 µL of 3 M sodium acetate (pH 5.2) and 6420 µL PB-Buffer (Qiagen) to the PCR solution. The mixture was then purified using a MinElute Spin Column (Qiagen). Purified cDNA was extracted into 40 µL of nuclease-free water. We additionally purified the cDNA with 32 µL of Ampure XP beads. Finally, we obtained 32 µL of purified cDNA. We checked the length distribution of amplified cDNA with an Agilent High Sensitivity DNA Kit (Agilent). The typical average size of the amplified cDNA in Quartz-Seq2 was approximately 1,400 bp.   In the case of the usage of the v3.2 RT primer in Quartz-Seq2, we modified the above steps as follows. After reverse transcription, we collected cDNA solution into a disposable reservoir from four sets of 384-well plates, which corresponded to 1,536 wells. We purified and concentrated the cDNA solution using eight purification columns for four 384-well PCR plates in the case of the v3.2 RT primer system. In the PCR step, we amplified cDNA for nine cycles with the following conditions: 98ºC for 10 s, 65ºC for 15 s, and 68ºC for 5 min. Finally, we transferred all of the PCR solution derived from four 384-well plates to a 50-mL Polypropylene Centrifuge Tube (Watson). Typically, we obtained approximately 3.5 mL of PCR solution per four 384-well PCR plates. We added 88 µL of 3 M sodium acetate (pH 5.2) and 17.6 mL of PB-Buffer (Qiagen) to the PCR solution. The mixture was purified using the MinElute Spin Column (Qiagen). Subsequently, cDNA was again purified using Ampure XP magnetic beads.   For the Quartz-Seq1-like reaction, we performed the following procedure in accordance with our previous study. We added 1 µL of RT premix (2x PCR buffer, 5 units/µL SuperScript III, 0.55 units/µL RNasin plus) to 1 µL of lysis buffer. We then performed reverse transcription at 35ºC for 5 min and 45ºC for 20 min. This reverse transcription was stopped at 70ºC for 15 min. We added 5 µL of ExoIB solution (1.6x Exonuclease I buffer, 3.2x PCR buffer, 16 mM DTT) and 20 µL of TdT solution [1x PCR buffer, 3 mM dATP, 0.0384 units/µL RNase H (Invitrogen), 33.6 units/µL terminal transferase (Roche)] into 20 µL of extracted cDNA using a pipette at 0ºC. The PCR tubes were placed on the block of the thermal cycle

value: J1 ES single-cells

tag: cell line

tag: the number of single-cell/well contained in each data

tag: method name of single-cell rna-seq

value: Quartz-Seq2 (v3.1, RT25)

tag: type of sequence library preparation

value: Ligation

tag: sequencer

tag: the number of fastq reads for dge-matrix (in million)

tag: read2 length for dge-matrix (nt)


title: male_naive_neurons_7

description:
S219_male
Supplementary Table 1 (cufflinks), Supplementary Table 3 (deseq2), Supplementary Table 6 (QC)


title: female_naive_neurons_1

description:
S213_female
Supplementary Table 1 (cufflinks), Supplementary Table 3 (deseq2), Supplementary Table 6 (QC)


title: female_naive_neurons_5

description:
S221_female
Supplementary Table 1 (cufflinks), Supplementary Table 3 (deseq2), Supplementary Table 6 (QC)


title: XMA.z_M20mo_S_0pt

city: San Marcos

first: Yuan

last: Lu

state: TX

address: university drive

country: USA

institute: Xiphophorus Genetic Stock Center

data_processing: No processed data was generated.

organism: Xiphophorus maculatus

source_name: Xiphophorus Genetic Stock Center

extract_protocol:
X. maculatus at varies at varies age were sacrificed for RNA isolation. Total RNA was isolated using TriReagent (Sigma Inc., St. Louis, MO, USA). Tissues from each individual fish was homogenized in Tri-reagent followed by addition of 200 μl chloroform and the samples vigorously shaken and subjected to centrifugation at 12,000 g for 15min at 4 °c. Total RNA was further purified using RNeasy mini RNA isolation kit (Qiagen, Valencia, CA, USA). Residual DNA was eliminated by performing on column DNase digestion at 37°C for 30 min. RNA integrity was assessed by gel electrophoresis (2% agarose in TAE running buffer) and total RNA concentration was determined using a spectrophotometer (NanoDrop Technologies, Wilmington, DE, USA). RNA quality was verified on an Agilent Bioanalyzer (Agilent Technologies, Santa Clara, CA) to confirm that RIN scores were above 8 prior to sequencing.
RNA libraries were prepared for sequencing using standard Illumina protocols. Libraries were constructed using the Illumina TrueSeq library preparation system that employs a polyA selection

value: skin

tag: age

value: 20 months

tag: gender


title: XMA.z_M20mo_S_Sham


title: male_naive_neurons_6

description:
S218_male
Supplementary Table 1 (cufflinks), Supplementary Table 3 (deseq2), Supplementary Table 6 (QC)

tag: cell type


title: XMA.z-r_M20mo_L_0min

tag: strain

value: liver


title: RT25PCR11_1004J1ES_p3

tag: Sex


title: RT25PCR7_1004J1ES_p7


title: RT25PCR11_1004J1ES_p1


title: male_naive_neurons_5

description:
S217_male
Supplementary Table 1 (cufflinks), Supplementary Table 3 (deseq2), Supplementary Table 6 (QC)

tag: injury condition


title: E10.5_SETD8-null_B

city: Rochester

first: Laurie

middle: A

last: Steiner

state: NY

address: 601 Elmwood Avenue

department: Center for Pediatric Biomedical Research

institute: University of Rochester

data_processing:
Quality evaluation with FastQC
Quality trim/filter with Trimmomatic (v0.32)
Alignment with Tophat(v.2.1.0)
Read quantification with HTSeq-count (v0.6.1)
EdgeR for differential expression analysis
Genome_build: mm9
Supplementary_files_format_and_content: compiled read counts for each sample generated from HTSeq-count

molecule: polyA RNA

source_name: Fetal liver

growth_protocol: Embryos were harvested following timed pregnancies at either E9.5 or E10.5 (E= Embryonic Day)

extract_protocol:
Life Technologies Arcturus PicoPure RNA isolation kit
RNA libraries were prepared for sequencing using standard Illumina protocols

treatment_protocol: cells were stained with Ter119 and CD71 antibodies and live Ter119+CD71+ cells were collected via FACS

tag: developmental stage

value: erythroblasts

tag: genotype

value: SETD8fl/fl EpoR Cre


title: XMA.z_M20mo_L_0min


title: male_naive_neurons_3

description:
S211_male
Supplementary Table 1 (cufflinks), Supplementary Table 3 (deseq2), Supplementary Table 6 (QC)

value: DRG neurons

tag: estrus identification

tag: contaminant


title: E10.5_SETD8-null_A


title: male_naive_neurons_4

description:
S212_male
Supplementary Table 1 (cufflinks), Supplementary Table 3 (deseq2), Supplementary Table 6 (QC)


title: XMA.z_M20mo_Int_0pt

tag: tissue

value: intestine


title: XMA.z_M20mo_Int_Sham


title: male_naive_neurons_2

description:
S210_male
Supplementary Table 1 (cufflinks), Supplementary Table 3 (deseq2), Supplementary Table 6 (QC)

value: naïve


title: E10.5_control-Cre_D

value: Setd8 +/+ EpoRCre


title: XMA.x_M20mo_S_Sham


title: RT25PCR11_1004J1ES_p6

tag: cell line

tag: the number of single-cell/well contained in each data


title: male_naive_neurons_1

description:
S209_male
Supplementary Table 1 (cufflinks), Supplementary Table 3 (deseq2), Supplementary Table 6 (QC)

value: no

value: satellite glial cells


title: XMA.x_M20mo_S_0pt


title: XMA.x_M20mo_L_0min


title: E10.5_control-Flox

value: Setd8 fl/fl EpoR WT


title: E10.5_SETD8-null


title: JWCB005Δcra - NaCl - 2

city: Oak Ridge

first: Kyle

last: Sander

state: TN

address: 1 Bethel Valley Road, MS 6038

department: Biosciences

institute: University of Tennessee/Oak Ridge National Laboratory

data_processing:
Fastq files were downloaded from HudsonAlpha’s servers and verified for integrity by computing their checksum.  Read quality was checked using FastQC (v. 0.11.5) [279].  RNAseq reads were trimmed the using Trimmomatic (v. 0.33) MAXINFO method with target length and strictness parameters set to 40 and 0.8, respectively [280]. Reads were mapped to the genome of Caldicellulosiruptor bescii JWCB005 (GCF_900166995.1, last modified 2017/03/09) using Bowtie2 (v. 2.2.9) with the same parameters as the very-sensitive preset option expect the number of mismatches was set to 1 [281], and reads were counted with HTSeq (v. 0.6.1p1) [282].
Raw read counts for each coding sequence were used as input for differential expression analysis using the DEseq2 package as part of the Bioconductor Suite in R
Genome_build: NZ_FUZN01000002
Supplementary_files_format_and_content: binary_comparisons_glu_gly_nacl_fapR_cra.xlsx, .xlsx format - Microsoft Excel spreadsheet version 2016.  Spreadsheet contains six tabs.  Each tab contains differential expression output from the DEseq2 package comparing any of the six [deletion strain-osmolyte] combinations with that of the genetic parent strain JWCB005 grown in the same osmolyte condition (e.g. JWCB005ΔB5X54_RS01260 vs. JWCB005, both strains grown in elevated glycerol).  The strain comparisons and osmolyte conditions are indicated on the top of each worksheet.

organism: Caldicellulosiruptor bescii

source_name: Caldicellulosiruptor bescii strain JWCB005 with the cra gene deleted cell biomass

growth_protocol: Cells were cultured in 135 mL serum bottles in a working/culture volume of 50 mL at a growth temperature of 75oC.  Media was prepared and allowed to deoxygenate overnight in an anaerobic chamber containing 5% H2, 10% CO2 and 85% N2.  Appropriate amounts of various osmolytes were added to media to give each media its total calcuated initial osmolarity.  Bottles were inoculated to equivalent OD680’s of 0.001-0.01 using batch grown cultures grown to mid-log phase.  Cultures were incubated in a shaking incubator oscillating at 200 rpm.  Three cultures were maintained and sampled for optical density measurement to track growth.  Six parallel bottles were maintained and harvested when they achieved mid-log (OD680 = ~0.170-0.190).  The three harvested replicates with OD680 closest to 0.180 were processed for RNAseq analysis.

