title: S4.2_NS

city: tianjin

first: Zhongai

last: Gao

address: qixiangtailu

institute: Tianjin Medical University

description: Gene expression in urine supernatant after centrifuge at 2000g for 10 minutes

hyb_protocol: 1.5 ug of Cy3-labelled cRNA (specific activity >10.0 pmol Cy3/ug cRNA) was fragmented at 60°C for 30 minutes in a reaction volume of 250 ml containing 1x Agilent fragmentation buffer and 2x Agilent blocking agent following the manufacturers instructions. On completion of the fragmentation reaction, 250 ml of 2x Agilent hybridization buffer was added to the fragmentation mixture and hybridized to Agilent Whole Human Genome Oligo Microarrays (G4112A) for 17 hours at 65°C in a rotating Agilent hybridization oven. After hybridization, microarrays were washed 1 minute at room temperature with GE Wash Buffer 1 (Agilent) and 1 minute with 37°C GE Wash buffer 2 (Agilent), then dried immediately by brief centrifugation.

scan_protocol: Slides were scanned immediately after washing on the Agilent DNA Microarray Scanner (G2505B) using one color scan setting for 1x44k array slides (Scan Area 61x21.6 mm, Scan resolution 10um, Dye channel is set to Green and Green PMT is set to 100%).

data_processing: The scanned images were analyzed with Feature Extraction Software 9.1 (Agilent) using default parameters to obtain background subtracted and spatially detrended Processed Signal intensities. Features flagged in Feature Extraction as Feature Non-uniform outliers were excluded.

source_name: S4, urine

label_protocol: Cyanine-3 (Cy3) labeled cRNA was prepared from 0.5 ug RNA using the One-Color Low RNA Input Linear Amplification PLUS kit (Agilent) according to the manufacturer's instructions, followed by RNAeasy column purification (QIAGEN, Valencia, CA). Dye incorporation and cRNA yield were checked with the NanoDrop ND-1000 Spectrophotometer.

extract_protocol: RNA was prepared using the Qiagen serum/Plasma Kit- Qiagen (Germany) following the manufacturer's recommendations. RNA was quantified using a NanoDrop-1000 spectrophotometer and quality was monitored with the Agilent 2100 Bioanalyzer (Agilent Technologies, Santa Clara, CA).

treatment_protocol: We collected urine samples in populations who underwent annual physical examination. The urine samples were centrifuged at 2000g for ten minutes in order to remove cells and debris, and then stored in -80 degree refrigerator for later use .

tag: disease state

value: diabetic kidney disease

tag: rna source

value: urine supernatant

value: female


title: Control N2 animals, Independent Trial 2

status: Public on Jun 05 2020

type: RNA

city: Murcia

first: Fernando

last: Gandía-Herrero

state: Murcia

address: Universidad de Murcia, Campus Espinardo - Edificio Veterinaria

department: Biochemistry and Moleular Biology

country: Spain

institute: University of Murcia

description: Gene expression data from control (non treated) animals

hyb_protocol: 3.75 µg of fragmented and biotinylated ss-cDNA were included in the hybridization mix, using the GenAtlas Hybridization, Wash and Stain kit for WT Array Strips (Affymetrix, P/N 901667) according to the recommendations of the manufacturer. The resulting preparations were hybridized to GeneChip® C. elegans Gene 1.1 ST Array Strip (Affymetrix, 902157) with 26 unique sequences of each transcript. After applying hybridization (spike controls) and labeling tests it was observed that the 20 chips had fulfilled the quality criteria.

scan_protocol: GeneAtlas Microarray System from Affymetrix

data_processing:
After scanning, microarrays data were processed using Affymetrix Expression Command Console (Affymetrix) and all samples were within bounds for hybridization and labeling tests. Three independent RNA samples were employed. Samples from worms treated with betalains were grouped as “treatment” and worms without betalains exposition were grouped as “control”. Data analysis was then performed with RMA (Robust Multiarray Average) allowing raw intensity values to be background corrected, log2 transformed and then quantile normalized in order to obtain an individual intensity value for each probe set.
log2 transformed and then quantile normalized in order to obtain an individual intensity value for each probe set.

label: BIOTIN-STREPTAVIDIN

molecule: total RNA

organism: Caenorhabditis elegans

source_name: C. elegans Control

label_protocol: Ss-cDNA was synthetized from 3.5 ng of each sample using the GeneChip WT Pico Reagent kit (Affymetrix, P/N 703262). The amount and quality of ss-cDNA were checked by Nanodrop and Bioanalyzer. ss-cDNA targets were cleaned up, and after fragmentation and terminal labelling with biotin

growth_protocol: N2 worm were synchronized using the bleaching protocol, from adultworms maintained in standard NGM agar plates at 20 ºC

extract_protocol: TRIzol extraction of total RNA was performed according to the manufacturer's instructions, followed by a purification using PureLink TM RNA Mini Kit from Invitrogen

treatment_protocol: 16-18 hours after haching L1 larvae were treated with 25 uM of each betalain in S. medium for 48 hours at 20 ºC

tag: tissue

value: Whole animal

tag: genotype

value: Wild-type N2

tag: age

value: 3 days after hatching, young adults

tag: treatment

value: control (non treated)


title: Control N2 animals, Independent Trial 1


title: PCNSL33

city: Kyoto

first: Ryuya

last: Yamanaka

address: 465 Kajii-cho, Kamigyoku

country: Japan

institute: Kyoto Prefectural University of Medicine

description: PCNSL33_AF_9463_(miRNA-4_0).rma-dabg-Signal

hyb_protocol: Following fragmentation, 1 ug of cRNA were hybridized for 17hr at 45C on GeneChip miRNA 4.0 Array. GeneChips were washed and stained in the Affymetrix Fluidics Station 450.

scan_protocol: GeneChips were scanned using the Hewlett-Packard GeneArray Scanner 3000.

data_processing: The data were analyzed with Affymetrix Expression Console™  using Affymetrix default analysis settings and global scaling as normalization method. The trimmed mean target intensity of each array was arbitrarily set to 100.

label: biotin

organism: Homo sapiens

source_name: PCNSL tissue

label_protocol: Biotinylated cRNA were prepared according to the standard Affymetrix protocol from 1 ug total RNA (Expression Analysis Technical Manual, 2001, Affymetrix).

extract_protocol: Isogen extraction of total RNA was performed according to the manufacturer's instructions.

tag: subject age

tag: gender

value: Male

value: primary central nervous system lymphomas (PCNSL)

tag: molecule subtype

value: miRNA


title: PCNSL36

description: PCNSL36_AF_9463_(miRNA-4_0).rma-dabg-Signal

value: Female


title: Indicaxanthin treated N2 animals, Independent Trial 1

description: Gene expression data from indicaxanthin treated animals

source_name: C. elegans treated with indicaxanthin (25 uM)

value: indicaxanthin


title: D21 uninjured A10

type: SRA

city: Los Angeles

first: Yerbol

middle: Z.

last: Kurmangaliyev

state: CA

address: 675 Charles E. Young Drive South

country: USA

institute: University of California, Los Angeles

description: rpkm.txt

library_source: transcriptomic

data_processing:
RNA sequencing reads were aligned to the mouse reference genome (mm10) using STAR (Dobin et al., 2013).
Raw expression levels were quantified for all annotated genes (Ensembl 86).
Raw expression levels were normalized to RPKM values using edgeR (Robinson et al., 2010)
Genome_build: mm10
Supplementary_files_format_and_content: tab-delimited text file include RPKM values for all samples

organism: Mus musculus

source_name: heart

extract_protocol:
Injured and uninjured regions of the heart were harvested at different time points following injury
Total RNA extracted using RNeasy Mini kit (Qiagen); RNA-Seq libraries were generated using TrueSeq RNA Library Prep Kit (Illumina)

tag: strain

value: cardiac muscle

value: uninjured

tag: time point

value: day 21 after injury


title: PCNSL32

description: PCNSL32_AF_9463_(miRNA-4_0).rma-dabg-Signal


title: Old

city: Singapore

first: Brijesh

middle: Kumar

last: Singh

address: CVMD Level-8, 8 College Road

department: Cardiovascular and Metabolic Disorders Program

country: Singapore

institute: Duke-NUS Gradiate Medical School

data_processing:
Raw data processing and quality assessment: Raw image data file from high-throughput sequencing(like Illumina ) was transformed to Sequenced Reads(called Raw Data or Raw Reads) by CASAVA base recognition (Base Calling). Raw Data is stored in FASTQ(fq) format files, which contain reads sequence and corresponding base quality
Error rate distribution:"e" represents sequencing error rate, "Qphred" represents base quality values of Illunima, the Qphred=-10log10(e).
GC-content:For RNA-seq, in view of the random interrupted and the principle of G/C, A/T content are respectively equal, G and C, A and T should be equal and content is stable on the whole through the whole sequencing process for non-stranded library (If the library is strand-specific, it may occur AT separation or GC separation). A large variation of sequencing error in the first 6-7 bases is allowed considering the usage of random primer in library construction. It's normal that the first few bases have certain preference in existing high-throughput sequencing technology, for the usage of 6bp random primer in library construction.
Data filtering:The Sequenced Reads/raw reads often contain low quality reads or reads with adaptors. As we can see in Fig 1.3, which will affect the further analysis results. In order to avoid this, it's necessary to filter the raw reads under conditions to get the clean reads. Raw reads filtering conditions are as follows:  (1) Remove reads containing adaptors;  (2) Remove reads containing N > 10% (N represents base that could not be determined);  (3) The Qscore (Quality value) of over 50% bases of the read is <= 5.  RNA-seq Adapter information:  NEBNext® UltraTM RNA Library Prep Kit  RNA 5' Adapter(5'Adaptor)  5' AATGATACGGCGACCACCGAGATCTACACTCTTTCCCTACACGACGCTCTTCCGATCT  RNA 3' Adapter(3'Adaptor,The underlined 6bp bases is Index)  5' GATCGGAAGAGCACACGTCTGAACTCCAGTCACATCACGATCTCGTATGCCGTCTTCTGCTTG
Mapping reads to reference genome: Mapping the clean reads to the reference genome or the transcriptome is the basis for the next following analysis. Novogene use STAR to accomplish the mapping. For the spieces(Human and mice) whose gene annotation information are more completed, if only need the differential expression significant genes, can map reads to the transcriptome directly. If research the study of alternative splicing, variation, the fusion gene, the alignment should be carried out at the genome level. In order to avoid losing a lot of effective junction reads, Novogene use STAR software to alignment for RNA-seq sequencing data analysis, the junction reads can be positioned precisely.
Reads distribution on genome: Reads will be mapped to exon, intron and intergenic.According to the mapping result, summary the percentage of statistics for reads in the genome exon, intron and intergenic region seperately. In general, the gene annotation of the species is more complete (such as mouse，human), the ratio of reads mapped to exon is supposed to be the highest. Reads mapped to intron may result from the remaining pre-mRNA or alternative splicing event. Reads mapped to intergenic may due to some ncRNA or DNA fragment contamination or the incomplete gene annotation. After getting the information about distribution of mapped reads on genome area, we will focus the distribution of mapped reads on chromosome. According to the mapping result, the 1M base is used as the sliding window, and the readings covered in the statistical window are drawn, and the density distribution map of the reads in each chromosome of the genome is plotted
Visualization of mapping results: We provide mapping result in the format of BAM together with reference genome and annotation file. BAM file can be visualized by using IGV (Integrative Genomics Viewer). The Integrative Genomics Viewer (IGV) is a high-performance visualization tool for interactive exploration of large, integrated genomic datasets. It supports a wide variety of data types, including array-based and next-generation sequence data, and genomic annotations. By using IGV we can (1)visualize reads position on genome, including reads distribution on chromosomes, the distribution of exon, intron, junction, intergenic and so on at different scale (2) visualize reads density in different area at different scale (3) visualize annotation of gene and its isoform(4)visualize other annotation information.
Gene expression level analysis: The abundance of transcript reflects gene expression level directly. In RNA-seq experiment, we estimate gene expression level by the abundance of transcripts(count of sequencing) that mapped to genome or exon. Reads count is proportional to be proportional to gene expression level, gene length and sequencing depth. FPKM (short for the expected number of Fragments Per Kilobase of transcript sequence per Millions base pairs sequenced) is the most common method of estimating gene expression levels, which takes the effects into consideration of both sequencing depth and gene length on counting of fragments.(Mortazavi et al. 2008)
Differential expression analysis: Readcount obtained from Gene Expression Analysis is used to do differential expression analysis. For the samples with biological replicates, differential expression analysis of two conditions/groups was performed using the DESeq2 R package (Anders et al., 2010). It provides statistical routines for determining differential expression in digital gene expression data using a model based on the negative binomial distribution. So, if the readcount of the i-th gene in j-th sample is Kij, there is: Kij ～ NB(μij,σij2) And the resulting P values were adjusted using the Benjamini and Hochberg's approach for controlling the false discovery rate. Kij ～ NB(μij,σij2), For the samples without biological replicates, readcount was adjusted by TMM, then differential expression analysis was performed by using the EdgeR R package
Genome_build: mm10
Supplementary_files_format_and_content: tab-delimited text files include FPKM values for each Sample ...

source_name: AML12 cells

growth_protocol: AML12 cells were maintained in DMED:F12 nutrient mixture supplemented with 10% FBS and

extract_protocol:
Total RNA was prepared from AML12 cells using InviTrap Spin Cell RNA Mini Kit (Stratec) according to the manufacturer's instructions. Quality control was performed with Agilent Bioanalyser.
mRNA from Eukaryote organisms is purified from total RNA using poly-T oligo-attached magnetic beads (For prokaryotes, mRNA was purified through the removal of rRNA by using kit). The mRNA is first fragmented randomly by addition of fragmentation buffer. Then first strand cDNA is synthesized using random hexamer primer and M-MuLV Reverse Transcriptase (RNase H-). Second strand cDNA synthesis is subsequently performed using DNA Polymerase I and RNase H. Double-stranded cDNA is purified using AMPure XP beads. Remaining overhangs of the purified double-stranded cDNA are converted into blunt ends via exonuclease/polymerase activities. After adenylation of 3' ends of DNA fragments, NEBNext Adaptor with hairpin loop structure is ligated to prepare for hybridization(1). In order to select cDNA fragments of preferentially 150~200 bp in length, the library fragments are purified with AMPure XP system (Beckman Coulter, Beverly, USA). Finally, the final library is gotten by PCR amplification and purification of PCR products by AMPure XP beads.

treatment_protocol: AML12 cells were treated with hydrogen peroxide to induce premature senescence and used as Old cells. Young are untreated control cells.

value: Liver

value: 3 months


title: S4.1_NS

country: China

value: diabetic kidney disease

value: male


title: D21 injured A4

description: rpkm.txt

value: injured


title: Young


title: S3.1_NS

source_name: S3, urine

value: diabetes

value: urine supernatant


title: PCNSL28

description: PCNSL28._AF_9463_(miRNA-4_0).rma-dabg-Signal


title: PCNSL29

description: PCNSL29_AF_9463_(miRNA-4_0).rma-dabg-Signal


title: S3.2_NS

value: female


title: PCNSL30

description: PCNSL30_AF_9463_(miRNA-4_0).rma-dabg-Signal


title: D21 uninjured A3

description: rpkm.txt


title: S2.2_NS

source_name: S2, urine

value: IGT with renal impairment


title: D21 injured A3

description: rpkm.txt


title: skin_7 day 2

city: Bethesda

first: Junji

last: Xu

address: 9000 Rockville Pike

institute: NIH, NIDCR

data_processing:
The reads were mapped to ensemble mouse reference transcriptome GRCm38v79 with kallisto v0.43.
Tab-delimited text file is summarized as transcript per million (TPM) or raw counts with tximport R package
Supplementary_files_format_and_content: tab-delimited text files include TPM values for each Sample

source_name: skin

extract_protocol:
Skin was cleaned of muscle and fat tissue, flash frozen on dry ice, and RNA was harvested using total RNA was extracted using the RNeasy mini kits (Qiagen), purified using Direct-zol RNA MiniPrep kit (Zymo Research), and was used with 1 ug of total RNA for the construction of sequencing libraries.
Next-generation libraries were prepared using the VAHTS TM mRNA-seq V2 Library Prep Kit for Illumina (Vazyme, #NR601)

tag: stress

value: wound

value: skin

tag: time

value: 7 days


title: skin_7 day 1


title: skin_5 day 2

value: 5 days


title: D14 uninjured A11

description: rpkm.txt

value: day14 after injury


title: skin_5 day 1


title: D21 uninjured A4

description: rpkm.txt

value: cardiac muscle


title: scATAC_Spata7_KO_6M

city: Houston

first: sangbae

last: Kim

address: 1 baylor plaza

institute: Baylor College of Medicine

library_source: genomic

data_processing:
After libraries were sequenced, the data from both reagent chemistries were analyzed using the  Cell Ranger ATAC pipeline v1.1.0 (10x Genomics, https://support.10xgenomics.com/single-cell-atac).
We processed scATAC-seq data using the Cell ranger ATAC pipeline. Briefly, this included barcode  processing, alignment, duplicate marking, peak calling, cell calling, dimensionality reduction,  clustering, t-SNE projection, peak annotation, and differential accessibility analyses. Using the Cell  Ranger ATAC pipeline we performed data aggregation and generated post-normalization  unique fragments for wild-type and mutants.
Open chromatin regions in a genome-wide manner were determined by peak calling using the ZINBA-line mixture model in the Cell Ranger ATAC pipeline. Next, a peak-barcode (cell) matrix was generated consisting of the counts of fragment ends within each peak region for each cell for downstream analysis such as dimensional reduction, clustering and visualization.
To find differentially accessible motifs between groups of cells, Cell Ranger ATAC uses a Negative Binomial generalized linear model to discover the cluster specific means and their standard deviations and then employs a Wald test for inference.
Genome_build: mm10
Supplementary_files_format_and_content: The files inclduing raw peak barcode matrix in hdf5 format and  annotation of peaks in tsv format for each sample condition

molecule: genomic DNA

growth_protocol: For all experiments, male Spata7 (Eblimit et al., 2015) mutant mice were compared with age-matched male C57BL6/J mice, to avoid sex as a biological variable.

extract_protocol:
The 10x Chromium Demonstrated Protocol for Nuclei Isolation for Single Cell ATAC Sequencing (10x genomics, Document CG000169) was followed to isolate nuclei.
Libraries were constructed with Single Cell ATAC reagent and chip kits following the procedure in the user guides for the Chromium Single Cell ATAC Library & Gel Bead Kit and the Chromium Chip E Single  Cell ATAC Kit (10x genomics, PN-1000110 and PN-1000084). Single cell ATAC library was quantified using a KAPA DNA Quantification kit (KAPABiosystems- KK4960) and sequenced with 1% PhiX on Illumin Novaseq with the following parameter: read1: 50bp, index1: 8bp, index2: 16bp, read2: 49bp.

treatment_protocol: Mice for all the time points were housed on a standard diet and in a 12 -hr light to 12 -hr dark cycle.

tag: Sex

value: Male

value: Spata7 mutant (Eblimit et al., 2015, PMID:25398945)

value: Retina

value: 6 month


title: D14 injured A11

description: rpkm.txt


title: skin_5 day 3


title: scATAC_Spata7_WT_7M

value: 7 month


title: skin_3 day 3

value: 3 days


title: S1.1_NS

source_name: S1, urine

value: control

value: male


title: CLjuzhi4

city: shanghai

first: bin

last: sun

address: zhizaojulu

institute: Shanghai Jiaotong University School of Medicine

data_processing:
Illumina Casava1.7 software used for basecalling.
Sequenced reads were trimmed for adaptor sequence, and masked for low-complexity or low-quality sequence, then mapped to hg38 whole genome using Hisat2 version 2.1.0 with default parameters
Gene Ontology (GO) analysis was performed with R package. Top GO categories were selected according to the P values of expression changes.
Genome_build: hg38
Supplementary_files_format_and_content: tab-delimited text files include FPKM values for each Sample ...