extract_protocol:
Total RNA was extracted by first incubating cell pellets in 250 µL of 20 mg/mL Lysozyme (Sigma Aldrich part number L-7651, St. Louis, MO) resuspended in SET buffer (50 mM Tris-HCl pH 8.0 50 mM EDTA, 20% w/v Sucrose) and incubated in a dry stationary bath at 37oC for eight minutes, vortexing briefly every two minutes.  900 µL of buffer RLT from a Qiagen RNeasy Kit (Qiagen, Hilden, Germany) was added and the sample was split into two equal aliquots of ~575 µL each.  325 µL of 100% ethanol was added to each aliquot and the contents were mixed by pipetting.  Contents from both tubes were spun through an RNeasy spin column (Qiagen, Hilden, Germany).  Spin columns were washed once with buffer RW1 (Qiagen, Hilden, Germany).  10 µL of DNAse stock I (Qiagen, Hilden, Germany) and 70 uL of buffer RDD (Qiagen, Hilden, Germany) were combined, pipetted onto the spin column and allowed to incubate for 15 minutes at room temperature to digest residual DNA.  The columns were washed once more with buffer RW1 (Qiagen, Hilden, Germany), followed by two washes of 500 µL buffer RDE (Qiagen, Hilden, Germany) and eluted in 35 µL RNAse free H20 (Qiagen, Hilden, Germany).  RNA concentration was quantified using a Nanodrop 1000 instrument (ThermoScientific, Waltham, MA) and RNA quality was verified by obtaining RNA Integrity Numbers (RIN) using an Agilent 2100 Bioanalyzer and corresponding RNAchip (Agilent Technologies, Santa Clara, CA).
Depleted RNA was used as the starting material for the Epicentre ScriptSeq™ v2 RNA-Seq Library preparation kit (Illumina compatible) utilising Epicentres Fail Safe PCR Enzyme mix (Epicentre, WI) and following the manufacturer’s protocol. cDNA, tagged with ScriptSeq adaptors (1-12), was purified with Agencourt AMPure XP magnetic beads (Beckman Coulter, IN ) and  eluted in water according to the ScriptSeq™ v2 RNA -Seq Library preparation kit protocol.  Twelve PCR cycles were used during library amplification and samples were purified with Agencourt AmPureXP magnetic beads and eluted with 20 µl of water. The final mRNA Seq library was quantified with a Qubit Fluorometer (Invitrogen, CA) and library quality was assessed with an Agilent Bioanalyzer DNA Chip (Agilent, CA). Samples were pooled and the concentration determined.  Pools of barcoded samples were sent to Hudson Alpha for SR50 sequencing run on an Illumina Hiseq 2500 platform.

treatment_protocol: Each single gene deletion strain was incubated separately in three elevated osmolarity conditions; added sodium chloride to a total caluclated initial osmolarity of 300 mOsm/L, added glucose to a total caluclated initial osmolarity of 500 mOsm/L, and added glucose to a total starting calculated osmolarity of 300 mOsm/L.

tag: osmolyte added to media

value: Sodium Chloride

value: delta-pyrF delta-cra

tag: total media starting osmolarity

value: 300 mOsm/L

tag: primary osmolyte

value: sodium chloride


title: XMA.x_M20mo_Int_0pt

value: male


title: JWCB005Δcra - NaCl - 3

value: Caldicellulosiruptor bescii strain JWCB005 with the cra gene deleted


title: XMA.n_M20mo_S_Sham

value: skin

value: 20 months


title: JWCB005Δcra - NaCl - 1


title: XMA.n_M20mo_L_Sham

value: liver


title: XMA.n_M20mo_Int_Sham


title: XMA_MS_istg25-embryo_WF

description: XMA_MS_istg25-embryo_WF_none_UO_2010-05-02

value: whole fish

value: embryo(stage 15)

value: missing


title: JWCB005Δcra - Glycerol - 2

value: Glycerol

value: 500 mOsm/L

value: glycerol


title: JWCB005Δcra - Glycerol - 1


title: JWCB005Δcra - Glucose - 2

value: Glucose

value: glucose


title: JWCB005Δcra - Glucose - 3


title: E10.5_control-Cre

value: erythroblasts

value: Setd8 +/+ EpoRCre


title: XMA.ac_M24mo_S_5AM

value: 24 months


title: XMA.a_F20mo_B_2PM

value: brain

value: female


title: XMA.ab_M24mo_S_5AM

value: male


title: JWCB005ΔfapR - NaCl - 3

source_name: Caldicellulosiruptor bescii strain JWCB005 with the fapR gene deleted cell biomass

value: Sodium Chloride

value: delta-pyrF delta-fapR

value: 300 mOsm/L

value: sodium chloride


title: XMA.ac_M24mo_B_5AM


title: JWCB005Δcra - Glucose - 1

value: Caldicellulosiruptor bescii strain JWCB005 with the cra gene deleted

value: delta-pyrF delta-cra


title: JWCB005ΔfapR - NaCl - 2

value: Caldicellulosiruptor bescii strain JWCB005 with the fapR gene deleted


title: JWCB005ΔfapR - NaCl - 1


title: JWCB005Δcra - Glycerol - 3

value: Glycerol

value: 500 mOsm/L


title: JWCB005ΔfapR - Glycerol - 3

value: glycerol


title: XMA.ab_M24mo_B_5AM


title: JWCB005ΔfapR - Glycerol - 2


title: JWCB005ΔfapR - Glycerol - 1


title: 24 hr Rep 3

city: Dallas

first: Pamela

middle: C

last: Ce La Cruz

address: 6000 Harry Hines Blvd

department: Dept. Microbiology

institute: UT Southwestern Medical Center

description:
Cells treated with universal type I IFN ( 50 units/mL) and RNA harvested at 24 hr.
24_hrs_C

data_processing:
Paired-end 100 bp read length Fastq files were checked for quality using fastqc/v0.11.2 and fastq_screen/v0.4.4
Reads were quality trimmed using fastq-mcf (ea-utils/v1.1.2-806).
Trimmed fastq files were mapped to Pteropus alecto (black flying fox) assembly ASM32557v1 (ftp://ftp.ncbi.nih.gov/genomes/Pteropus_alecto) using TopHat/v2.0.12
Duplicates were marked using picard-tools/v1.127
Reference annotation based transcript assembly was done using cufflinks/v2.2.1, and read counts were generated using featureCounts (subread/v1.4.6).
Normalized read counts generated using edgeR/v3.2.4
Genome_build: ASM32557v1
Supplementary_files_format_and_content: tab delimited txt files that includes raw read counts for each sample per gene

organism: Pteropus alecto

source_name: PaKi

growth_protocol: PaKi cells were grown in DMEM/F-12 media (GIBCO) supplemented with 10% FBS.

extract_protocol:
RNA was harvested using RNeasy Mini Kit (Qiagen #74106) following the manufacturer's instructions. Integrity was verified using the Agilent 2100 Bioanalyzer. All samples had a RIN score >9.5.
One microgram of total DNAse-treated RNA was prepared with the TruSeq Stranded Total RNA LT Sample Prep Kit (Illumina  #RS-122). rRNA-depleted total RNA was fragmented and used for strand-specific cDNA synthesis.  cDNA was then A-tailed and ligated with indexed adapters. Samples were then PCR amplified, purified with AmpureXP beads, and validated on the Agilent 2100 Bioanalyzer and Qubit.  Normalized and pooled samples were run on the Illumina Hiseq 2500 using SBS v3 reagents.

treatment_protocol: Cells were seeded at a density of 3x10^5 cells/well of a six-well plate.The following day, cells were treated with 50U/mL universal type I IFN (PBL# #11200) added to the media. Cells were harvested for RNA extraction at several time points.

value: Kidney

value: WT

tag: time point


title: XMA_MS_istg15-embryo_WF

description: XMA_MS_istg15-embryo_WF_none_UO_2010-05-02

value: whole fish

tag: age

value: embryo(stage 15)

value: missing


title: Rhesus_Placenta1

city: Bethesda

first: Peng

last: Yang

state: Maryland

address: 6 Center Drive 6B-211

department: DDB

institute: NICHD/NIH

data_processing:
Illumina Casava software used for basecalling.
Fastq files were aligned by Bowtie2(v2.25) with default premeters to generate bam files.
Bam files were converted to tdf files by using IGVtools (2.3.78)
Genome_build: rheMac3
Supplementary_files_format_and_content: tdf files generated from IGVtools

organism: Macaca mulatta

source_name: Rhesus monkey placenta

extract_protocol:
RNA was prepared from Rhesus monkey placenta using the RNeasy Plus Mini kit  (Qiagen).
RNA-seq libraries were constructed according to Illumima protocol and sequenced with the Illumina Hiseq 2000.

value: Placenta


title: 24 hr Rep 1

description:
Cells treated with universal type I IFN ( 50 units/mL) and RNA harvested at 24 hr.
24_hrs_A


title: Rhesus_Placenta2


title: 24 hr Rep 2

description:
Cells treated with universal type I IFN ( 50 units/mL) and RNA harvested at 24 hr.
24_hrs_B


title: 12 hr Rep 3

description:
Cells treated with universal type I IFN ( 50 units/mL) and RNA harvested at 12 hr.
12_hrs_C


title: JWCB005ΔfapR - Glucose - 3

value: Glucose

value: glucose


title: JWCB005ΔfapR - Glucose - 2

value: delta-pyrF delta-fapR


title: XMA.a_F20mo_S_2PM

value: skin

value: 20 months

tag: gender

value: female


title: XMA_106R_u_H_none

value: heart


title: XMA_106R_u_B_none

value: brain


title: 12 hr Rep 1

description:
Cells treated with universal type I IFN ( 50 units/mL) and RNA harvested at 12 hr.
12_hrs_A


title: 8 hr Rep 3

description:
Cells treated with universal type I IFN ( 50 units/mL) and RNA harvested at 8 hr.
8_hrs_C


title: JWCB005ΔfapR - Glucose - 1

value: Caldicellulosiruptor bescii strain JWCB005 with the fapR gene deleted

value: 300 mOsm/L


title: w12

city: beijing

first: wang

last: ying

address: beichen

institute: State Key Laboratory of Mycology

data_processing:
Illumina Casava software used for basecalling.
RNA-Seq were aligned to the reference genome using hisat2 version 2.0.5
fpkm was calculated by cufflink version 2.2.1
differential gene expression was analysed by NOISeq version 2.22.0
Genome_build: Acremonium chrysogenum ATCC 11550 (ascomycetes)
Supplementary_files_format_and_content: gene fpkm

organism: Hapsidospora chrysogena

source_name: AcmybAOE 96 hour

extract_protocol:
flash frozen on liquid nitrogen, and RNA was harvested using Trizol reagent.
RNA libraries were prepared for sequencing using standard Illumina protocols

value: mycelium

tag: age (hours)

value: AcmybA overexpress


title: 12 hr Rep 2

description:
Cells treated with universal type I IFN ( 50 units/mL) and RNA harvested at 12 hr.
12_hrs_B

value: Kidney

value: WT


title: JWCB005 - NaCl - 2

source_name: Caldicellulosiruptor bescii strain JWCB005 cell biomass

tag: strain

value: Sodium Chloride

value: delta-pyrF

value: sodium chloride


title: JWCB005 - NaCl - 3

value: Caldicellulosiruptor bescii strain JWCB005


title: w11

source_name: AcmybAOE 72 hour


title: XMA_111e.b_i_1.5mo_WF

value: whole fish

value: 1.5 months

value: missing


title: 8 hr Rep 2

description:
Cells treated with universal type I IFN ( 50 units/mL) and RNA harvested at 8 hr.
8_hrs_B


title: JWCB005 - NaCl - 1


title: XMA_111e.a_i_1.5mo_WF


title: XMA_106R_u_L_none

value: liver


title: 8 hr Rep1

description:
Cells treated with universal type I IFN ( 50 units/mL) and RNA harvested at 8 hr.
8_hrs_A


title: w10

source_name: AcmybAOE 48 hour


title: w8

source_name: ∆AcmybA 96 hour

value: AcmybA knockout


title: JWCB005 - Glycerol - 3

value: Glycerol

value: 500 mOsm/L

value: glycerol


title: 4 hr Rep 2

description:
Cells treated with universal type I IFN ( 50 units/mL) and RNA harvested at 4 hr.
4_hrs_B


title: w7

source_name: ∆AcmybA 72 hour


title: JWCB005 - Glycerol - 1


title: JWCB005 - Glycerol - 2


title: 4 hr Rep 3

description:
Cells treated with universal type I IFN ( 50 units/mL) and RNA harvested at 4 hr.
4_hrs_C


title: 4 hr Rep1

description:
Cells treated with universal type I IFN ( 50 units/mL) and RNA harvested at 4 hr.
4_hrs_A


title: w4

source_name: WT 96 hour

value: wild type


title: 0 hr Rep 3

description:
Cells treated with universal type I IFN ( 50 units/mL) and RNA harvested immediately.
0_hrs_C


title: 0 hr Rep 2

description:
Cells treated with universal type I IFN ( 50 units/mL) and RNA harvested immediately.
0_hrs_B


title: w3

source_name: WT 72 hour

value: mycelium


title: JWCB005 - Glucose - 3

value: Glucose

value: 300 mOsm/L

value: glucose


title: JWCB005 - Glucose - 2


title: 3ETSleuZ

city: SHERBROOKE

first: Eric

last: Massé

state: Québec

address: 3201, Jean Mignault

department: Biochemistry

country: Canada

institute: University of Sherbrooke

data_processing:
Basecalls performed using MCS Version 2.4.1.3
Reads were aligned to the Escherichia coli K12 genome assembly using Bowtie for Illumina (Galaxy Project)
Read counts were normalized by coverage according to Oshlack et al., 2010
Genome_build: Escherichia coli K12
Supplementary_files_format_and_content: tab-delimited text file include normalized reads for each sample and MS2-3'ETSleuZ/3'ETSleuZ ratio

organism: Escherichia coli K-12

source_name: Bacterium

growth_protocol: Cells were harvested in exponential (OD600nm=0.5; 100 mL LB medium) and in stationary phase (OD600nm=1.3; 100 mL LB medium).