source_name: mac-ADSCs

extract_protocol:
ADSCs were removed, flash frozen on dry ice, and RNA was harvested using Trizol reagent. Illumina TruSeq RNA Sample Prep Kit (Cat#FC-122-1001) was used with 1 ug of total RNA for the construction of sequencing libraries
RNA libraries were prepared for sequencing using standard Illumina protocols

tag: cell type

value: adipose derived stem cell of macrodactyly + BYL -719


title: CLjuzhi5


title: skin_3 day 2


title: PCNSL31

description: PCNSL31_AF_9463_(miRNA-4_0).rma-dabg-Signal

value: Female

value: primary central nervous system lymphomas (PCNSL)

value: miRNA


title: S1.2_NS

value: urine supernatant

value: female


title: skin_7 day 3

value: wound

value: skin

value: 7 days


title: skin_1 day 3

value: 1 day


title: CLjuzhi1


title: D14 uninjured A6

description: rpkm.txt

value: cardiac muscle

value: uninjured

value: day14 after injury


title: D14 injured A6

description: rpkm.txt

value: injured


title: S2.1_NS

value: IGT with renal impairment


title: GXBduozhi4

source_name: pol-ADSCs

value: adipose derived stem cell from polydactyly


title: skin_3 day 1

value: 3 days


title: skin_1 day 1


title: skin_0 day 3

value: 0 day


title: D14 uninjured A5

description: rpkm.txt


title: D14 injured A5

description: rpkm.txt


title: GXBduozhi3


title: skin_0 day 1


title: D14 uninjured A4

description: rpkm.txt


title: GXBduozhi1


title: GXBjuzhi4

value: adipose derived stem cell of macrodactyly


title: D7 uninjured A12

description: rpkm.txt

value: day 7 after injury


title: GXBjuzhi1


title: GXBjuzhi3


title: skin_1 day 2


title: skin_0 day 2


title: D7 uninjured A10

description: rpkm.txt


title: D7 injured A12

description: rpkm.txt


title: Muscle6

city: Chengdu

first: Liming

last: Chang

address: No.9 Section 4,Renmin Nan Road,Chengdu, Sichuan, P.R. China

department: Chengdu Institute of Biology

institute: Chinese Academy of Sciences

data_processing:
Illumina Casava1.7 software used for basecalling.
Sequenced reads were trimmed for adaptor sequence, and masked for low-complexity or low-quality sequence;The total clean reads from all libraries were assembled using Trinity as a reference transcriptome.
The expression level of each unigene was expressed as fragments per kilobase of exon model per million mapped reads (FPKM).
Genome_build: de novo
Supplementary_files_format_and_content: XLSX file includes FPKM values for all Samples
Supplementary_files_format_and_content: fasta file includes the sequences of all unigenes

organism: Pelophylax nigromaculatus

source_name: muscle

extract_protocol:
Total RNA of each sample was extracted and purified using Trizol (Invitrogen, Carlsbad, CA, USA) following the manufacturer’s instructions. After purified with poly-T oligo-attached magnetic beads, the mRNAs were fragmented.
First-strand cDNA was synthesized using random hexamer primers and M-MuLV Reverse Transcriptase (RNase H−). Second-strand cDNA synthesis was subsequently performed using DNA Polymerase I and RNase H. The remaining overhangs were converted into blunt ends via exonuclease/polymerase activities. After adenylation of the 3′ ends of the DNA fragments, NEBNext Adaptors with a hairpin loop structure were ligated to prepare for hybridization. To preferentially select cDNA fragments of 150–200 bp in length, the library fragments were purified with AMPure XP system (Beckman Coulter, Beverly, USA). Then, 3 μl USER Enzyme (NEB, USA) was used with size-selected, adaptor-ligated cDNA at 37 °C for 15 min followed by 5 min at 95 °C before PCR. Then, PCR was performed with a Phusion High-Fidelity DNA polymerase, universal PCR primers and Index (X) primer. Finally, PCR products were purified (AMPure XP system), and library quality was assessed on the Agilent Bioanalyzer 2100 system. The clustering of the index-coded samples was performed on a cBot Cluster Generation System using the TruSeq PE Cluster Kit v3-cBot-HS (Illumina) according to the manufacturer’s instructions. After cluster generation, the library preparations were sequenced on an Illumina HiSeq 2500 platform by NovoGene (Beijing), and paired-end reads were generated.

tag: developmental stage

value: adult

value: muscle

tag: disease

value: torticollis


title: Brain6

source_name: brain

value: brain


title: Muscle5


title: D7 uninjured A9

description: rpkm.txt


title: Liver6

source_name: liver

value: liver


title: D7 injured A10

description: rpkm.txt


title: ∆metr-1_nhr114_RNAi_rep2

city: Worcestor

first: Gabrielle

last: Giese

state: MA

address: 368 Plantation St.

department: Program in Systems Biology

institute: University of Massachusetts Medical School

description: grown on NGM + 20 nM vitamin B12 + E. coli HT115 nhr-114 RNAi

data_processing:
Homemade python script was used for demultiplexing
Sequenced reads were trimmed using trimmomatic-0.32
Trimmed reads were aligned to the reference genome using STAR with the following parameters: —runThreadN 4 --alignIntronMax 25000 --outFilterIntronMotifs RemoveNoncanonicalUnannotated
Features were counted by ESAT (Derr et al Genome Res. 2016) with parameter “-task score3p -wLen 100 -wOlap 50 -wExt 1000 -sigTest .01 -multimap normal -scPrep -umiMin 1”. Features in “c_elegans.PRJNA13758.WS271.canonical_geneset.gtf” were used as the annotation table input for ESAT, but pseudogenes were discard
Genome_build: c_elegans.PRJNA13758 (WormBase WS271)
Supplementary_files_format_and_content: tab dilimeted file containing TPM matrix for all samples and all genes

source_name: whole animal

growth_protocol: Animals were maintained on NGM-agar with soy peptone and 0.64 nM vitamin B12 prior to experiment. They were synchronized by an 18 hour L1 arrest post-buffered bleach treatment.

extract_protocol:
A minumum of 200 staged animals were collected and washed thrice in M9 buffer. Total RNA was extracted via Trizol and BCP followed by Dnase I treatment. Zymo Direct-zol RNA miniprep kit was used for clean up.
A modified Celseq2 protocol was used.

treatment_protocol: Experment 1: Synchronized animals were grown on NGM-agar containing soy peptone plus or minus methionie. Experiment 2: Synchronized animals were grown on NGM-agar plus IPTG and Amp and seeded with E. coli HT115 containing either vector or nhr-114 RNAi.

value: whole animal

tag: Stage


title: metr-1(ww52)_nhr114_RNAi_rep1


title: metr-1(ww52)_vector_rep1

description: grown on NGM + 20 nM vitamin B12 + E. coli HT115 vector RNAi


title: metr-1(ww52)_nhr114_RNAi_rep2


title: Brain5


title: D14 injured A4

description: rpkm.txt


title: 3T3-L1 Cells_24hr_X-1_rep1

city: Shanghai

first: jun

last: Han

address: 800 Xiangyin Rd.

institute: Second Military Medical University, Shanghai, China

description: Gene expression after 24hr in X-1-treated 3T3-L1 preadipocytes

hyb_protocol: 875 ng of Cy3-labelled cRNA (specific activity >10.0 pmol Cy3/ug cRNA) was fragmented at 60°C for 30 minutes in a reaction volume of 55 μl containing 25x Agilent fragmentation buffer and 10 x Agilent blocking agent following the manufacturers instructions. On completion of the fragmentation reaction, 55 μl of 2x Agilent hybridization buffer was added to the fragmentation mixture and hybridized to Agilent Whole Mouse Genome Oligo Microarrays (4×44K) for 17 hours at 65°C in a rotating Agilent hybridization oven.  After hybridization, microarrays were washed 1 minute at room temperature with GE Wash Buffer 1 (Agilent) and 1 minute with 37°C GE Wash buffer 2 (Agilent), then dried immediately by brief centrifugation.

scan_protocol: Slides were scanned immediately after washing on the Agilent DNA Microarray Scanner (G2565BA) using one color scan setting for 4x44k array slides (Scan resolution 5um, Dye channel is set to Green, Green PMT is set to 100% and 10%,one scan each time).

data_processing: The scanned images were analyzed with Feature Extraction Software 9.1 (Agilent) using default parameters (protocol GE1-v1_91 and Grid: 014868_D_F_20100617 or 014868_D_F_20090416) to obtain background subtracted and spatially detrended Processed Signal intensities.  Features flagged in Feature Extraction as Feature Non-uniform outliers were excluded.

source_name: 3T3-L1 cells, 24hr, 70 µM X-1

label_protocol: Cyanine-3 (Cy3) labeled cRNA was prepared from 0.5 ug RNA using the One-Color Low RNA Input Linear Amplification PLUS kit (Agilent) according to the manufacturer's instructions, followed by RNAeasy column purification (QIAGEN, Valencia, CA).  Dye incorporation and cRNA yield were checked with the NanoDrop ND-1000 Spectrophotometer.

extract_protocol: The total RNA was prepared using the RNeasy Mini kit (Qiagen) following the manufacturer's recommendations.  The protocol includes lysis of cells, and an on-column DNase digestion. RNA was quantified using a NanoDrop-1000 spectrophotometer and quality was monitored with the Agilent 2100 Bioanalyzer (Agilent Technologies, Santa Clara, CA).

treatment_protocol: 3T3-L1 cells. Cells were incubated with DMEM containing 10% (vol/vol) fetal bovine serum, and subsequently serum-starved 48h. X-1 or X-3 at 70 µM was added and incubated for further 24 h at 37 ºC in a humidified incubator with 5% CO2.

tag: cell line

tag: agent

tag: replicate


title: 3T3-L1 Cells_24hr_X-3_rep3

description: Gene expression after 24hr in X-3-treated 3T3-L1 preadipocytes

source_name: 3T3-L1 cells, 24hr, 70 µM X-3


title: ∆metr-1_nhr114_RNAi_rep1

tag: tissue

tag: genotype


title: 3T3-L1 Cells_24hr_X-3_rep1


title: metr-1(ww52)_vector_rep2


title: D7 uninjured A7

description: rpkm.txt

tag: treatment


title: 3T3-L1 Cells_24hr_X-3_rep2


title: 3T3-L1 Cells_24hr_control_rep3

description: Gene expression after 24hr in untreated 3T3-L1 preadipocytes

source_name: 3T3-L1 cells, 24hr, control

value: control


title: 3T3-L1 Cells_24hr_control_rep1


title: 3T3-L1 Cells_24hr_X-1_rep3


title: 3T3-L1 Cells_24hr_X-1_rep2


title: D7 injured A7

description: rpkm.txt

value: cardiac muscle


title: D7 injured A9

description: rpkm.txt


title: 3T3-L1 Cells_24hr_control_rep2


title: Liver5


title: Brain4


title: D3 injured A8

description: rpkm.txt

value: day 3 after injury


title: D3 uninjured A8

description: rpkm.txt

value: uninjured


title: K31: control 17

city: Homburg

first: Nicole

last: Ludwig

address: Kirrberger Strasse

department: Human Genetics

country: Germany

institute: Saarland University

description: miRNAs in embryonic culture medium

hyb_protocol: The expression profile of 2549 human miRNAs was determined using SurePrint G3 8x60k miRNA microarrays (Agilent) according to the manufacturers recommendations

scan_protocol: Array´s were scanned with the Agilent Scanner with default settings.

data_processing: Raw expression values were extracted using the Feature Extraction Software of Agilent. Raw signal intensities were quantil-normalized and log2 transformed.

source_name: conjunctival cells

label_protocol: miRNA Complete Labeling and Hyb Kit (Agilent, Cat. No 5190-0456)

extract_protocol: Total RNA including miRNAs was isolated using the Qiagen miRNeasy Kit according to manufacturers recommendations.