extract_protocol:
Cells were resuspended in 3 mL of buffer A and lysed using a French Press (430 psi, four times). Lysate was then cleared by centrifugation. The soluble fraction was subjected to affinity chromatography. The column was prepared by adding 100 µL of amylose resin (New England Biolabs) to Bio-Spin disposable chromatography columns (Bio-Rad). The column was washed with 3 mL of buffer A. Next, 200 pmol of MS2-MPB protein was immobilized on the amylose resin, and the column was washed with 2 mL of buffer A. The cleared lysate was then loaded onto the column, which was washed with 8 mL of buffer A. RNA was eluted from the column with 1 mL of buffer A containing 15 mM maltose. Eluted RNA was extracted with phenol-chloroform, followed by ethanol (3 vol) precipitation of the aqueous phase in the presence of 20 mg of glycogen. RNA samples were treated with TURBO DNA-free (DNase treatment) and precipitated again.
cDNA librairies were prepared with NEBNext Small RNA Library Prep set E7330S kit to target small RNA fragments.

treatment_protocol: 0.1% arabinose was added to induce the expression of MS2-3'ETSleuZ/3'ETSleuZ during 10 min. Cells were chilled for 10 min on ice. Cells were then centrifuged, resuspended in 1 mL of buffer A (20 mMTris-HCl at pH 8.0, 150 mM KCl, 1 mM MgCl2, 1 mM DTT), and centrifuged again.

tag: growth phase

value: exponential and stationary phases

tag: genotype/variation


title: w2

source_name: WT 48 hour


title: MS2-3ETSleuZ


title: 0 hr Rep 1

description:
Cells treated with universal type I IFN ( 50 units/mL) and RNA harvested immediately.
0_hrs_A

value: Kidney

value: WT


title: Sample_54973

status: Public on Nov 27 2017

city: Ann Arbor

first: Richard

last: McEachin

state: United States

address: 2800 Plymouth Road

department: DCM&B

institute: University of Michigan

data_processing:
QC: We checked the quality of the raw reads data for each sample using FastQC [1] (version 0.10.1) to identify features of the data that may indicate quality problems (e.g. low quality scores, over-represented sequences, inappropriate GC content, etc.).
Aligment: We used the software package Tuxedo Suite for alignment, differential expression analysis, and post-analysis diagnostics [2, 3, 4]. Briefly, we aligned reads to the reference transcriptome including both mRNAs and lncRNSs (UCSC mm10) [5] using TopHat (version 2.0.9) and Bowtie (version 2.1.0.). We used default parameter settings for alignment, with the exception of: “--b2-very-sensitive” telling the software to spend extra time searching for valid alignments.
QC: We used FastQC for a second round of quality control (post-alignment), to ensure that only high quality data would be input to expression quantitation and differential expression analysis.
Normalization and Quantitation: We used Cufflinks/CuffDiff (version 2.1.1) for expression quantitation, normalization, and differential expression analysis, using UCSC mm10.fa as the reference genome sequence. For this analysis, we used parameter settings: “--multi-read-correct” to adjust expression calculations for reads that map in more than one locus, as well as “--compatible-hits-norm” and “--upper-quartile –norm” for normalization of expression values. We generated diagnostic plots using the CummeRbund package.
Differential Expression: We used locally developed scripts to format and annotate the differential expression data output from CuffDiff. Briefly, we identified genes and transcripts as being differentially expressed based on three criteria: test status = “OK”, FDR < 0.05 and fold change ≥ ± 1.5.
Annotation: We annotated genes and isoforms with NCBI Entrez GeneIDs and text descriptions. We further annotated differentially expressed genes with Gene Ontology (GO) [6] terms using NCBI annotation.
Enrichment Testing: We used DAVID (version 6.7) [7] for enrichment analysis of the set of differentially expressed genes to identify significantly enriched functional categories.
Genome_build: MM10
Supplementary_files_format_and_content: .xls table of normalized expression values for each gene for each sample

source_name: Isolated pancreatic acinar cells

growth_protocol: To induce acinar specific recombination, Ptf1a-CreERT, or Pdx1f/f;Ptf1aCreERT mice were administered tamoxifen (Sigma T5648) P.O. once daily for five days at 5 mg per 20g of body weight.

extract_protocol:
RNA was harvested using the RNEasy-Plus Kit (Qiagen;74136) and assessed for quality using the TapeStation (Agilent). Samples with RNA Integrity Number of 8 or greater were rRNA depleted using Ribo Minus (Invitrogen;K1550-04).
rRNA-depleted samples were prepared using the TruSeq mRNA SamplePrep v2 kit (Illumina;RS-122-2001, RS-122-2002). The entire fraction of 0.1-3 ug of rRNA depleted total RNA was fragmented and copied into first-strand cDNA using reverse transcriptase and random primers. cDNA 3'-ends were adenylated and adapters were ligated. One of the adapters had a 6-nucleotide barcode that was unique for each sample, allowing us to multiplex in a HiSeq flow cell (Illumina). Products were purified and enriched by PCR to create the final cDNA library. Libraries were checked for quality and quantity by TapeStation (Agilent) and qPCR using a library quantification kit for Illumina platforms (Kapa Biosystems;KK4835). They were clustered on the cBot (illumina) and sequenced 24 samples per lane on 6 lanes of a 50-cycle single end HiSeq 4000. HiSeq Control Software version 3.3.52 was used according to manufacturer?s protocols. Demultiplexing and Fastq file generation was done using bcl2fastq version 2.17.1.14.

treatment_protocol: Pancreata were harvested, washed twice in HBSS, minced, and incubated with 0.2 mg/mL Collagenase P (Roche;11249002001) at 37 degrees C for 15 minutes. Tissue was washed 3 times in HBSS containing 5% FBS and filtered through 500-um and 105-um polypropylene mesh (Spectrum Laboratories;106436 and 106418). Filtrate was centrifuged through 30% FBS in HBSS and resuspended in complete Waymouth media (1x Waymouth MB 752/1 medium, 50 mg/mL gentamycin, 0.4 mg/mL soybean trypsin inhibitor, and 1 mg/mL dexamethasone).  For RNAseq experiments, suspension was plated in non-adherent plates, treated with TGF? (50 ng/mL) and maintained at 370C in a 5% CO2 atmosphere for 1-3 days.

tag: strain background

tag: cell type

value: Isolated pancreatic acinar cells

tag: treatment

value: TGFa Day 3

tag: molecule subtype

value: Ribo depleted RNA


title: Sample_54972

value: TGFa Day 2


title: Sample_54971

value: TGFa Day 1


title: w6

source_name: ∆AcmybA 48 hour

tag: tissue

tag: genotype

value: AcmybA knockout


title: Sample_54969


title: Sample_54970

value: TGFa Day 0


title: Sample_54968


title: Sample_54965


title: Sample_54967


title: Sample_54966


title: Sample_54961


title: Sample_54962


title: Sample_54960


title: Sample_54963


title: Sample_54959


title: Sample_54957


title: Sample_54955


title: Sample_54954


title: Sample_54951


title: Sample_54953


title: Sample_54950


title: JWCB005 - Glucose - 1

value: Caldicellulosiruptor bescii strain JWCB005

value: delta-pyrF


title: Meningioma tumor 79

city: San Francisco

first: Harish

last: Vasudevan

state: CA

address: 555 Mission Bay Blvd South, Room 452H

department: Radiation Oncology

institute: University of California San Francisco

data_processing:
Adapters were trimmed and reads were filtered to remove bases that did not have an average quality score of 20 within a sliding window across 4 bases.
Aligned to hg19 human reference geneome using HISAT2 with default parameters
FPKM value estimates were obtained from Cufflinks
Genome_build: hg19
Supplementary_files_format_and_content: FPKM values from Cufflinks

organism: Homo sapiens

source_name: Meningioma tumor

extract_protocol:
Extracted in Trizol and Rneasy Mini Kit cleanup (Qiagen)
Libraries were prepared using the TruSeq RNA Library Prep Kit v2 from Illumina following standard protocol

value: Meningioma tumor


title: Meningioma tumor 81


title: Meningioma tumor 75


title: Meningioma tumor 61


title: Meningioma tumor 77


title: Meningioma tumor 74


title: Meningioma tumor 62


title: Meningioma tumor 50


title: Sample_54964


title: Meningioma tumor 70


title: Meningioma tumor 59


title: Sample_54958


title: Meningioma tumor 69


title: Sample_54956


title: Meningioma tumor 58


title: Sample_54952


title: Meningioma tumor 47


title: Meningioma tumor 43


title: Meningioma tumor 38


title: Meningioma tumor 45


title: Meningioma tumor 44


title: Meningioma tumor 40


title: Meningioma tumor 37


title: Meningioma tumor 34


title: Meningioma tumor 54


title: Meningioma tumor 32


title: Meningioma tumor 33


title: Meningioma tumor 57


title: Meningioma tumor 53


title: Meningioma tumor 31


title: Meningioma tumor 30


title: Meningioma tumor 24


title: Meningioma tumor 22


title: Meningioma tumor 19


title: Meningioma tumor 23


title: Meningioma tumor 14


title: Meningioma tumor 15


title: Meningioma tumor 12


title: Meningioma tumor 17


title: Meningioma tumor 9


title: Meningioma tumor 7


title: Meningioma tumor 2


title: Meningioma tumor 4


title: Control_3

city: Marburg

first: Anna

last: Antoniou

address: Karl-von-Frisch str.1

country: Germany

institute: Philipps University Marburg

data_processing:
Illumina Casava1.7 software used for basecalling.
Raw sequencing reads were trimmed from 3’ adapter (AGATCGGAAGAGCACACGTCT) and filtered according to quality using “percent = 90 and cut-off = 30” parameters of Fastx- Toolkit for fastq data on Galaxy (https://usegalaxy.org/). Reads were excluded if they only contained the adapter sequence, did not contain the adapter sequence prior to trimming, or were shorter than 15 nucleotides.
The remaining reads were mapped to the rat mature miRNAs (miRBase v21) using default parameters (one mismatch, 3 nt in the 3′ or 5′-trimming variants, 3 nt in the 3′-addition variants) of Miraligner software.
Genome_build: miRBase v21 (rat)
Supplementary_files_format_and_content: *.txt: Tab-delimited text files include raw values of miRNA for each Sample.