tag: age

tag: gender

value: W

value: healthy control

value: conjunctival cells


title: K36: control 19

description: miRNAs in embryonic culture medium


title: K32: control 18

description: miRNAs in embryonic culture medium

value: M


title: K5: control 20

description: miRNAs in embryonic culture medium


title: WT_nhr-114_RNAi_rep2

tag: strain


title: Liver4


title: WT_nhr-114_RNAi_rep1


title: Muscle4


title: ∆metr-1_vector_rep2


title: D3 uninjured A7

description: rpkm.txt

tag: time point


title: D3 injured A7

description: rpkm.txt

value: injured


title: K29: control 15

description: miRNAs in embryonic culture medium


title: K3: control 16

description: miRNAs in embryonic culture medium


title: WT_vector_rep2


title: WT_vector_rep1


title: ∆metr-1_vector_rep1


title: Liver3

value: healthy


title: K28: control 14

description: miRNAs in embryonic culture medium


title: Muscle3


title: D3 uninjured A5

description: rpkm.txt


title: ∆nhr-114_soy_Met_rep2

description: growrn on NGM(soy peptone) + 25 mM methionine + E. coli OP50


title: D3 injured A5

description: rpkm.txt


title: K27: control 13

description: miRNAs in embryonic culture medium


title: D3 uninjured A4

description: rpkm.txt


title: ∆nhr-114_soy_Met_rep1


title: Brain2


title: K21: control 11

description: miRNAs in embryonic culture medium


title: Muscle2


title: Brain3


title: ∆nhr-114_soy_rep2

description: grown on NGM(soy peptone) + E. coli OP50


title: ∆nhr-114_soy_rep1


title: D3 injured A4

description: rpkm.txt


title: Liver2


title: WT_soy_Met_rep2


title: PA-VMHvl180315

city: New York

first: Takashi

last: Yamaguchi

state: New York

address: 435 E 30th St

department: Neuroscience Institute

institute: New York University School of Medicine

data_processing:
The RNA-seq libraries were then prepared using Clontech SMARTer Stranded Total RNA-Seq Kit (cat. no. 635006) and purified using AMPure beads (Beckman Coulter). The quantity of the library was examined using Qubit RNA HS assay Kit (Thermo Fisher Scientific) and qPCR. The quality of the library was determined using Bioanalyzer (Agilent). Once the libraries were deemed high quality (free of adaptor dimers and in concentration >2 nM), we pooled eight samples equimolarly and sequenced them by Illumina HiSeq 2500 using high output mode to achieve greater depth of coverage. Read alignment was carried out using HISAT2 (https://github.com/infphilo/hisat2) against the mouse mm10 reference genome obtained from Ensembl database (http://ensembl.org/)[
The alignment results were then exported to the open-source software Featurecounts to determine the number of reads for each gene.
Genome_build: mm10
Supplementary_files_format_and_content: normalized counts per gene per sample

source_name: ventromedial hypothalamus ventro-lateral part- projecting neurons in posterior amygdala

extract_protocol:
coronal frozen sections of C57BL/6N male mice. MPN- and VMHvl projecting neurons labeled by green or red retrobeads in posterior amygdala and basomedial amygdala posterior part were microdissected using LCM. RNA extracted using picopure (ThermoFisher).
Clontech SMARTer Stranded Total RNA-Seq Kit

treatment_protocol: injecting red or green retrobeads into the posterior amygdala

value: 8-9 weeks old

value: male


title: Muscle1

value: adult

value: muscle


title: Brain1

value: brain


title: PA-VMHvl180313


title: K2: control 9

description: miRNAs in embryonic culture medium


title: PA-VMHvl180316


title: Liver1

value: liver


title: K22: control 12

description: miRNAs in embryonic culture medium


title: K19: control 8

description: miRNAs in embryonic culture medium


title: K20: control 10

description: miRNAs in embryonic culture medium


title: PA-MPN180316

source_name: medial preoptic nucleus-projecting neurons in posterior amygdala


title: Brain9

value: torticollis with cataracts


title: WT_soy_Met_rep1

value: whole animal


title: WT_soy_rep1


title: WT_soy_rep2


title: K18: control 7

description: miRNAs in embryonic culture medium


title: PA-MPN180315


title: K16: control 6

description: miRNAs in embryonic culture medium


title: PA-MPN180313


title: H3K27me3_IP_Sperm

city: Manchester

first: Angela

last: Simeone

address: 200 Hathersage Rd

country: United Kingdom

institute: Genomic Services QIAGEN

description:
ICeChIP, paired-ended
macs2_nodup_190_Alan3_H3K27me3_narrowpeaks.HMD.bed

data_processing:
Adatpert trimming (cutadapt)
duplicate marked with Picard(2.14 - markDupicates) and removed; removal of reads with quality below 20 (samtools view -q20)
BAM files sorted, unmapped reads and secondary alignment removed; properly paired reads were joined to have fragments information and stored as BEDPE
BED files extracted from BEDPE
Given the length of fragments, BED for each fragment group have been created (150 - 110 - 70)
Peak calling with MACS2 (-q 0.01) with default options for H3K4me3 in frog ,and H3K4me3 H3K27me3 in Human, with broad option in H3K27me3 frog.
HMD was estimated as reported in paper considering coverages in windows 50-bp wide across the genome.
Genome_build: Xenopus laevis 6.1
Genome_build: HG38
Supplementary_files_format_and_content: BED files for peaks, bigwig files for tracks

growth_protocol: no growth protocol

extract_protocol:
Preparation sperm and egg extract treated sperm chromatin for ChIP-seq: Chromatin fractionation and chromatin immunoprecipitation (ChIP) were performed as described before (Hisano et al., 2013 and Teperek et al., 2016) with slight modifications. Xenopus sperm were purified from their testes and permeabilised before ChIP. Sperm or egg extract treated sperm chromatin was resuspended in 50 l of Buffer 1 (0.3 M Sucrose, 15 mM Tris pH 7.5, 60 mM KCl, 15 mM NaCl, 5 mM MgCl2, 0.1 mM EGTA, 0.5 mM DTT) and added 50 l of Buffer 1 with detergent (Buffer 1 including 0.5% NP40 and 1% NaDOC). Samples were incubated for 10 min on ice. 100 l of MNase buffer (0.3 M Sucrose, 85 mM Tris, 3 mM MgCl2, 2　mM CaCl2, 2.5U of micrococcal nuclease: Roche 10107921001) was added in to each tube (0.5 million of cells per tube). Tubes were incubated at 37C for 30 min in pre-warmed water bath. Reaction was stopped by adding 2 l of 0.5 M EDTA pH8.0 in the same order as started. Tubes were vortexed and placed on ice for at least 5 min. Supernatant and pellet were separated by centrifugation at 13,000 rpm for 10 min at room temperature. Supernatant was transferred to new tube and stored on ice. For ICeChIP, semi-synthetic nucleosomes were spiked in as described (Grzybowski et al., 2015) before MNase digestion. Preparation Chromatin for embryo ChIP-seq: ChIP was performed as described previously (Gentsch and Smith, 2014) with the following modifications. Blastula (stage 7) embryos were generated by in vitro fertilization. For each ChIP experiment, 50 embryos were fixed in 2 mL of 1% Formaldehyde in 1x MMR for 25 min at room temperature, followed by 4 washes with 1 ml of 1xMMR and equilibration in 500 μl HEG solution (50 mM HEPES-KOH pH 7.5, 1 mM EDTA, 20% Glycerol) at 4°C, then excess buffer was removed and samples were frozen at −80°C. To extract chromatin, the samples were transferred to 2 ml tube or 15 ml Falcon tube and homogenized in 200-250 μl of sonication buffer (20 mM Tris-HCl pH 8.0, 70 mM KCl, 1 mM EDTA pH 8.0, 10% Glycerol, 5 mM DTT, 0.125% NP40, 1x complete protease inhibitors), by pipetting up and down in a 1 ml pipette tip. 250 μl of diagenode beads were transferred to diagenode falcon tube (Diagenode, C1020031). 750 μl of embryo lysates were added and diagnode falcons were transferred to the sonicator bath (diagnode bioruptor). Sonication is carried out in 30 cycles (with 30 s on/off cycles). Sonicated embryo extract was transferred into eppendorf tubes. Chromatin was collected by centrifugation for 5 min at top speed in tabletop centrifuge at 4°C. Chromatin extract was transferred to new tube and stored on ice.
ChIP-seq library preparation was performed using the TruSeq DNA kit (Illumina, FC-121-2001) according to the manufacturer’s protocol.

value: sperm

tag: chip antibody


title: K15: control 5

description: miRNAs in embryonic culture medium


title: K14: control 4

description: miRNAs in embryonic culture medium


title: Liver9


title: H3K4me3_IP_Sperm

description:
ICeChIP, paired-ended
macs2_nodup_190_Alan3_H3K4me3_narrowpeaks.HMD.bed


title: BMAp180316

source_name: basomedial amygdala posterior part


title: Brain8


title: K12: control 3

description: miRNAs in embryonic culture medium


title: BMAp180315


title: Extracted RNA 3-day old chicken IAHC cells

city: Utrecht

first: Catherine

last: Robin

address: Uppsalalaan 8

country: Netherlands

institute: Hubrecht Institute

data_processing:
CEL-seq were mapped with bwa using custom scripts as previously described (Grün et al., Nature Methods, 2014)
Genome_build: mouse: mm10; chicken: galGal4, with improved 3’UTR annotations (see Junker et al., Cell, 2014); zebrafish: Zv9, with improved 3’UTR annotations (see Junker et al., Cell, 2014); human: hg19
Supplementary_files_format_and_content: *.coutc.csv: Tab separated data file for each sequencing library. The first column lists the official gene symbol followed by the chromosome name, separated by a double underscore. Column names indicate the number of sections/primers, separated by an underscore. For 'ggcells2.coutc.csv' Tab, which belongs to the chicken IAHC sorted cells, the two last columns indicate the number of reads obtained for experiment#1 and experiment#2 (primer#95 and primer#96, respectively). The order of the columns/primers follows the description of the sample from the title.  For instance: "Extracted RNA 3-day old chicken embryo from posterior-to-anterior axis":  the column 1 = most posterior section while the column 96 = most anterior. This is the case for all the samples listed.

organism: Gallus gallus

source_name: sorted IAHC cells

growth_protocol: Zebrafish were maintained and propagated according to standard laboratory conditions. Kdrl:mcherry/cd41:eGFP reporter fish line was used for tomo-seq at both 28hpf and 40hpf. Fertilized chicken eggs (Bovans Brown) were incubated at 37±1°C in a humidified atmosphere until they reached the appropriate developmental stage. E3 and E4 chicken embryos were used for tomo-seq. C57BL/6 mouse strain was housed according to institutional guidelines, and procedures were performed in compliance with Standards for Care and Use of Laboratory Animals with approval from the Hubrecht Institute ethical review board. E10.5 and E11.5 mouse embryos were used for tomo-seq. Human embryos were obtained from Dr. Chuva de Sousa Lopes laboratory (LUMC, Leiden, Netherlands). The Medical Ethical Committee of the Leiden University Medical Center (P08.087) approved all procedures regarding the collection and use of human embryonic material.