organism: Rattus norvegicus

source_name: primary cortical neurons

growth_protocol: Neurons were maintained in Neurobasal media containing 2% B27, 2mM GlutaMax, and 100μg/ml streptomycin and 100U/ml penicillin (Gibco, Invitrogen) in a humidified incubator with 5% CO2 at 37°C.

extract_protocol:
Total RNA was isolated using mirVana miRNA isolation kit (AM1561, Invitrogen).
Libraries were prepared using NEBNext® Multiplex Small RNA Library Prep Set for Illumina® (Set 1) kit (New England BioLabs) as per manufacturer’s instructions.

treatment_protocol: 7DIV primary neurons were stimulated with BDNF (100ng/ml) or vehicle control (0.1%BSA) for 20 minutes.

tag: strain/background

value: Sprague-Dawley

value: cortical neurons derived from embryos (E18)

tag: days in vitro

value: control


title: Meningioma tumor 5


title: Bdnf_2

value: brain-derived neurotrophic factor (BDNF)


title: Bdnf_3


title: Control_2


title: Bdnf_1


title: Control_1


title: SU_scRNA_seq_cell_94

city: Montréal

first: Claire

middle: Elizabeth

last: Nash

state: QC

address: 1001 Decarie Blvd

department: Urology

institute: Research Institute of McGill University Health Centre

data_processing:
Raw paired-end reads were trimmed using Trimmomatic v0.33, to a minimum length of 30 nucleotides. Illumina Nextera XT adapters were removed in palindrome mode. A minimum Phred quality score of 30 was required for the 3’end. Single end reads as well as paired end reads failing previous minimum quality controls were discarded. Individual read groups were aligned, using TopHat first against the rat transcriptome as defined by the Ensembl gene models version 83, with default parameters and the remaining unmapped genes to the Ensembl Rnor_6.0 reference rat genome from Illumina iGenomes web site.  Trimming rates and insert length were controlled on each read group based on metrics reported by Trimmomatic, and Picard v1.128 respectively. Aligned reads from multiple read groups belonging to the same sample were indexed, sorted and merged using sambamba v0.5.1, a faster implementation of the Samtools algorithms. Amplification duplicates were removed using Picard v1.128. Various quality controls from the RNA-SeQC package were used, including the genes detected, mapping rates, duplication rates, and intronic rate, based on metrics collected for each sample used.
Read counts were quantified using HTSeq Count using Ensembl Rnor_6.0 v83 GTF
Genome_build: Ensembl Rnor_6.0
Supplementary_files_format_and_content: Raw read count csv file was generated using HTSeq Count.

source_name: Female urogenital sinus

extract_protocol:
Urogenital tracts were removed from day of birth Wistar rats and microdissected into VMP and SU components using a Leica MZ6 dissection microscope. VMP and SU tissues were disscociated with 2mg/mL Collagenase 1A (Sigma-Aldrich, Missouri, USA) for 60 minutes at 37 degrees C. Dispersed fibroblasts were separated by centrifugation and passing through a 0.7um cell strainer (Falcon, Corning, New York, USA). Cell suspensions were centrifuged for 10 minutes at 500g and resuspended in LIVE/DEAD Cell Viability/Cytotoxicity Assay for mammalian cells (ThermoFisher, L-3224).   After a 10 minute incubation at room temperature, cells were centrifuged and resuspended in Cell Wash Buffer (Fluidigm).  Cell concentration, size and viability were verified on hemocytometers (Incyto DHC-N01-5) through bright field, GFP and RFP on a EVOS FL Auto microscope (ThermoFisher).  Single cell RNA libraries were constructed according to the Fluidigm protocol using C1 to generate libraries for RNA sequencing (PN 100-7168).
Single cell RNA libraries were constructed according to the Fluidigm protocol using C1 to generate libraries for RNA sequencing (PN 100-7168).  Briefly, full length mRNA-Seq libraries were generated from single-cells captured on the Fluidigm C1 platform using SMARTer Ultra Low RNA Kit (P/N 634936 Clontech). ERCC RNA Spike-In mix (P/N 4456740 ThermoFisher) was added to the lysis mix for normalization and quality control purposes. Full length cDNAs were converted into sequence ready libraries using Nextera XT DNA Sample Preparation Kit (P/N FC-131-1096 Illumina), and sequenced on an Illumina HiSeq 2000 with paired-end 100bp read option. The Fluidigm C1 platform captured 52 and 70 single cells from VMP and SU respectively. The average full length cDNA yield/min/max were 6.06ng(+/-0.12)/2.25ng/14.81ng for VMP and 8.51ng(+/-0.21)/2.55ng/26.6ng for SU.  Libraries from 52 VMP and 63 SU single cells were sequenced.

tag: tissue subset

value: Smooth muscle, peri-urethral mesenchyme and urethral epithelium (SU)

value: P0 (day of birth)

tag: sequencing method

value: single-cell RNA-sequencing


title: SU_scRNA_seq_cell_96


title: SU_scRNA_seq_cell_45


title: SU_scRNA_seq_cell_43


title: SU_scRNA_seq_cell_88


title: Meningioma tumor 6


title: SU_scRNA_seq_cell_89


title: SU_scRNA_seq_cell_42


title: SU_scRNA_seq_cell_40


title: SU_scRNA_seq_cell_93


title: SU_scRNA_seq_cell_92


title: SU_scRNA_seq_cell_90


title: SU_scRNA_seq_cell_41


title: SU_scRNA_seq_cell_87


title: SU_scRNA_seq_cell_38


title: SU_scRNA_seq_cell_37


title: SU_scRNA_seq_cell_39


title: SU_scRNA_seq_cell_85


title: SU_scRNA_seq_cell_84


title: SU_scRNA_seq_cell_83


title: SU_scRNA_seq_cell_82


title: SU_scRNA_seq_cell_34


title: SU_scRNA_seq_cell_36


title: SU_scRNA_seq_cell_79


title: SU_scRNA_seq_cell_95


title: SU_scRNA_seq_cell_44


title: SU_scRNA_seq_cell_80


title: SU_scRNA_seq_cell_81


title: SU_scRNA_seq_cell_33


title: SU_scRNA_seq_cell_32


title: SU_scRNA_seq_cell_78


title: SU_scRNA_seq_cell_76


title: SU_scRNA_seq_cell_30


title: SU_scRNA_seq_cell_74


title: SU_scRNA_seq_cell_28


title: SU_scRNA_seq_cell_75


title: SU_scRNA_seq_cell_25


title: SU_scRNA_seq_cell_23


title: SU_scRNA_seq_cell_27


title: SU_scRNA_seq_cell_70


title: SU_scRNA_seq_cell_68


title: SU_scRNA_seq_cell_67


title: SU_scRNA_seq_cell_21


title: SU_scRNA_seq_cell_16


title: SU_scRNA_seq_cell_66


title: SU_scRNA_seq_cell_18


title: SU_scRNA_seq_cell_62


title: SU_scRNA_seq_cell_63


title: SU_scRNA_seq_cell_13


title: SU_scRNA_seq_cell_14


title: SU_scRNA_seq_cell_59


title: SU_scRNA_seq_cell_15


title: SU_scRNA_seq_cell_58


title: SU_scRNA_seq_cell_11


title: SU_scRNA_seq_cell_24


title: SU_scRNA_seq_cell_56


title: SU_scRNA_seq_cell_12


title: SU_scRNA_seq_cell_09


title: SU_scRNA_seq_cell_05


title: SU_scRNA_seq_cell_54


title: SU_scRNA_seq_cell_50


title: SU_scRNA_seq_cell_08


title: SU_scRNA_seq_cell_01


title: SU_scRNA_seq_cell_02


title: Sample_48581

description: processed data file: POOLREP_MACS_PDX1_A_q0.01_treat_pileup.bw

library_source: genomic

data_processing:
Reads were downloaded and concatenated to yield one .fastq.gz file for each sample, and FastQC v0.10.0 was used to assess data quality for each file.
Reads were aligned to mm10 (UCSC) using bowtie2 with the following parameter settings: bowtie2 --phred33 -p 2 --very-sensitive -x genome -U Sample_43843_R1.fastq -S /ccmb/BioinfCore/Projects/Crawford_howcraw_mceachin_HI_CS3_1446/mapped_reads/Sample_43843_R1_k2.sam
MACS2 (v2.0.10) was used to identify peaks with the following parameter settings: macs2 callpeak -t /ccmb/BioinfCore/Projects/Crawford_howcraw_mceachin_HI_CS3_1446/mapped_reads/ChIPR_A/POOLREP_ChIPR_A.bam -c /ccmb/BioinfCore/Projects/Crawford_howcraw_mceachin_HI_CS3_1446/mapped_reads/INPUT_A/POOLINPUT_A.bam -n POOLREP_MACS_ChIPR_A_q0.01 -f BAM -g mm -B -q 0.01. Blacklisted islands were removed from the list of peaks (https://sites.google.com/site/anshulkundaje/projects/blacklists)
Peaks were annotated by HOMER (v4.6) for their locations relative to nearby genes.
Bigwig files were generated by counting read pile-ups using MACS.
Genome_build: mm10
Supplementary_files_format_and_content: BigWig

molecule: genomic DNA

source_name: NB490 cell line

growth_protocol: NB490 cells (kindly provided by Nabeel Bardeesy) were grown in DMEM supplemented with 10% FBS and 50 mg/mL gentamycin.

extract_protocol:
Chromatin was prepared from the NB490 cell line using the truChIP Chromatin Shearing Reagent Kit (Covaris;520154). Chromatin was sheared in 1 mL millitubes (Covaris;520135) using a Covaris S200 (5% duty cycle, intensity of 4, 200 cycles per burst, for 8 minutes) to obtain chromatin fragments 100-700 base-pairs in length. 50 ug of fragmented chromatin was incubated overnight with 2.5 uL of rabbit anti-PDX1 antibody (Millipore;07-696). Complexes were isolated using 50 uL of Dynabeads (Invitrogen;10004D) previously blocked with BSA (1 mg/mL).
2 ng of immuno-precipitated DNA was prepared as a standard Illumina library using the ThruPLEX DNA-seq Kit (Rubicon), according to manufacturer's protocol. Samples were PCR-amplified and pooled. Final libraries were checked for quality and quantity by TapeStation (Agilent) and qPCR using Kapa?s library quantification kit for Illumina Sequencing platforms (Kapa Biosystems;KK4835).  They were clustered on the cBot (Illumina) and sequenced 3 samples per lane on a 50-cycle single-end on a HiSeq 2000 in High Output mode using version 3 reagents.

tag: cell line

tag: chip antibody

value: PDX1 (Millipore 07-696)

tag: sample type

value: PDX1 ChIP


title: VMP_scRNA_seq_cell_94

value: Ventral mesenchymal pad (VMP)

tag: age


title: Sample_48582

value: Input


title: SU_scRNA_seq_cell_49


title: Sample_48580


title: Sample_43843

extract_protocol:
Chromatin was prepared from the NB490 cell line using the truChIP Chromatin Shearing Reagent Kit (Covaris;520154). Chromatin was sheared in 1 mL millitubes (Covaris;520135) using a Covaris S200 (5% duty cycle, intensity of 4, 200 cycles per burst, for 8 minutes) to obtain chromatin fragments 100 ? 700 base-pairs in length. 50 ug of fragmented chromatin was incubated overnight with 2.5 uL of rabbit anti-PDX1 antibody (Millipore;07-696). Complexes were isolated using 50 uL of Dynabeads (Invitrogen;10004D) previously blocked with BSA (1 mg/mL).
2 ng of immuno-precipitated DNA was prepared as a standard Illumina library using the ThruPLEX DNA-seq Kit (Rubicon), according to manufacturer?s protocol. Samples were PCR-amplified and pooled. Final libraries were checked for quality and quantity by TapeStation (Agilent) and qPCR using Kapa?s library quantification kit for Illumina Sequencing platforms (Kapa Biosystems;KK4835).  They were clustered on the cBot (Illumina) and sequenced 2 samples per lane on a 50-cycle single-end on a HiSeq 2000 in High Output mode using version 3 reagents.