extract_protocol:
Total RNA was isolated from individual cryosection using TRIzol reagent (Ambion), according to the manufacturer’s manual. RNA extraction, amplification and sequencing libraries were performed as described in {Junker et al., 2014}. Briefly, RNA samples were treated according to the CEL-seq protocol {Hashimshony, 2012 #36}. Illumina sequencing libraries were prepared with the TruSeq small RNA sample prep kit (Illumina).
Libraries were sequenced on an Illumina HiSeq2500 using 50bp paired end sequencing and Illumina Illumina NextSeq 500 using 75bp paired end sequencing.

value: bovans brown

value: embryonic cells

tag: tissue preparation

value: sorted IAHC cells

value: 3 days old


title: Muscle8


title: K11: control 2

description: miRNAs in embryonic culture medium

value: healthy control

value: conjunctival cells


title: BMAp180313


title: D21 injured A10

description: rpkm.txt

source_name: heart

value: cardiac muscle

value: injured

value: day 21 after injury


title: Extracted RNA 3-day old chicken anterior slice #1 from dorsal-to-ventral axis

source_name: transversal slice

value: embryo

value: transversal slice


title: Extracted RNA 4-day old chicken posterior slice #1 from dorsal-to-ventral axis

value: 4 days old


title: Brain7

value: adult

value: brain


title: Extracted RNA 3-day old chicken posterior slice #1 from dorsal-to-ventral axis


title: Liver8

value: liver

value: torticollis with cataracts


title: Muscle9

value: muscle


title: Extracted RNA 4-day old chicken  anterior slice #1 from dorsal-to-ventral axis

value: bovans brown


title: K10: control 1

description: miRNAs in embryonic culture medium

value: W


title: AN6: patient 36

description: miRNAs in embryonic culture medium

value: F

value: AAK

value: Severe


title: HR_3 (2)

city: Baltimore

first: Suvarna

last: Nadendla

state: MD

address: 670 W.Baltimore Street

department: Institute for Genome Sciences

institute: University of Maryland School of Medicine

description: same as GSM2597300

data_processing:
FASTQ files were mapped to their respective genomes using HISAT2 (Kim, Langmead et al. 2015) for Mus musculus and Bowtie (Langmead, Trapnell et al. 2009) for pneumococcal genomes of D39, TIGR4, and 6A10. Reads mapped to each respective genome were tabulated in Supplemental Table 1. Gene counts for all samples were then estimated using HTseq (Anders, Pyl et al. 2015). All sample counts for the mouse data were tabulated in Supplemental Table 2. For bacterial counts, core genes without paralogs were determined for D39, TIGR4, and 6A10 using PanOCT (Fouts, Brinkac et al. 2012). Counts were then tabularized for analysis based on core genes, using TIGR4 locus tags as identifiers in Supplemental Table 2. Counts tables were then imported into Rstudio for analyses and estimation of differentially expressed (DE) genes. For mouse DE gene estimation, infected/colonized tissues were compared to their respective uninfected tissue control samples using DEseq2 (Love, Huber et al. 2014) and filtered using an FDR cutoff of 0.05. For bacterial DE gene estimation, each infected tissue was compared to the nasopharyngeal colonization state, while accounting for strain differences within core genes using DEseq2 and filtered using an FDR cutoff of 0.05.
Genome_build: Streptococcus pneumoniae D39 (NC_008533.1), Streptococcus pneumoniae TIGR4 (NC_003028.3), Streptococcus pneumoniae 6A10 (CP053210), Mus musculus (GCA_000001635.5).

source_name: Heart

growth_protocol: For mouse infection, pneumococci were grown in Todd Hewitt Broth (THB) (Acumedia, Neogen) with 0.5% yeast extract (THY) at 37°C in 5% CO2 until reaching mid-exponential phase [Optical density, O.D.620nm = 0.5; ~1.0 x 108 colony forming units (CFU)/mL] and serially diluted in sterile phosphate buffered saline (PBS). Animal experiments were carried out using male and female mice 6 to 12-weeks of age. Wildtype C57BL/6 were supplied from The Jackson Labs (Sacramento, California) and housed at the University of Alabama at Birmingham Animal Facilities for at least 1 week prior to their use. All animal experiments were performed using protocols approved by the University of Alabama at Birmingham IACUC (protocols #21152 and #21231). For the in vitro biofilm and planktonic samples: planktonic mid-log phase (OD620nm = 0.5) TIGR4 grown in THB were used to seed continuous once-flow through biofilm reactors. Biofilms were allowed to grow for 48 hours prior to collection of bacteria. RNA was isolated from the planktonic seed cultures and their respective biofilms. In total, 3 independent biological samples were collected with planktonic and biofilm RNAs from each experiment paired.

extract_protocol:
For nasopharynx samples, euthanized mice were decapitated, exterior skin removed, brain and cerebral column excised, and the skull defleshed. The skull and oropharynx were subsequently crushed in RNA Protect Bacteria Reagent (Qiagen) and stored at -80°C. For blood, samples were obtained by exsanguination via retro-orbital bleed and mixed with RNAprotect Bacteria Reagent and stored at -80°C. For lungs, heart, and kidneys, organs were collected, rinsed with PBS, minced, and placed into RNAprotect Bacteria Reagent and stored at -80°C. On the day of RNA extractions, samples were thawed and spun down to discard the supernatant. Pellets were then incubated in 100 μL of lysis buffer (10 μL of mutanolysin, 20 μL of proteinase K, 30 μL of lysozyme, 40 μL of TE buffer) for 10 minutes. Pellets were mechanically disrupted in 600 μL RLT buffer (RNeasy Mini Kit) containing 1% beta-mercaptoethanol, using a motorized mortar and pestle for 30 seconds. Samples were then spun through a Qiashredder followed by addition of 590 μL of 80% ethanol, and RNA was captured on the RNeasy Mini Kit columns with DNase treatment on column, as per their protocol, followed by DNase treatment in solution. Extracted RNA was quantitated using a Bioanalyzer. Ribosomal RNA was depleted using the Ribo-Zero rRNA Removal Kit for Gram-positive bacteria and/or for human/mouse/rat. RNA was isolated from the planktonic seed cultures and their respective biofilms. In total, 3 independent biological samples were collected with planktonic and biofilm RNAs from each experiment paired. Total RNA was extracted from each replicate separately using enzymatic lysis of pneumococcal cells (10 μL mutanolysin, 20 μL proteinase K, 15 μL lysozyme, and 55 μL TE) followed by RNA extraction using the same protocol mentioned above.
300 bp-insert RNA-seq Illumina libraries were constructed using 0.01 – 2.0 μg of enriched mRNA that was fragmented then used for synthesis of strand-specific cDNA using the NEBNext Ultra Directional RNA Library Prep Kit for Illumina (NEB-E7420L). The cDNA was purified between enzymatic reactions and the size selection of the library performed with AMPure SpriSelect Beads (Beckman Coulter Genomics). The titer and size of the libraries was assessed on the LabChip GX (Perkin Elmer) and with the Library Quantification Kit (Kapa Biosciences).

treatment_protocol: Mice were anesthetized using vaporized isoflurane at 2.5% during experimental challenge. For asymptomatic nasopharyngeal colonization mice were inoculated with 105 CFU of Spn in 10 µl saline with samples collected on day 7 (Riegler, Brissac et al. 2019). For pneumonia, mice were challenged with 105 CFU of Spn in 100 µl saline by forced aspiration with lungs collected on day 2 (Gonzalez-Juarbe, Bradley et al. 2018). For invasive disease, mice were challenged intraperitoneally with 104 CFU of Spn in 100 µl saline with blood, heart, and kidneys collected between 30-36h post-challenge. Data generated from TIGR4 infected blood and heart samples were from a previously published study by our group, with mice infected in same fashion (Shenoy, Brissac et al. 2017).

value: Heart


title: AN48: patient 40

description: miRNAs in embryonic culture medium

value: conjunctival cells


title: HR_2 (2)

description: same as GSM2597299


title: AN49: patient 32

description: miRNAs in embryonic culture medium

value: Mild


title: Extracted RNA 3-day old chicken sub-dissected aorta from posterior-to-anterior axis

source_name: sub-dissected aorta

value: sub-dissected aorta

value: 3 days old


title: Liver7


title: HR_1 (2)

description: same as GSM2597298


title: AN46: patient 33

description: miRNAs in embryonic culture medium

value: M


title: Extracted RNA 3-day old chicken embryo from posterior-to-anterior axis

source_name: whole embryo

value: embryo

value: whole embryo


title: AN47: patient 5

description: miRNAs in embryonic culture medium


title: Muscle7

source_name: muscle

value: adult


title: ∆tri6/TRI6-NPN-BR2

city: Ottawa

first: Kristina

last: Shostak

address: 960 Carling Avenue

country: Canada

institute: Agriculture and Agri-Food Canada

data_processing:
Raw read QC and trimming (CLC Genomics V12). Parameters: ambiguous limit 2, quality limit 0.05, min number of nucleotides in reads 20, discard short reads.
RNAseq Analysis function in CLC Genomics V12. Parameters: one reference sequence per transcript, no spike-in controls, mismatch cost 2, indel cost 3, length fraction 0.9, similarity fraction 0.8, auto-detect paired distances, strand specific - both, max number of hits for a read 10, do not count paired reads as two, expression value in unique counts.
DESeq2 to normalize unique gene read counts and to obtain differential expression.
Genome_build: The reference used was the CDS of genes predicted in SPRZ00000000 in the NCBI WGS. The CDS fasta is provided in the processed files.
Supplementary_files_format_and_content: CSV file provides unique gene reads for each sample originated from CLC Genomics RNAseq analysis (Column Headers = Strain-Condition-Replicate#; Row Headers = Gene IDs).

organism: Fusarium graminearum

source_name: Fungal mycelia

growth_protocol: 20,000 spores in 4 ml of PN medium were grown in 6-well plates (in the dark, at 28 degrees C, rotating at 160rpm); Samples destined for in NPN medium were then transferred to the NPN medium and grown for additional 24 hours under the same conditions. Mycelia was collected and frozen at -80 degrees C.

extract_protocol:
Mycelia was ground in liquid nitrogen; total RNA was purified using Trizol extraction method and cleaned using Invitrogen Stratec kit (Ref# 1061100200).
RNA libraries were prepared for sequencing using standard Illumina protocols.

tag: environmental conditions

value: non-preferred nutrient (NPN)

tag: timepoint

value: 24 hrs

tag: sequencing

value: Illumina NextSeq


title: ∆tri6/TRI6-NPN-BR3


title: ∆tri6/TRI6-NPN-BR1


title: miR29ab1

city: Nanjing

first: Yi

last: Sun

address: Tianyuandong Road 818

institute: Nanjing medical university

data_processing:
HTSeq v0.6.0 was used to count the reads numbers mapped to each gene. And then FPKM of each gene was calculated based on the length of the gene and reads count mapped to this gene. FPKM, expected number of Fragments Per Kilobase of transcript sequence per Millions base pairs sequenced, considers the effect of sequencing depth and gene length for the reads count at the same time, and is currently the most commonly used method for estimating gene expression levels.
Genome_build: GRCm38
Supplementary_files_format_and_content: Matrix table with raw gene counts for every gene and every sample

source_name: pancreatic islets

growth_protocol: 10-12 week male mice

extract_protocol:
Pancreatic islets were isolated, flash frozen on dry ice, and RNA was harvested using Trizol reagent. Illumina TruSeq RNA Sample Prep Kit was used with 1 ug of total RNA for the construction of sequencing libraries.
RNA libraries were prepared for sequencing using standard Illumina protocols

value: pancreatic islets

value: miR-29a/b-1 transgene

value: 10-12 weeks

tag: Sex

value: male


title: Uninf_BIP_2

source_name: Blood

value: Blood


title: Uninf_naso_2

source_name: Nasopharynx

value: Nasopharynx


title: Uninf_naso_1


title: Uninf_BIP_1


title: miR29ab2


title: Uninf_lung_F2

source_name: Lung

value: Lung


title: wt1

value: wild type


title: ∆tri6-NPN-BR2


title: Uninf_lung_F1


title: AN44: patient 27

description: miRNAs in embryonic culture medium

value: F

tag: disease

value: AAK

tag: Stage

value: Severe

value: conjunctival cells


title: AN45: patient 46

description: miRNAs in embryonic culture medium


title: AN40: patient 31

description: miRNAs in embryonic culture medium


title: Uninf_kidney_F2

source_name: Kidney

value: Kidney


title: hMG_200IU_rep6

city: Beijing

first: Ying

last: Huo

address: HuanYuan street No.38

institute: Peking University Third Hospital

data_processing:
Illumina CASAVA version 1.8 were used to the basecalling.
Trim Galore (v0.4.1) were used to remove the first 11 bases of each read. Adaptor sequences and low quality bases were also trimmed with the command ‘trim_galore --clip_R1 11 --quality 20 --phred33 --stringency 3 --gzip --length 30’. The paired-end reads were trimed as single-end reads.
Bismark (v0.18.2) were used to map the clean reads to mouse genome mm9 with single-end and non-directional module (parameters: bismark --bowtie2 --non_directional ).
After alignment, PCR duplicates were removed with the picard tools.
CG methylation level of cytosines was estimated using comstomized scripts.
Genome_build: mm9
Supplementary_files_format_and_content: Each bigWig file contains methylation level of each CG sites.

source_name: oocyte

extract_protocol:
All the denuded oocytes were treated with pronase to remove the zona pellucida and polar body. The MII oocytes from each mouse were treated separately and the certain oocyte with the best morphology and status was selected for subsequent DNA methylation library preparation.
Libraries were prepared using pBAT-sequencing protocol.

treatment_protocol: Natural control MII oocytes (n = 6) were taken direct from the ovary and fallopian tube of estrous female mice. Each experimental group of mice were intraperitoneal-injected with different dosages of FSH (FSH 5 IU, n = 6; FSH 50 IU, n = 6; FSH 200 IU, n = 6) or hMG (hMG 5 IU, n = 6; hMG 50 IU, n = 6; hMG 200 IU, n = 7) for 48 h and primed with 0.1 mL 5 IU hCG respectively. hCG control group was also designed as this group of mice were treated with only 0.1 mL 5 IU hCG (n = 6). Then, all these female mice were sacrificed about 14 h later by cervical dislocation to obtain ampullae. Cumulus-oocyte complexes were collected from oviduct ampullae and digested by 0.2% hyaluronidase for a short time.

value: hMG 200IU

value: oocyte

tag: develomental stage

value: MII oocyte


title: AN42: patient 30

description: miRNAs in embryonic culture medium

source_name: conjunctival cells

value: M

value: Mild


title: hMG_200IU_rep5


title: AN4: patient 38

description: miRNAs in embryonic culture medium


title: Uninf_kidney_F1


title: ∆tri6-NPN-BR3

value: non-preferred nutrient (NPN)

value: 24 hrs

value: Illumina NextSeq


title: WT-NPN-BR2

value: WT


title: wt2

value: pancreatic islets

value: 10-12 weeks

value: male


title: hMG_200IU_rep3


title: hMG_200IU_rep4


title: AN33: patient 9

description: miRNAs in embryonic culture medium

value: F


title: ∆tri6/TRI6-PN-BR3

value: preferred nutrient (PN)


title: AN38: patient 23

description: miRNAs in embryonic culture medium

value: AAK

value: Severe


title: WT-NPN-BR1


title: ∆tri6/TRI6-PN-BR2


title: hMG_200IU_rep2


title: ∆tri6/TRI6-PN-BR1


title: ∆tri6-NPN-BR1


title: AN31: patient 39

description: miRNAs in embryonic culture medium

value: conjunctival cells


title: AN3: patient 4

description: miRNAs in embryonic culture medium


title: AN26: patient 34

description: miRNAs in embryonic culture medium


title: WT-NPN-BR3


title: ∆tri6-PN-BR3


title: hMG_50IU_rep5

value: hMG 50IU


title: hMG_50IU_rep4

value: oocyte


title: ∆tri6-PN-BR2


title: hMG_50IU_rep3

value: MII oocyte


title: AN23: patient 10

description: miRNAs in embryonic culture medium


title: AN14: patient 26

description: miRNAs in embryonic culture medium


title: AN17: patient 45

description: miRNAs in embryonic culture medium


title: ∆tri6-PN-BR1


title: hMG_50IU_rep2


title: WT-PN-BR4


title: hMG_200IU_rep1

value: hMG 200IU


title: AN11: patient 8

description: miRNAs in embryonic culture medium

value: Mild


title: AN10: patient 25

description: miRNAs in embryonic culture medium


title: WT-PN-BR2


title: WT-PN-BR3


title: hMG_5IU_rep6

value: hMG 5IU


title: hMG_50IU_rep1


title: Broodiness period_3

first: Chunyou

last: Ning

address: No. 211, Huimin road

institute: SICAU

description:
The_FPKM_of_lncRNA.txt
The_FPKM_of_mRNA.txt
BP3

data_processing:
Total RNA was extracted using  RNAiso Plus reagent (TaKaRa, Japan) according to the manufacturer’s instruction. RNA degradation and contamination were monitored using 1% agarose gels.
The integrity and quality of total RNA samples were analyzed with an RNA 6000 Nano kit (Agilent Technologies, Palo Alto, CA) on a Bioanalyzer 2100 system. Approximately 1 μg total RNA and the Ribo-ZeroTM kit (Epicentre, Madison, WI, USA) were used to generate RNA-Seq cDNA libraries from each sample.
RNA sequencing was performed following the manufacturer’s standard procedures. cDNA size and purity were checked using a high sensitivity DNA 1000 kit and AMPure XP beads and was amplified using PCR. High-quality strand-specific libraries were sequenced on the Illumina HiSeq X Ten platform and 150 bp paired-end reads were generated.
The filtered clean reads were mapped to the chicken reference genome (Gallus gallu-5.0) using HISAT2 2.1.0. The mapped reads were assembled by Stringtie v1.3.3.
Assembled transcripts were then merged into consensus transcripts using custom Python scripts after filtering reads with length less than 200 nt. Next, transcripts annotated as “C” and “=” were removed by Cuffcompare v2.2.1. Remaining transcripts that contained a known protein-coding domain were removed by Hmmscan and BLASTX
LncRNAs were classified based on their genomic characterization by FEELnc.
The expression levels of transcripts were quantified as FPKM values of mRNAs and lncRNAs using StringTie v1.3.3.
Genome_build: Gallus_gallus-5.0
Supplementary_files_format_and_content: tab-delimited text files include FPKM values for each Sample

source_name: liver

growth_protocol: Collection of samples from four developmental stages of chicken liver

extract_protocol:
Total RNA was extracted using RNAiso Plus reagent (TaKaRa, Japan)
RNA libraries were prepared for sequencing using standard Illumina protocols

treatment_protocol: Stored in liquid nitrogen

value: female

value: day 400


title: K5: control 20

hyb_protocol: The mRNA expression profile was determined using Agilent SurePrint G3 Human GE v3 8x60K Microarray according to the manufacturers recommendations

data_processing: Raw expression values were extracted using the Feature Extraction Software. Data was quantil normalized and log transformed.

label_protocol: Low Input Quick Amp Labeling Kit, one-color (Agilent Order ID 5190-2305)

extract_protocol: Total RNA including miRNAs was isolated using theQiagen miRNeasy Kit according to manufacturers recommendations.  RNA concentration was measured using NanoDrop2000 (ThermoScientific). RNA integrity was measured using the NanoRNA kit (Agilent) on a Bioanalyzer2100.

value: W

value: healthy control


title: Broodiness period_1

description:
The_FPKM_of_lncRNA.txt
The_FPKM_of_mRNA.txt
BP1


title: Broodiness period_2

description:
The_FPKM_of_lncRNA.txt
The_FPKM_of_mRNA.txt
BP2


title: K3: control 16


title: Peak laying period_3

description:
The_FPKM_of_lncRNA.txt
The_FPKM_of_mRNA.txt
PL3

value: day 220


title: K36: control 19


title: K32: control 18

value: M


title: WT-PN-BR1

value: WT

value: 24 hrs

value: Illumina NextSeq


title: hMG_5IU_rep5


title: K28: control 14


title: K22: control 12


title: hMG_5IU_rep4


title: K31: control 17


title: K27: control 13


title: K21: control 11


title: K29: control 15


title: Lung organoid - poly(I:C)_rep4 [LO-IC-4]

city: Brno

first: Marco

last: De Zuani

address: Pekarska, 53

department: CTM

country: Czech Republic

institute: FNUSA - ICRC

data_processing:
Raw reads were quality checked (FastQC, MultiQC, minion, swan), preprocessed (seqtk, Trimmomatic, FastQC, MultiQC) and mapped (STAR, Samtools, MultiQC) to the reference genome with gene annotation (genome version: Ensembl GRCh38, gene annotation: Ensembl v94)
Mapped reads were counted and summarized to genes (featureCounts)
Datasets were filtered for low count reads (<10), normalized using a TMM-approach and transformed to logCPM
Genome_build: Ensembl GRCh38.p10
Supplementary_files_format_and_content: For each file, matrix table of featureCount outputs for every gene including raw count values prior to data filtration and normalization
Supplementary_files_format_and_content: Summary matrix table containing raw counts for each file ans each gene, prior to data filtration and normalization

source_name: Lung organoid - poly(I:C)

extract_protocol:
Total RNA was isolated by column separation using a RNeasy Plus Micro Kit with gDNA elimination (Qiagen, Hilden, Germany), according to manufacturer’s protocol. RNA quality was assessed with Bioanalyzer2100 RNA Nano 6000 chips (Agilent technologies, Santa Clara, California, USA) and samples with RIN > 9 were used for sequencing.
An Illumina sequencing library was prepared using the NEBNext Ultra II Directional RNA Library Prep Kit (New England Biolabs, MA, USA) following manufacturer’s protocols.