title: Sample_43844


title: VMP_scRNA_seq_cell_93


title: Sample_54901

description: processed data file: POOLREP_MACS_ChIPR_A_q0.01_treat_pileup.bw

data_processing:
Reads were downloaded and concatenated to yield one .fastq.gz file for each sample, and FastQC v0.10.0 was used to assess data quality for each file.
Reads were aligned to mm10 (UCSC) using bowtie2 with the following parameter settings: bowtie2 --phred33 -p 2 --very-sensitive -x genome -U Sample_54899_R1.fastq -S /ccmb/BioinfCore/Projects/Crawford_howcraw_mceachin_HI_CS3_1446/mapped_reads/Sample_54899_R1_k2.sam
MACS2 (v2.0.10) was used to identify peaks with the following parameter settings: macs2 callpeak -t /ccmb/BioinfCore/Projects/Crawford_howcraw_mceachin_HI_CS3_1446/mapped_reads/ChIPR_A/POOLREP_ChIPR_A.bam -c /ccmb/BioinfCore/Projects/Crawford_howcraw_mceachin_HI_CS3_1446/mapped_reads/INPUT_A/POOLINPUT_A.bam -n POOLREP_MACS_ChIPR_A_q0.01 -f BAM -g mm -B -q 0.01. Blacklisted islands were removed from the list of peaks (https://sites.google.com/site/anshulkundaje/projects/blacklists)
Peaks were annotated by HOMER (v4.6) for their locations relative to nearby genes.
Bigwig files were generated by counting read pile-ups using MACS.
Genome_build: mm10
Supplementary_files_format_and_content: BigWig

extract_protocol:
Chromatin was prepared from isolated acinar cells using the truChIP Chromatin Shearing Reagent Kit (Covaris;520154). Chromatin was sheared in 1 mL millitubes (Covaris;520135) using a Covaris S200 (5% duty cycle, intensity of 4, 200 cycles per burst, for 8 minutes) to obtain chromatin fragments 100 ? 700 base-pairs in length. 50 ug of fragmented chromatin was incubated overnight with 2.5 uL of rabbit anti-PDX1 antibody (Millipore;07-696) or goat anti-PDX1 antibody (Abcam;ab47383). Complexes were isolated using 50 uL of Dynabeads (Invitrogen;10004D) previously blocked with BSA (1 mg/mL).
2 ng of immuno-precipitated DNA was prepared as a standard Illumina library using the ThruPLEX DNA-seq Kit (Rubicon), according to manufacturer's protocol. Samples were PCR-amplified and pooled. Final libraries were checked for quality and quantity by TapeStation (Agilent) and qPCR using Kapa?s library quantification kit for Illumina Sequencing platforms (Kapa Biosystems;KK4835).  They were clustered on the cBot (Illumina) and sequenced 3 samples per lane on a 50-cycle single-end on a HiSeq 2000 in High Output mode using version 3 reagents.

treatment_protocol: Pancreata were harvested, washed twice in HBSS, minced, and incubated with 0.2 mg/mL Collagenase P (Roche;11249002001) at 37 degrees C for 15 minutes. Tissue was washed 3 times in HBSS containing 5% FBS and filtered through 500-um and 105-um polypropylene mesh (Spectrum Laboratories;106436 and 106418). Filtrate was centrifuged through 30% FBS in HBSS and resuspended in complete Waymouth media (1x Waymouth MB 752/1 medium, 50 mg/mL gentamycin, 0.4 mg/mL soybean trypsin inhibitor, and 1 mg/mL dexamethasone).

value: Isolated pancreatic acinar cells

value: PDX1 (Abcam ab47383)


title: VMP_scRNA_seq_cell_43


title: VMP_scRNA_seq_cell_44


title: Sample_54900


title: SU_scRNA_seq_cell_52


title: VMP_scRNA_seq_cell_89


title: Sample_54899


title: VMP_scRNA_seq_cell_87


title: VMP_scRNA_seq_cell_38


title: PMG-Fingo-LPS-1

city: Ansan

first: Young Gyu

last: Chai

state: Gyeonggi-do

address: 55 Hanyangdaehak-ro

department: Molecualr & Life Science

country: South Korea

institute: Hanyang Univ.

data_processing:
Illumina CASAVA v1.8.2 software was used for basecalling.
To avoid low-quality data, we clipped and trimmed the reads using Trimmomatic (v0.33)
Quality controlled FASTQ files were alignment to Mus musculus UCSC mm10 reference genome sequence using the STAR (version 2.5.1) aligner software. To measure differential gene expression, DESeq2 was used.
The number of reads mapped to gene ID features were counted using STAR.
Genome_build: Mus musculus UCSC mm10
Supplementary_files_format_and_content: tab-delimited text files including normalized read counts for each sample obtained using STAR program

source_name: primary microglia cells

growth_protocol: PM were isolated from 3-day-old ICR mice as previously described with minor modifications. All experimental protocols were performed in accordance with Institutional Animal Care and Use Committee (IACUC) guidelines and approved by the IACUC committee of Hanyang University (HY-IACUC-2014-0164A and HY-IACUC-2015-0075). Briefly, whole brains of neonatal mice were dissected out of the skull, and blood vessels and meninges were carefully removed. Then, the tissues from whole brains obtained from 12 mice were pooled together, finely minced, and digested using a Neural Tissue Dissociation Kit-Postnatal Neurons (Miltenyi Biotec, Germany, 130-094-802). Next, the digested cells were passed through a 70-µm nylon cell strainer (BD Bioscience, Franklin Lakes, NJ) and seeded in poly-L-lysine coated T-75 flasks in DMEM/nutrient mixture F-12 (DMEM/F12, 1:1) containing 20% FBS (catalog # 26140; Gibco, Waltham, MA), 100 IU/ml penicillin and 10 µg/ml streptomycin (catalog # 15140) obtained from Invitrogen (Waltham, MA). The cells were maintained in a humidified incubator with a 95% air/5% CO2 atmosphere at 37°C. The medium was changed every 2-3 days. After two weeks in culture, the mixed glial cell cultures were shaken at 110 rpm at 37°C for 55 min, and the glial cell suspensions were collected from each flask and seeded on poly-L-lysine coated cell culture plates. Microglial cells were sub-plated and used for further experiments. More than 92 % of cells obtained were PM microglia as quantified by CD11b (rat monoclonal immunoglobulin G2b (IgG2b), clone: M1/70.15.11.5, Miltenyi Biotec Germany) FACS analysis (Figure 1C). Mouse microglial BV2 microglial cell lines were grown in high-glucose Dulbecco’s modified Eagle’s medium (DMEM) supplemented with 10% fetal bovine serum (FBS) (catalog # 26140; Gibco, Waltham, MA), 100 IU/ml penicillin, and 10 µg/ml streptomycin (catalog # 15140; Invitrogen, Waltham, MA). The cells were maintained in a humidified incubator with 95% air and a 5% CO2 atmosphere at 37°C. Medium containing the appropriate agents was replaced every other day. The cells were maintained in a humidified incubator with 95% air and a 5% CO2 atmosphere                            at 37°C and FTY720 (2-amino-2-[2-(4-octylphenyl) ethyl] propane-1, 3-diol) were purchased from Selleckchem, Houston, TX and dissolved in dimethyl sulfoxide (DMSO, Sigma- Aldrich, St. Louis, MO) as a 10 mM stock solution; the stock solution was diluted in DMEM for experiments

extract_protocol:
Total RNA was extracted using RNAiso Plus (Takara Bio Inc., Shiga, Japan) and a QIAGEN RNeasy® Mini kit (QIAGEN, Hilden, Germany). PM cells were completely lysed using RNAiso Plus, and then 200 μl of chloroform was added. The tubes were then inverted for 5 min. The mixture was centrifuged at 12,000 x g for 15 min at 4°C, and the upper phase was placed into a new tube. A 600 μl volume of 70% ethanol was added, and the mixture was applied to an RNeasy mini column. The column was washed with wash buffer. To elute the RNA, RNase-free water (30 μl) was added directly onto the RNase mini column, which was then centrifuged at 12,000 x g for 3 min at 4°C. To deplete ribosomal RNA (rRNA) from the total RNA preparations, a RiboMinus Eukaryote kit (Life Technologies, Carlsbad, CA) was used according to the manufacturer’s instructions. RNA libraries were created using a NEBNext® Ultra™ directional RNA library preparation kit for Illumina® (New England Biolabs, Ipswich, MA). The obtained rRNA-depleted total RNA was fragmented into small pieces using divalent cations at elevated temperatures. First-strand cDNA was synthesized using reverse transcriptase and random primers, and second-strand cDNA synthesis was then performed using DNA polymerase I and RNase H. The cDNA fragments were processed using an end-repair reaction after the addition of a single ‘A’ base, followed by adapter ligation. These products were purified and amplified using PCR to generate the final cDNA library. The cDNA fragments were sequenced using an Illumina HiSeq2000. Biological triplicate RNA sequencing was performed on independent RNA samples from the FTY720-stimulated PM cells: FTY720_4 h (3 samples) and LPS+ FTY720_4 h (3 samples).
RNA libraries were prepared for sequencing using standard Illumina protocols

treatment_protocol: Murine microglial PM, were treated with FTY720 (1 uM), and with LPS (10 ng/ml) for 4 hrs.

value: primary microglia cells

tag: agent

value: FTY720 (1 uM) and LPS (10 ng/ml) for 4 hrs


title: PMG-Fingo-LPS-3


title: VMP_scRNA_seq_cell_37


title: PMG-Fingo-LPS-2


title: VMP_scRNA_seq_cell_86


title: VMP_scRNA_seq_cell_91


title: VMP_scRNA_seq_cell_34


title: PMG-Fingo-3

value: FTY720 (1 uM) for 4 hrs


title: VMP_scRNA_seq_cell_84


title: PMG-Fingo-1


title: PMG-Fingo-2


title: VMP_scRNA_seq_cell_79


title: VMP_scRNA_seq_cell_81


title: VMP_scRNA_seq_cell_76


title: VMP_scRNA_seq_cell_77


title: SatuaratedMut_intronless_output1

city: Bar Harbor

first: Shengdong

last: Ke

state: ME

address: 600 Main Street

institute: The Jackson Laboratory

data_processing:
Sequencing was carried out on an Illumina platform with reads of 74 nt. Reads were filtered for accuracy by accepting only those that exactly matched the expected sequence of the non-mutated stretch of 6 nt beyond the mutated region and that contained no changes other than those designed. This filter typically removed ~50% of the reads. Approximately 10 million reads were collected for the input DNA and approximately 3 million from each of the 2 independent transfections experiments. The average number of reads for the input DNA was 1806 and all but 2 mutant sequences had at least 50 reads. The average number of reads for the output cDNA was 1105 but these ranged widely as expected (0 to 21340). Nine percent of the output reads were zero, informative of a very poor splicing efficiency; these mutants were given a pseudo count of 1 when logarithms were calculated, as for LEI. Thus very low splicing values may be overestimated. The overall dynamic range of mRNA detection was over 3 orders of magnitude.
Reads for each molecule were transformed into EIs by summing the outputs of the 2 replicate transfections (which agreed to an R2 of >0.99), converting the sum to a proportion of total output reads and dividing by the analogous proportion for input DNA. Psi values were calculated from a calibration curve (Supplemental_Fig_S19) relating the traditionally measured psi values from transfections with the EI values of the 10 relative wild type molecules as previously described (Ke et al., 2011).
For RBP-IP experiments, the fastq files represent raw combined R1 and R2 paired end reads for the protein/input/control, as specified in each file name. First, sequencing data was classified by demanding the presence of a  perfect bar code, which was an 8mer sequence serving as a unique identifier for a protein and run (ex: CGTCTAGA for U2AF65, run 1); Then, paired-ends were combined by finding the greatest sequence homology (between the two paired ends) over a 7 base window. Once the index that achieved  maximum homology was identified, the final read was generated, nt by nt, by taking a base from the end (R1 vs R2) with the highest quality score and assigning that base the corresponding quality score. The summary.txt files contain a summary of the final total reads per sequence. For each run, a read was considered valid (and incorporated into the count) when the sequence contained, at any position, a perfect match to a designed exon from our experiment. Given the strict criteria, Phred quality scores were not taken into consideration for this step.
Supplementary_files_format_and_content: txt files, two columes: sequence content and counts