treatment_protocol: iPSC-derived organoids derived from four independent differentiations were stimulated with either 10 μg/mL LPS, 20 μg/mL poly(I:C) or left untreated.

value: human iPSC-derived lung organoid

value: poly(I:C) 20ug/ml


title: hMG_5IU_rep3


title: Peak laying period_2

description:
The_FPKM_of_lncRNA.txt
The_FPKM_of_mRNA.txt
PL2


title: Peak laying period_1

description:
The_FPKM_of_lncRNA.txt
The_FPKM_of_mRNA.txt
PL1


title: Lung organoid - poly(I:C)_rep3 [LO-IC-3]


title: Sexual maturity period_3

description:
The_FPKM_of_lncRNA.txt
The_FPKM_of_mRNA.txt
SM3

value: day 133


title: hMG_5IU_rep2

value: oocyte


title: Lung organoid - poly(I:C)_rep2 [LO-IC-2]


title: hMG_5IU_rep1

value: MII oocyte


title: Lung organoid - poly(I:C)_rep1 [LO-IC-1]


title: Sexual maturity period_2

description:
The_FPKM_of_lncRNA.txt
The_FPKM_of_mRNA.txt
SM2


title: Lung organoid - LPS_rep4 [LO-LPS-4]

source_name: Lung organoid - LPS

value: LPS 10ug/ml


title: FSH_200IU_rep5

value: FSH 200IU


title: K20: control 10

value: conjunctival cells


title: K2: control 9


title: Lung organoid - LPS_rep3 [LO-LPS-3]


title: Lung organoid - LPS_rep2 [LO-LPS-2]


title: FSH_200IU_rep4


title: Sexual maturity period_1

description:
The_FPKM_of_lncRNA.txt
The_FPKM_of_mRNA.txt
SM1

value: female


title: K18: control 7


title: FSH_200IU_rep2


title: Juvenile period_2

description:
The_FPKM_of_lncRNA.txt
The_FPKM_of_mRNA.txt
JP2

value: day 60


title: FSH_200IU_rep6


title: Juvenile period_1

description:
The_FPKM_of_lncRNA.txt
The_FPKM_of_mRNA.txt
JP1


title: K15: control 5

value: W


title: Lung organoid - untreated_rep4 [LO-NT-4]

source_name: Lung organoid - untreated

value: untreated


title: K12: control 3

value: healthy control


title: FSH_50IU_rep4

value: FSH 50IU


title: K19: control 8


title: K16: control 6


title: Lung organoid - untreated_rep3 [LO-NT-3]


title: FSH_200IU_rep1


title: K11: control 2

value: M


title: Juvenile period_3

description:
The_FPKM_of_lncRNA.txt
The_FPKM_of_mRNA.txt
JP3


title: Lung organoid - untreated_rep2 [LO-NT-2]


title: FSH_50IU_rep3


title: FSH_200IU_rep3


title: NO2_lim

city: Nijmegen

first: Stijn

last: Peeters

state: Gelderland

address: Heyendaalseweg 135

institute: Radboud University

data_processing:
Base calling performed by Illumnina Miseq basecall protocol
Read trimming performed with CLC Genomics Workbench 11.0.1 (Qiagen bioinformatics, Denmark). Quality limit 0.01, maximum of 2 ambiguous nucleotides, no adapter trimming, and removal of 10 nucleotides at the 5’ and 5 nucleotides at the 3’ end. Reads smaller than 50 bp were discarded.
Read filtering performed with CLC Genomics Workbench 11.0.1 (Qiagen bioinformatics, Denmark). Non-mRNA sequences were removed by mapping the reads to all non-coding RNAs obtained from the K. stuttgartiensis MBR1 genome.
Read mapping performed with CLC Genomics Workbench 11.0.1 (Qiagen bioinformatics, Denmark). The non-mapped reads, consisting of the mRNA reads, were subsequently mapped to the annotated K. stuttgartiensis MBR1 genome and mapping results were expressed in RPKM values.
Genome_build: GCA_900232105.1
Supplementary_files_format_and_content: csv file contains the RPKM values per gene in comma separated format for both NO2 and NH4 limited growth.

organism: Candidatus Kuenenia stuttgartensis

source_name: Membrane bioreactor

growth_protocol: Kuenenia stuttgartiensis MBR1 was cultivated in a single cell membrane bioreactor (MBR) (Applikon (Applikon B.V. Schiedam, The Netherlands) as described previously (Kartal, Geerts, and Jetten 2011). The growth medium consisted of (per litre): 1.25 g KHCO3, 0.025 g NaH2PO4, 0.5 ml 1.2 M HCl, 0.15 g CaCl2·2H2O, 0.1 g MgSO4·7H2O and 6.25 mg FeSO4·7 H2O. The reactor volume was 2 L. The pH was maintained at 7.3 using 100 g/L KHCO3 buffer solution. To ensure an anaerobic system both the reactor and the main medium bottle were flushed with a mixture of Argon/CO2 (95%/5%) at 10 ml /min. Excess biomass was removed at 0.138 L per day, resulting in a dilution rate of 0.07 (doubling time of 10 days). For nitrite limited conditions, the reactor received 15 mmol/day NaNO2 and 7.5 mmol/day (NH4)2SO4, resulting in 15 mmol/day NO2- and 15 mmol/day NH4+. For ammonium limited conditions, the NaNO2 load of the reactor was increased in 2 two steps to 18 and finally to 21 mmol/day NaNO2, without changing the (NH4)2SO4 load. A secondary medium bottle containing all medium salts as described above but without NaNO2, only 5 mM (NH4)2SO4 and with reduced FeSO4·7 H2O (3.13 mg/L) to prevent precipitation, was used to add approximately 0.45 mmol (NH4)2SO4 per day. This allowed for fine tuning of the ammonium:nitrite ratio and enabled the nitrite levels to be kept below toxic levels, but between 2 and 20 mg/L (40 and 400 uM).

extract_protocol:
Total RNA was isolated using both the TRIzol™ Reagent and the RiboPure™ bacteria kit (ThermoFisher Scientific, Waltham, USA) according to the manufacturer’s instructions with the exception that 100-fold higher cell numbers were used, since the recommended cell number resulted in very low to no RNA yields. RNA was eluted in DEPC treated water in volumes between 20-25 μl for the TRIzol™ method and 50-70 μl for the RiboPure™ method, of which 1-2 μl was used to quantify the isolated RNA. The isolated RNA was treated with DNAse 1, as described and provided by the RiboPure™ kit.
cDNA libraries were made using TruSeq stranded mRNA Library Prep Kit v2, according to the protocol supplied by the manufacturer (Illumina, San Diego, California U.S.A.).

tag: condition

value: nitrite limited growth

tag: nr of replicates


title: FSH_50IU_rep2

tag: tissue


title: Lung organoid - LPS_rep1 [LO-LPS-1]

tag: treatment


title: Lung organoid - untreated_rep1 [LO-NT-1]


title: K10: control 1

tag: age


title: NH4_lim

value: ammonium limited growth


title: K14: control 4

tag: gender


title: Intestinal organoid - poly(I:C)_rep4 [IO-IC-4]

source_name: Intestinal organoid - poly(I:C)

value: human iPSC-derived intestinal organoid


title: L7d-1B [RNA-Seq]

city: Harbin

first: Jian

last: Wang

address: 600 Changjiang Road

institute: Northeast Agricultural University

data_processing:
The sequencing data was filtered with SOAPnuke (v1.5.2)  by Removing reads  containing sequencing adapter; Removing reads whose low-quality base ratio (base  quality less than or equal to 5) is more than 20%; Removing reads whose unknown  base ('N' base) ratio is more than 5%, afterwards clean reads were obtained and stored  in FASTQ format. The clean reads were mapped to the reference genome using HISAT2  (v2.0.4) . After that, Ericscript (v0.5.5)   and rMATS (V3.2.5) were used to detect  fusion genes and differential splicing genes (DSGs), respectively. Bowtie2 (v2.2.5)   was applied to align the clean reads to the gene set, a database built by BGI (Beijing  Genomic Institute in ShenZhen), in which known and novel, coding and noncoding  transcripts were included, then expression level of gene was calculated by RSEM  (v1.2.12)   . The heatmap was drawn by pheatmap (v1.0.8)    according to the gene  expression in different samples. Essentially, differential expression analysis was  performed using the DESeq2(v1.4.5)  with Q value ≤ 0.05. To take insight to the  change of phenotype, GO (http://www.geneontology.org/) and KEGG  (https://www.kegg.jp/) enrichment analysis of annotated different expression gene  was performed by Phyper (https://en.wikipedia.org/wiki/Hypergeometric_distribution)  based on Hypergeometric test. The significant levels of terms and pathways were  corrected by Q value with a rigorous threshold (Q value ≤ 0.05) by Bonferroni   .
Genome_build:  Helianthus annuus (assembly HanXRQr1.0)(https://www.ncbi.nlm.nih.gov/genome/?term=Helianthus+annuus)
Supplementary_files_format_and_content: EXCEL files include FPKM values for each Sample

organism: Helianthus annuus

growth_protocol: L-1-OL-1 were planted in the experimental field under natural conditions at the industrial crops institute, Heilongjiang Academy of Agricultural Sciences (Harbin, China).Seed samples were collected at 7-, 22- and 37 day.

extract_protocol:
Ethanol precipitation protocol and CTAB-PBIOZOL reagent was used for the purification of total RNA from the plant tissue according to the manual instructions. Grind tissue samples about 80mg with liquid nitrogen into powder and transfer the powder samples in 1.5ml of preheated 65°C CTAB-pBIOZOL reagents. The samples were incubated by Thermo mixer for 15min at 65°C to permit the complete dissociation of nucleoprotein complexes. After centrifuge at 12000×g for 5min at 4°C, the supernatant was added 400ul of chloroform per 1.5ml of CTAB-pBIOZOL reagent and was centrifuged at 12000×g for 10min at 4°C. The supernatant was transferred to a new 2.0ml tube that added 700ul acidic phenol and 200ul chloroform followed by centrifuging 12000×g for 10 min at 4°C. The aqueous phase was added equal volume of aqueous phase of chloroform and centrifuged at 12000×g for 10 min at 4°C. The supernatant was added equal volume of supernatant of isopropyl alcohol and placed at -20°C for 2 hours for precipitation. After that, the mix was centrifuged at 12000×g for 20min at 4°C and then removed the supernatant. After washed with 1ml of 75% ethanol, the RNA pellet was air-dried in the biosafety cabinet and was dissolved by adding 50µL of DEPC-treated water. Subsequently, total RNA was qualified and quantified using a Nano Drop and Agilent 2100 bioanalyzer (Thermo Fisher Scientific, MA, USA).
RNA libraries were prepared for sequencing using standard BGI500 protocols
According to the manufacturer’s instructions, the first step involves the removal of ribosomal RNA  (rRNA) using target-specific oligos and RNase H reagents to deplete both cytoplasmic (5S rRNA,  5.8S rRNA, 18S rRNA and 28S rRNA) and mitochodrial ribosomal RNA (12S rRNA and 16S  rRNA) from total RNA preparations. Following SPRI beads purification, the RNA is fragmented  into small pieces using divalent cations under elevated temperature. The cleaved RNA fragments  are copied into first strand cDNA using reverse transcriptase and random primers, followed by  second strand cDNA synthesis using DNA Polymerase I and RNase H. This process removes the  RNA template and synthesizes a replacement strand, incorporating dUTP in place of dTTP to  generate ds cDNA. These cDNA fragments then have the addition of a single 'A' base and  subsequent ligation of the adapter. After UDG treatment, the incorporation of dUTP quenches the  second strand during amplification. The products are enriched with PCR to create the final cDNA  library. The libraries were assessed quality and quantity in two methods: check the distribution of  the fragments size using the Agilent 2100 bioanalyzer, and quantify the library using real-time  quantitative PCR (QPCR) (TaqMan Probe). The qualified libraries were sequenced pair end on the  BGISEQ-500/ MGISEQ-2000 System (BGI-Shenzhen, China).