source_name: sythesized minigene

extract_protocol:
Exon-RBP complexes were assembled in a splicing competent nuclear extract prepared from HeLa cells; in transient transfection HeLa cells differentiated a sample of cloned mutant minigenes in a manner similar to HEK293 cells (Supplemental Fig_S20.jpg).  The saturation mutagenesis minigene library was PCR amplified with Phusion High-Fidelity DNA Polymerase (New England Biolabs) for a limited number of cycles (15) using the primers T7LibFwd TAATACGACTCACTATAGGACCTCTTCTTCTTTTCTAGA and LibRev gccagctagcACTCACTTG. The resulting T7 PCR DNA library was transcribed in vitro with MegaScript (Ambion) according to the manufacturer’s protocol. The resulting saturation mutagenesis RNA library was cleaned with Quick Spin Sephadex Columns for RNA (Roche), purified by phenol/chloroform extraction and precipitated with ammonium acetate. Nuclear extract preparation was adapted from Hartmuth et al. . Briefly, HeLa cells were grown in suspension to 5 to 7 x105 cells per ml. Pelleted cells were Dounce homogenized in buffer A (10mM Hepes KOH pH7.9, 10mM KCl, 1.5mM MgCl2, 0.5mM DTT, 0.25mM PMSF). Resulting pelleted nuclei were dispersed in a Dounce homogenizer with 15 strokes of the B-type pestle in buffer C (20mM Hepes KOH ph7.9, 600mM KCl, 1.5mM MgCl2, 0.2mM EDTA pH8, 25% glycerol, 0.5mM DTT, 0.25mM PMSF). The nuclear extract obtained after centrifugation at 30,000xg for 40 min was dialyzed 2 x 2h against a minimum 50-fold volume of buffer D (20mM Hepes KOH pH 7.9, 0.1 M KCl, 0.2mM EDTA pH 8.0, 10% (v/v) glycerol, 0.5mM DTT, 0.25mM PMSF) , flash frozen in liquid nitrogen, stored at -80ºC and tested for splicing before performing RNA immunoprecipitation (IP).
The microarray primer extension method and the subsequent double-stranded DNA linker ligation were described by Ray et al. (2009). A custom 60 bp 176,000 (4X44K) element microarray was purchased from Agilent. It was comprised of 5550 equally represented mutant sequences (555 per Hexmut) and 10 relative wild type sequences equally overrepresented at ~30 times the number of a mutant sequence. The 3’ end of each 60‑mer was constant, representing 13 positions from -1 to +12 relative to the 3’ end of the WT-1 exon 5 (Ke et al, 2011) with the sequence of “cttcttttctagA”. The remaining 47 nucleotides were specified as wild types or mutants, as described in Fig. 1 of the Results. The 19 nt double-stranded DNA linker with the sequence of “CAAgtgagtggacaatgcg” was ligated to the free end of the 60 nt double-stranded DNA probes. The eluted 79nt single-stranded DNA library (e.g. cttcttttctagAGTTGCTGCTGGGAGCTCCAGCACAGTGAAATGGACAGAAGGGCAGAGCAAgtgagtggacaatgcg) was then PCR amplified using the flanking primer pair F: ccccacctcttcttcttttctagA; R: the reverse complement of CAAgtgagtggacaatgcg), generating the 90nt double-stranded DNA library that was used to construct the 3kb full mini-gene library by overlap extension PCR (Fig 18). The relative wild type 60‑mer probe sequence of HMA is shown on line 1 below. All possible single and tandem double base changes were designed as shown in Fig. 1 and below: Line 1 shows the wild type sequence of HMA where the exon is underlined and the variable 6‑mer than distinguishes Hexmuts is double underlined. Starting at positon 2 of the exon, all 12 dinucleotides that changed the first base of the wild type dimer sequence were created (bold, lines 2 to 13). There are 15 possible mutant dinucleotides that can start at position 2; the 3 dimers missing in lines 2 to 13 appear when position 3 dimers are created, as seen underlined on lines 17, 21 and 25. The complete set of mutations covering the region 2 to 48 comprise 555 mutants, with the last position, 48, being necessarily comprised of only 3 single base substitutions.
Incubations of the microarray were carried out in an Illumina hybridization oven with a rotating slide carrier. A 5’‑Cy3-labeled primer complementary to the 13 nt at 3’ end of all microarray bound probes was extended with T4 DNA polymerase. The Cy3 label afforded confirmation of the double strandedness by examining the slide in a microarray scanner. The double stranded probes were then ligated  to a double stranded oligomer representing the 20 nt upstream of the 5’ end of the sense strand of the probe. The sense strand of the 20‑mer carried a Cy5 label at the 5’ end and the antisense strand carried a ddC appended to the 3’ end to prevent primer self ligation and a phosphate included on the 5’ end for ligation to the probe. The Cy5 label afforded confirmation of the ligation efficiency by scanning. The single stranded extension-ligation products were stripped from the slide with NaOH  at 65C for 20 minutes and used as a mega-primer for minigene construction by overlap extension PCR as shown in Supplemental_Fig_S18.jpg.

treatment_protocol: The ~3 kB minigenes (Fig. 1) were prepared by 3 fragment overlap PCR and used directly (not cloned) for transfection of HEK293 cells exactly as described previously (Ke et al, 2001). There we showed that linear PCR products are expressed in transient transfections as well as plasmids. The framework dhfr minigene was pMA-Universal (Arias et al., 2015). RNA was isolated 24 h after transfection of two 100 mm dishes per biological replicate and was converted to cDNA and amplified by PCR (Ke et al., 2011). Both the input DNA and the amplified cDNA from two independent transfections were prepared using primers shown schematically in Fig. S17.

tag: experiment

value: output for saturated mutatgensis control

value: sythesized DNA


title: VMP_scRNA_seq_cell_78


title: SatuaratedMut_intronless_input3

value: input for saturated mutatgensis control


title: SatuaratedMut_intronless_input2


title: SatuaratedMut_intronless_output2


title: SatuaratedMut_intronless_input1


title: VMP_scRNA_seq_cell_29


title: SatuaratedMut_output3

value: output for saturated mutatgensis screening


title: SatuaratedMut_output2


title: VMP_scRNA_seq_cell_27


title: VMP_scRNA_seq_cell_26


title: SatuaratedMut_output1


title: SatuaratedMut_input4

value: input for saturated mutatgensis screening


title: SatuaratedMut_input3


title: SatuaratedMut_input2


title: SatuaratedMut_input1


title: VMP_scRNA_seq_cell_31


title: VMP_scRNA_seq_cell_72


title: SU_Tag_seq

data_processing:
Raw sequencing reads were trimmed to 17bp to remove adaptor sequences and restriction digestion sites. Reads were quality controlled using FastQC to keep only reads with a mean quality score of 20 and above. Reads were aligned to the rat genome (Ensembl Rnor_6.0) using the Bowtie2 algorithm (default settings). Reads aligned to random contigs and mitochondrial DNA were removed and only uniquely mapped reads with a mapping quality >= 25 were used for further analysis.
Read counts were quantified using the summarizeOverlaps function from the GenomicAlignments R package
Genome_build: Ensembl Rnor_6.0
Supplementary_files_format_and_content: Raw read count csv file was generated using summarizeOverlaps from GenomicAlignments.

extract_protocol:
Urogenital tracts were removed from day of birth Wistar rats and microdissected into VMP and SU components using a Leica MZ6 dissection microscope. For RNA-sequencing only, VMP and SU tissues were disscociated with 2mg/mL Collagenase 1A (Sigma-Aldrich, Missouri, USA) for 60 minutes at 37 degrees C. Dispersed fibroblasts were separated by centrifugation and passing through a 0.7um cell strainer (Falcon, Corning, New York, USA)
Microdissected tissues (VMP and SU) were processed for digital gene expression Tag-profiling using NlaIII and a protocol provided by Illumina followed by sequencing on an llumina GAIIX (1x 50 SE) at one lane per sample (25-30m reads). PolyA+ RNA was purified, cDNA synthesised, digested with NlaIII and ligated to Adaptor 1 (containing an Mme1 site). Samples were digested with Mme1, ligated to Adaptor 2, and PCR amplified, followed by gel electrophoresis and purification of 85bp fragments that were sequenced

tag: strain

value: Wistar

tag: Sex

value: female

value: urogenital sinus

value: Smooth muscle, peri-urethral mesenchyme and urethral epithelium (SU)

value: Tag-sequencing


title: VMP_scRNA_seq_cell_73

value: Ventral mesenchymal pad (VMP)

value: P0 (day of birth)

value: single-cell RNA-sequencing


title: VMP_scRNA_seq_cell_25


title: Meningioma tumor 83

value: Meningioma tumor


title: Sample_7764_TAC_diesel_exhaust_exposure

city: Seattle

first: James

middle: William

last: MacDonald

state: WA

address: 4225 Roosevelt Way NE

department: Environmental and Occupational Health Sciences

institute: University of Washington

description: common adaptor: GGCCAAGGCG

data_processing:
Sequences were trimmed of adaptor sequences (GGCCAAGGCG) using cutadapt version 1.6, excluding any resulting reads that were shorter than 16 nt in length. Trimmed reads were then aligned against the UCSC mm10 genome using the standalone version of the subread aligner version 1.4.2 using default arguments. Counts per gene were generated using the featureCounts function of the Bioconductor Rsubread package (version 1.12.1), using the built-in mm10 gene annotations.
Genome_build: mm10
Supplementary_files_format_and_content: Processed data consist of tab-delimited counts per gene for each sample. The first column contains the Entrez Gene ID, and the remaining columns are counts for each gene per sample.

source_name: Heart tissue, diesel exhaust exposure, TAC surgical procedure

extract_protocol:
RNA was extracted from frozen heart tissue using the Qiagen miRNeasy kit, following the manufacturer's recommendations. Quality and quantity were assessed using an Agilent 2100 Bioanalyzer, and NanoDrop 8000 spectrophotometer, respectively.
RNA samples were depleted of ribosomal RNA using the Low Input RiboMinus Eukaryotic System v2 kit (Thermo Fisher Scientific). The cDNA library was then created with the ribosomal depleted RNA using the Ion Total RNA-Seq Kit v2 (Thermo Fisher Scientific). This included fragmentation via RNase III, hybridizing and ligating the RNA, performing reverse transcription, and amplification.  Samples were labeled with the Ion Xpress RNA-Seq Barcode 1-16 Kit (Thermo Fisher Scientific) to enable multiplexing 4 samples per run.  Templating, loading and sequencing were carried out using the Ion PI Template OT2 200 Kit v2, the Ion PI Chip Kit v2, and the Ion PI Sequencing 200 Kit v2 respectively (all from Thermo Fisher Scientific).

value: heart

value: 15 weeks


title: VMP_scRNA_seq_cell_23


title: Sample_7767_TAC_diesel_exhaust_exposure

description: common adaptor: GGCCAAGGCG


title: Sample_7762_TAC_diesel_exhaust_exposure

description: common adaptor: GGCCAAGGCG


title: VMP_Tag_seq


title: VMP_scRNA_seq_cell_24


title: VMP_scRNA_seq_cell_33


title: IP2_U2AF35

value: output for U2AF35 IP2

tag: rip antibody

value: rabbit anti-U2AF35

tag: rip antibody vendor

value: Abcam

tag: rip antibody cat. #


title: Sample_7765_Sham_diesel_exhaust_exposure

description: common adaptor: GGCCAAGGCG

source_name: Heart tissue, diesel exhaust exposure, Sham surgical procedure

tag: tissue


title: VMP_scRNA_seq_cell_28


title: IP2_U170K

value: output for U170K IP2

value: goat anti-U170k

value: Santa Cruz Biotechnology

tag: molecule subtype

value: sythesized DNA


title: VMP_scRNA_seq_cell_20


title: VMP_scRNA_seq_cell_21


title: Sample_7760_Sham_diesel_exhaust_exposure

description: common adaptor: GGCCAAGGCG


title: VMP_scRNA_seq_cell_66


title: IP2_TIAR

value: output for TIAR IP2

value: rabbit anti-TIA1


title: Sample_7756_TAC_filtered_air_exposure

description: common adaptor: GGCCAAGGCG

source_name: Heart tissue, filtered air exposure, TAC surgical procedure


title: IP2_TIA1

value: output for TIA1 IP2


title: VMP_scRNA_seq_cell_65


title: Sample_7750_TAC_filtered_air_exposure

description: common adaptor: GGCCAAGGCG


title: IP2_RBM6

value: output for RBM6 IP2

value: rabbit anti-RBM6

value: Bethyl


title: Sample_7753_TAC_filtered_air_exposure

description: common adaptor: GGCCAAGGCG


title: VMP_scRNA_seq_cell_64

value: Ventral mesenchymal pad (VMP)

value: P0 (day of birth)

value: single-cell RNA-sequencing


title: IP2_RBM4

value: output for RBM4 IP2

value: rabbit anti-RBM4

value: Proteintech


title: IP2_DAZAP1

value: output for DAZAP1 IP2

value: rabbit anti-DAZAP1

value: Abcam


title: VMP_scRNA_seq_cell_63


title: VMP_scRNA_seq_cell_16


title: Sample_7752_Sham_filtered_air_exposure

description: common adaptor: GGCCAAGGCG

source_name: Heart tissue, filtered air exposure, Sham surgical procedure

value: heart

value: 15 weeks


title: Sample_7763_Sham_diesel_exhaust_exposure

description: common adaptor: GGCCAAGGCG


title: IP2_Input

value: input for IP2

value: sythesized DNA


title: IP2_Rabbit IgG

value: output for Rabbit IgG IP2

value: normal rabbit IgG

value: Santa Cruz Biotechnology


title: Sample_7754_Sham_filtered_air_exposure

description: common adaptor: GGCCAAGGCG


title: Sample_7757_Sham_filtered_air_exposure

description: common adaptor: GGCCAAGGCG


title: VMP_scRNA_seq_cell_62


title: post-germination

city: Berkeley

first: Sean

middle: D.

last: Gallaher

state: California

address: 176 Stanley Hall, room 408

department: California Institute for Quantitative Biosciences

institute: UC Berkeley

description: processed data file: fpkms.sexual.tsv

data_processing:
Basecalling performed using Illumina RTA software.
Conversion of qseq to fastq format and demultiplexing by in-house scripts
Removal of contaminating rRNA reads by alignment to rRNA+snoRNA.fasta with RNA-STAR (STAR_2.4.0j). Unmapped reads selected for further analysis with "--outReadsUnmapped Fastx" option of STAR.
Alignment to C.reinhardtii nuclear genome (Phytozome v5.5) plus chloroplast genome (CPv4) plus mitochondrial genome (MTv4) by RNA-STAR (STAR_2.4.0j) with default parameters except --alignIntronMax 5000
SAM alignments sorted and converted to BAM format by samtools (v1.3).
Counts per gene and FPKMs calculated by cuffdiff (v2.0.2) with "--multi-read-correct --max-bundle-frags 1000000000 --library-type fr-firststrand"
Genome_build: Cre.nuc.v5.5.cp.v4.4.mt.v4.4
Supplementary_files_format_and_content: tab-delimited text files (.tsv) containing transcript abundance determinations calculated as FPKMs

organism: Chlamydomonas reinhardtii

source_name: liquid culture

growth_protocol: Vegetatively growing cells following germination.

extract_protocol:
Total RNA was purified from pelleted cells by freezing in SDS/proteinase K digestion buffer followed extraction with Trizol reagent.
~2 ug total RNA was used as starting material for either rRNA-depletion by RiboZero Plant Leaf kit or for enrichment of poly-A transcripts by binding oligo-T beads (indicated by "library protocol" above). The resulting RNA was then used to generate strand-specific Illumina libraries.

value: CC-620 x CJU10

tag: media

value: tris acetate phosphate medium

tag: light regime

value: continuous light

tag: library protocol

value: RiboZero


title: IP2_Goat IgG

value: output for Goat IgG IP2

value: normal goat IgG


title: zygote

description: processed data file: fpkms.sexual.tsv

growth_protocol: Zygotes 1 d after cross of mt+ and mt- gametes.

value: high salt medium -N

value: continuous dark


title: IP1_U2AF65

value: output for U2AF65 IP1

value: mouse anti-U2AF65

value: Sigma


title: IP1_U1A

value: output for U1A IP1

value: mouse anti-U1A

value: Abcam


title: VMP_scRNA_seq_cell_10

tag: tissue subset

value: Ventral mesenchymal pad (VMP)

value: P0 (day of birth)

tag: sequencing method

value: single-cell RNA-sequencing


title: VMP_scRNA_seq_cell_60


title: mt- gamete

description: processed data file: fpkms.sexual.tsv

growth_protocol: Gametes of mt- strain.


title: mt+ gamete

description: processed data file: fpkms.sexual.tsv

growth_protocol: Gametes of mt+ strain.


title: IP1_SRSF7

value: output for SRSF7 IP1

value: rabbit anti-SRSF7

value: Bethyl

value: sythesized DNA


title: IP1_SRSF1

value: output for SRSF1 IP1

value: mouse anti-SRSF1

value: Santa Cruz Biotechnology


title: VMP_scRNA_seq_cell_56


title: mt- vegetative

description: processed data file: fpkms.sexual.tsv

growth_protocol: Vegetatively growing culture of mt- strain.

value: high salt medium

value: continuous light

value: RiboZero


title: mt+ vegetative

description: processed data file: fpkms.sexual.tsv

growth_protocol: Vegetatively growing culture of mt+ strain.


title: IP1_HNRNPL

value: output for hnRNPL IP1

value: rabbit anti-hnRNPL


title: VMP_scRNA_seq_cell_55


title: VMP_scRNA_seq_cell_08


title: whiB6 deletion mutant Complemented, Rep B

city: East Lansing

first: Robert

middle: B.

last: Abramovitch

state: MI

address: 567 Wilson Road

department: Microbiology and Molecular Genetics

institute: Michigan State University

data_processing:
Illumina bcl2fastq (v1.8.4) was used to de-multiplex and convert reads to FASTQ format
Reads from each lane were combined into individual files using standard UNIX commands
Trimmomatic (v0.33)  was used to trim low quality bases and remove adapter sequences with a 4 bp sliding window, cutting when the read quality dropped below 15 or read length was less than 36 bp.
Bowtie (v1.1.2) was used to align trimmed reads to the M. tuberculosis CDC1551 genome (NCBI accession AE000516) with the -S option to produce SAM files as output.
HTSeq software suite (v0.6.0) was used to analyze aligned and determine counts per gene feature in the M. tuberculosis CDC1551 genome
edgeR (v3.2) was used to normalize data by estimating effective library sizes using trimmed mean M-values
R was used for statistical analysis and differential gene expression by fitting a negative binomial model to each set of conditions and testing for differences utilizing the GLM functionality edgeR package
Genome_build: Mycobacterium marinum M reference genome (GCA_000018345.1)
Supplementary_files_format_and_content: Processed files are presented as .xlsx spreadsheets and represents the average of 2 biological replicates

organism: Mycobacterium marinum

source_name: Bacterial Cell

growth_protocol: M. marinum cultures were grown to saturation in 7H9 defined broth and diluted to an OD600 of 0.8 in 50 ml of Sauton's defined broth with 0.05% Tween 80. After 48 h of growth at 30°C, cells were harvested by centrifugation. Whole-cell lysates were generated by lysing the mycobacterial cells with a Mini-Bead Beater-24 (BioSpec).

extract_protocol:
Total RNA was extracted using an RNeasy minikit (Qiagen) according to the manufacturer's recommendation with the following modifications. After bacterial cells were harvested, they were resuspended in 1 ml of RLT buffer containing 0.1% 2-mercaptoethanol and transferred to a 2-ml screw-cap tube containing glass beads. The mixture was homogenized in a mini-bead beater (BioSpec Products) with three 30-s pulses. RNA samples were treated with DNase I. Total RNA quality was assessed using an Agilent Bioanalyzer 2100 and the Agilent RNA 6000 Nano Reagent kit (16S rRNA chip) in the University of Notre Dame Genomics Core Facility.
cDNA libraries were constructed using the Illumina TruSeq RNA library preparation kit (v2), omitting the polyA selection step. Multiplexing sequences are included in the title of the associated fastq.gz files.
After library quality control, sample libraries were pooled and sequenced in two lanes of an Illumina HiSeq 2500 Rapid Run flow cell (v1) in 50bp, single-end read format (SE50). After the sequencing run, reads were de-multiplexed and converted to FASTQ format using the Illumina bcl2fastq (v1.8.4) script.

value: M strain (ATCC BAA-535)

tag: genotype

value: whiB6 deletion mutant Complemented


title: VMP_scRNA_seq_cell_13

value: Ventral mesenchymal pad (VMP)


title: whiB6 deletion mutant Complemented, Rep A


title: VMP_scRNA_seq_cell_09

value: P0 (day of birth)


title: SU_scRNA_seq_cell_69

value: Smooth muscle, peri-urethral mesenchyme and urethral epithelium (SU)

value: single-cell RNA-sequencing


title: IP1_HNRNPI

value: output for hnRNPI IP1

value: mouse anti-hnRNPI


title: IP1_HNRNPD

value: output for hnRNPD IP1

value: rabbit anti-hnRNPD/AUF1

value: Abcam

value: sythesized DNA


title: whiB6 deletion mutant, Rep B

value: whiB6 deletion mutant


title: VMP_scRNA_seq_cell_15


title: VMP_scRNA_seq_cell_54


title: whiB6 deletion mutant, Rep A


title: IP1_HNRNPD0

value: output for hnRNPD0 IP1


title: media -Fe - poly-A

description: processed data file: fpkms.iron_PAvsRZ.tsv

growth_protocol: 4 h after transfer to medium without Fe, poly-A enriched library.

value: tris acetate phosphate medium -Fe

value: continuous light

value: poly-A


title: eccCb1 ochre mutant Complemented, Rep B

value: M strain (ATCC BAA-535)

value: eccCb1 ochre mutant Complemented


title: IP1_HNRNPA1

value: output for hnRNPA1 IP1

value: mouse anti-hnRNPA1

value: Sigma


title: media +Fe - poly-A

growth_protocol: Growth in media with Fe, poly-A enriched library.

value: tris acetate phosphate medium +Fe


title: eccCb1 ochre mutant Complemented, Rep A


title: media -Fe - RiboZero

growth_protocol: 4 h after transfer to medium without Fe, RiboZero library.

value: RiboZero


title: IP1_Input

value: input for IP1


title: VMP_scRNA_seq_cell_06

value: Ventral mesenchymal pad (VMP)

value: P0 (day of birth)

value: single-cell RNA-sequencing


title: eccCb1 ochre mutant , Rep B

value: eccCb1 ochre mutant


title: media -Cu

description: processed data file: fpkms.copper.tsv

growth_protocol: Growth in media without Cu.

value: tris acetate phosphate medium -Cu


title: media +Fe - RiboZero

growth_protocol: Growth in media with Fe, RiboZero library.


title: IP1_Rabbit IgG

value: output for Rabbit IgG IP1

value: normal rabbit IgG

value: Santa Cruz Biotechnology

value: sythesized DNA


title: VMP_scRNA_seq_cell_50


title: media +Cu

description: processed data file: fpkms.copper.tsv

growth_protocol: Growth in media with Cu.

value: tris acetate phosphate medium +Cu

value: continuous light


title: diurnal growth - light

description: processed data file: fpkms.diurnal.tsv

growth_protocol: Diurnally grown culture sampled 1 h into the light phase.

value: high salt medium

value: 12h light / 12h dark


title: IP1_Mouse IgG

value: output for Mouse IgG IP1

value: normal mouse IgG


title: diurnal growth - dark

description: processed data file: fpkms.diurnal.tsv

growth_protocol: Diurnally grown culture sampled at the end of the dark phase.

value: RiboZero


title: Wild Type, Rep B

value: M strain (ATCC BAA-535)

value: Wild Type


title: VMP_RNA-sequencing

extract_protocol:
Bulk samples of cells were taken prior to single-cell viability and lysis protocols. RNA was extracted using a Qiagen Rneasy kit as per manufacturers instructions.
Libraries were constructed as per single-cell protocols using 5ng of purified total RNA with cDNA and library preparation performed on a BioRad T100 thermal cycler.

value: Ventral mesenchymal pad (VMP)

value: P0 (day of birth)

value: RNA-sequencing


title: SU_RNA-sequencing

value: Smooth muscle, peri-urethral mesenchyme and urethral epithelium (SU)


title: IP1_Goat IgG

value: output for Goat IgG IP1

value: normal goat IgG


title: Wild Type, Rep A


title: VMP_scRNA_seq_cell_53

value: single-cell RNA-sequencing


title: VMP_scRNA_seq_cell_05


title: H12_plate4_396

status: Public on Nov 26 2017

city: Boston

first: Ruben

last: Dries

address: 360 Longwood Ave

department: Computational Biology

institute: DFCI

description: single cell RNAseq T cells after G1T28 treatment

data_processing:
Sequenced reads were mapped to the human gemome (hg19) by STAR for mapping information.
Estimated transcript counts were retrieved via the pseudo-aligner Kallisto using the mouse Gencode vM1 annotation.
Only single cell samples with at least 100.000 paired-end reads and a minimum of 20% aligning to the transcriptome were retained for further analysis.
PCA combined with density based clustering was performed to remove outlier cells based on a combination of quality metrics.
Genes were considered not expressed if TPM < 1 and were removed if not detected in at least 10% of remaining cells.
The R package SCONE was used to define the optimal normalization approach for the data in a 2 step approach. First we used the plates that contain ERCC spike -ins (plate 3 and plate4) and next we used the most stable genes identified in the previous step.
Density based clustering was used to identify different clusters after initial feature selection by selecting most variable and informative genes.
Pairwise comparisons to identify differentially distributed genes were performed with the R package scDD.
Genome_build: mm9
Supplementary_files_format_and_content: log_normalized_expr_plus1.txt: txt file with log normalized expression counts + 1
Supplementary_files_format_and_content: information_raw_data.xlsx
Supplementary_files_format_and_content: phenodata_samples.txt

source_name: T cell

growth_protocol: Genetically engineered mouse model (GEMM) of conditional activating mutation that harboring endogenous Kras (KrasLSL-G12D/+) were crossed with p53 conditional knockout Trp53fl/fl. CRE recombinase was induced through intranasal inhalation of 5x106 p.f.u. adeno-Cre (University of Iowa adenoviral core).

extract_protocol:
Live CD45+ and CD3+ cells were sorted in 2ul of Qiagen TCL buffer in 96-well plates
SmartSeq2 protocol with modifications (Trombetta et al., 2014)

treatment_protocol: G1T28, 100 mg/kg, PO, QD

value: Lung tumor

value: Mixed background

tag: gemm model

tag: cell type

value: tumor infiltrating T cells


title: H09_plate4_372


title: H08_plate4_364


title: H10_plate4_380


title: H11_plate4_388


title: H06_plate4_348

description: single cell RNAseq control T cells from tumor


title: H04_plate4_332


title: H03_plate4_324


title: H07_plate4_356


title: H05_plate4_340


title: H02_plate4_316


title: eccCb1 ochre mutant , Rep A

value: eccCb1 ochre mutant


title: G09_plate4_371


title: G12_plate4_395


title: H01_plate4_308


title: G08_plate4_363


title: G10_plate4_379


title: G05_plate4_339


title: G06_plate4_347


title: G03_plate4_323


title: G02_plate4_315


title: G01_plate4_307


title: F11_plate4_386


title: F12_plate4_394


title: F09_plate4_370


title: F08_plate4_362


title: F04_plate4_330


title: F03_plate4_322


title: G11_plate4_387


title: F05_plate4_338


title: F07_plate4_354


title: E12_plate4_393


title: F10_plate4_378


title: G07_plate4_355


title: E11_plate4_385


title: G04_plate4_331


title: E10_plate4_377


title: E07_plate4_353


title: E08_plate4_361


title: E06_plate4_345


title: E09_plate4_369


title: E04_plate4_329


title: D12_plate4_392


title: E03_plate4_321


title: E01_plate4_305


title: D11_plate4_384


title: D10_plate4_376


title: F06_plate4_346


title: F02_plate4_314


title: D08_plate4_360


title: D09_plate4_368


title: F01_plate4_306


title: D06_plate4_344


title: D07_plate4_352


title: D05_plate4_336


title: D03_plate4_320


title: E05_plate4_337


title: E02_plate4_313


title: D02_plate4_312


title: D01_plate4_304


title: C10_plate4_375


title: C12_plate4_391


title: C09_plate4_367


title: C02_plate4_311


title: C07_plate4_351


title: B12_plate4_390


title: C06_plate4_343


title: C01_plate4_303


title: D04_plate4_328


title: C03_plate4_319


title: C04_plate4_327


title: B10_plate4_374


title: B09_plate4_366


title: B11_plate4_382


title: C08_plate4_359


title: B07_plate4_350


title: B04_plate4_326


title: B06_plate4_342


title: B08_plate4_358


title: C11_plate4_383


title: B05_plate4_334


title: B02_plate4_310


title: C05_plate4_335


title: A12_plate4_389


title: A10_plate4_373


title: A09_plate4_365


title: B03_plate4_318


title: B01_plate4_302


title: A08_plate4_357


title: A01_plate4_301


title: H10_plate3_280


title: A04_plate4_325


title: H12_plate3_296


title: H05_plate3_240


title: H04_plate3_232


title: A03_plate4_317


title: A11_plate4_381


title: A05_plate4_333


title: H03_plate3_224


title: A07_plate4_349


title: H02_plate3_216


title: A06_plate4_341


title: H06_plate3_248


title: G12_plate3_295


title: G11_plate3_287


title: H01_plate3_208


title: H11_plate3_288


title: H08_plate3_264


title: H09_plate3_272


title: G10_plate3_279


title: G09_plate3_271


title: G08_plate3_263


title: G06_plate3_247


title: G07_plate3_255


title: G05_plate3_239


title: A02_plate4_309


title: H07_plate3_256


title: G03_plate3_223


title: G04_plate3_231


title: F11_plate3_286


title: G01_plate3_207


title: F10_plate3_278


title: G02_plate3_215


title: F09_plate3_270


title: F07_plate3_254


title: F08_plate3_262


title: E11_plate3_285


title: F04_plate3_230


title: F06_plate3_246


title: E09_plate3_269


title: F05_plate3_238


title: F03_plate3_222


title: E12_plate3_293


title: F02_plate3_214


title: F01_plate3_206


title: E05_plate3_237


title: E08_plate3_261


title: E07_plate3_253


title: E10_plate3_277


title: E06_plate3_245


title: E04_plate3_229


title: E02_plate3_213


title: E03_plate3_221


title: E01_plate3_205


title: F12_plate3_294


title: D12_plate3_292


title: D11_plate3_284


title: D10_plate3_276


title: D06_plate3_244


title: D07_plate3_252


title: D02_plate3_212


title: D04_plate3_228


title: D03_plate3_220


title: D08_plate3_260


title: C11_plate3_283


title: D09_plate3_268


title: C12_plate3_291


title: C07_plate3_251


title: D01_plate3_204


title: C06_plate3_243


title: C09_plate3_267


title: C10_plate3_275


title: C02_plate3_211


title: B12_plate3_290


title: B10_plate3_274


title: B11_plate3_282


title: C01_plate3_203


title: C03_plate3_219


title: C05_plate3_235


title: C04_plate3_227


title: B09_plate3_266


title: B08_plate3_258


title: B07_plate3_250


title: B05_plate3_234


title: B01_plate3_202


title: B04_plate3_226


title: D05_plate3_236


title: B06_plate3_242


title: A11_plate3_281


title: A12_plate3_289


title: B02_plate3_210


title: A10_plate3_273


title: C08_plate3_259


title: B03_plate3_218


title: A08_plate3_257


title: A07_plate3_249


title: A04_plate3_225


title: A09_plate3_265


title: A06_plate3_241


title: A03_plate3_217


title: H12_plate2_196


title: H11_plate2_188


title: H10_plate2_180


title: A01_plate3_201


title: A02_plate3_209


title: H09_plate2_172


title: H08_plate2_164


title: H06_plate2_148


title: H07_plate2_156


title: H04_plate2_132


title: H02_plate2_116


title: H05_plate2_140


title: H01_plate2_108


title: A05_plate3_233


title: G12_plate2_195


title: G11_plate2_187


title: G04_plate2_131


title: G10_plate2_179


title: G06_plate2_147


title: G07_plate2_155


title: G05_plate2_139


title: G09_plate2_171


title: G02_plate2_115


title: G08_plate2_163


title: G01_plate2_107


title: F12_plate2_194


title: F11_plate2_186


title: F10_plate2_178


title: H03_plate2_124


title: F04_plate2_130


title: F06_plate2_146


title: F07_plate2_154


title: F05_plate2_138


title: F09_plate2_170


title: F08_plate2_162


title: F02_plate2_114


title: E12_plate2_193


title: E11_plate2_185


title: F03_plate2_122


title: E10_plate2_177


title: F01_plate2_106


title: G03_plate2_123


title: E09_plate2_169


title: E08_plate2_161


title: E06_plate2_145


title: E05_plate2_137


title: E07_plate2_153


title: E04_plate2_129