treatment_protocol: no


title: L7d-1C [RNA-Seq]


title: FSH_50IU_rep1

tag: strain


title: L7d-1A [RNA-Seq]

tag: time point


title: FSH_5IU_rep5

value: FSH 5IU


title: L37d-1C [RNA-Seq]


title: FSH_5IU_rep4


title: L37d-1B [RNA-Seq]


title: FSH_5IU_rep3


title: Intestinal organoid - poly(I:C)_rep1 [IO-IC-1]

value: poly(I:C) 20ug/ml


title: Intestinal organoid - poly(I:C)_rep3 [IO-IC-3]


title: AN47: patient 5

value: AAK

value: Mild


title: AN49: patient 32

value: F


title: Intestinal organoid - LPS_rep4 [IO-LPS-4]

source_name: Intestinal organoid - LPS

value: LPS 10ug/ml


title: AN46: patient 33

value: conjunctival cells


title: AN6: patient 36

value: Severe


title: FSH_5IU_rep2

value: oocyte

value: MII oocyte


title: AN48: patient 40


title: Intestinal organoid - LPS_rep3 [IO-LPS-3]


title: L22d-1C [RNA-Seq]


title: L22d-1B [RNA-Seq]


title: L37d-1A [RNA-Seq]


title: FSH_5IU_rep1


title: Intestinal organoid - LPS_rep2 [IO-LPS-2]


title: Intestinal organoid - poly(I:C)_rep2 [IO-IC-2]


title: hCG_control_rep5

value: hCG control


title: L22d-1A [RNA-Seq]


title: Intestinal organoid - LPS_rep1 [IO-LPS-1]


title: AN44: patient 27


title: AN42: patient 30


title: AN40: patient 31


title: hCG_control_rep4


title: AN45: patient 46


title: AN38: patient 23


title: Intestinal organoid - untreated_rep4 [IO-NT-4]

source_name: Intestinal organoid - untreated

value: untreated


title: IR4 (2)

description: same as GSM2293983

organism:
Streptococcus pneumoniae
Mus musculus

source_name: Heart

value: Heart

tag: s. pneumoniae strain


title: hCG_control_rep3


title: IR3 (2)

description: same as GSM2293982


title: Intestinal organoid - untreated_rep3 [IO-NT-3]


title: hCG_control_rep2


title: hCG_control_rep1


title: IR1 (2)

description: same as GSM2293981


title: Intestinal organoid - untreated_rep1 [IO-NT-1]


title: Intestinal organoid - untreated_rep2 [IO-NT-2]


title: Natural_control_rep3

value: Natural control


title: AN31: patient 39

value: M


title: AN33: patient 9


title: BIP1 (2)

description: same as GSM2493799

value: Blood


title: BIP2 (2)

description: same as GSM2493800


title: AN4: patient 38


title: Natural_control_rep2


title: T4_BF_8_9_14_U (2)

description: same as GSM2293989

organism: Streptococcus pneumoniae

source_name: Streptococcus pneumoniae cells in vitro

value: Streptococcus pneumoniae cells in vitro


title: T4_BF_8_6_14_U (2)

description: same as GSM2293988


title: AN3: patient 4


title: AN26: patient 34


title: TIGR4_naso_2

value: Nasopharynx


title: AN23: patient 10


title: mut_litter_1425_m2

city: Saint Louis

first: Kristen

middle: L

last: Kroll

state: MO

address: 320 McDonnell Sciences/660 S. Euclid Ave.

department: Developmental Biology

institute: Washington University School of Medicine

data_processing:
Raw reads were aligned to mouse genome mm10 using STAR version 2.5.4b with annotation gencode vM9 and default paramters.
Aligned reads were quantified using subreads featureCounts version v1.4.6-p4
Genome_build: mm10
Supplementary_files_format_and_content: tab delimited featureCounts gene level expression data

source_name: mutant_forelimb bud

extract_protocol:
Six litter and somite-matched pairs of Gmnnf/f; Prx-Cre (mutant) and Gmnn+/f; Prx-Cre (control) E10.5 embryos were used to prepare RNA. RNA was quantified with the NanoDrop ND-1000 spectrophotometer (Thermo Scientific), and the quality of total RNA was further confirmed using the Agilent Bioanalyzer 2100. Samples were rejected if the RNA integrity number was below 8.
RNA-seq library preparation (Ribo-zero) and Illumina sequencing were performed by the Genome Technology Access Center at Washington University.

tag: strain background

tag: genotype/variation

value: Gmnn f/f; Prx-Cre

value: Forelimb bud


title: TIGR4_naso_1


title: T4_PK_8_9_14_U (2)

description: same as GSM2293986


title: AN17: patient 45

value: AAK


title: mut_litter_1420_m5


title: TIGR4_lung_F5

value: Lung


title: TIGR4_lung_F3


title: mut_litter_1424_m3


title: T4_BF_7_25_14_T (2)

description: same as GSM2293987


title: T4_kidney_2

value: Kidney


title: AN14: patient 26

value: F

value: Mild

value: conjunctival cells


title: AN11: patient 8


title: T4_PK_7_24_14_T (2)

description: same as GSM2293984


title: AF1

city: Yangling

first: Rui

last: Jiang

address: Litai street

institute: Northwest A&F University

description: circRNA

data_processing:
From the raw FASTQ files, adapter was removed using Trim Galore and filtered data were aligned to the Bos taurus reference genome (bosTau7) from UCSC (http://genome.ucsc.edu/) with TopHat2 (version 2.0.14)
Linear transcripts assembly and abundance estimation were first performed using CuffLinks, and then the reads of fusion transcripts, obtained from TopHat2, which did not align to the linear RNA were identified.
Then the program TopHat-Funsion was done with the fusion transcripts and the transcripts obtained from the assembled linear RNA to obtain back-spliced junctions reads. Only circRNAs within 100 kb, with two or more supporting reads, and with no more than two mismatches were kept.
CircRNAs were normalized as the number of uniquely mapped fragments per kilobase of exon per million fragments mapped (FPKM).
Supplementary_files_format_and_content: Additional file 1：The length distribution of all circRNA
Supplementary_files_format_and_content: Additional file 2：48 circRNA significantly reduced during calf initiation
Supplementary_files_format_and_content: Additional file 3：33 circRNA significantly upregulated during calf initiation
Supplementary_files_format_and_content: Additional file 4：1682 terms enriched in biological processes of Gene Ontology enrichment
Supplementary_files_format_and_content: Additional file 5：90 KEGG pathways were enriched in bovine fat tissues.
Supplementary_files_format_and_content: Additional file 6：List of all circRNAs identified in fat tissues.
Supplementary_files_format_and_content: Additional file 7：raw counts of all circRNAs identified in fat tissues.

organism: Bos taurus

source_name: Subcutaneous fat

growth_protocol: Six subcutaneous fat samples from Qinchuan cattles of two developmental states (calf, 6 M old; and adult, 24 M old) were collected from Yiming abattoir, a local slaughterhouse of Xi’An, P.R. China. Tissue samples were snap-frozen in liquid nitrogen and stored at –80°C freezer until use.

extract_protocol:
Total RNA was assessed by electrophoresis on a denaturing agarose gel and quantified by NanoDrop spectrophotometer (NanoDrop, USA) and Agilent 2100 Bioanalyzer (Agilent, Santa Clara, CA). The total RNA samples (3 mg) were treated with the epicentre Ribo-ZeroTM Kit to remove rRNA before constructing the RNA-seq libraries. For RNase R treatment, ribosome-depleted RNA was incubated for 15 min at 37°C with 5 units μg−1 of RNase R (Epicentre Technologies, Madison, WI).
Then the rRNA depleted RNA samples or circRNAs samples were fragmented and used to synthesize first- and second-strand complementary DNA (cDNA) with random hexamer primers, dNTPs and DNA Polymerase I. The cDNA fragments were cleaned and concentrated using AMPure XP beads, then the ends were repaired and modified with T4 DNA polymerase Klenow DNA polymerase to add A and adapter at the 3’ end of the DNA fragments. The ligated cDNA products were purified with AMPure XP beads and treated with uracil DNA glycosylase to remove the second-strand cDNA.
Purified first-strand cDNA was subjected to PCR amplification, and the libraries were quality controlled with an Agilent 2100 Bioanalyzer and sequenced by HiSeq 2500 (Illumina, San Diego, CA) on a 150 bp paired-end (PE) run.

value: adipose tissue

tag: developmental stage

value: adult

value: 24 months old


title: AF3

description: circRNA


title: Natural_control_rep1

value: oocyte

value: MII oocyte


title: mut_litter_1424_m1


title: M8

city: Madison

first: Francisco

last: Peñagaricano

state: Wisconsin

address: 1675 Observatory Drive

department: Animal and Dairy Sciences

institute: University of Wisconsin-Madison

data_processing:
The quality of the sequencing reads was evaluated using FastQC software (version 0.11.7). Read trimming was performed using Trim Galore software (version 0.4.4).
Reads were mapped to the latest bovine reference genome (ARS-UCD1.2) using Hisat2 sofware (version 2.1.0).
Transcript models were recontructed from read alignments using Cufflinks software (version 2.2.1).
The tool cuffmerge (Cufflinks) was used for merging together each of the assemblies with the reference bovine annotation file in order to combine novel isoforms with known isoforms.
The number of reads that mapped to each gene was obtained using the script htseq-count (mode intersection-nonempty).
Genome_build: Bovine genome assembly: (ARS-UCD1.2)
Supplementary_files_format_and_content: Tab-delimited text files including ensembl gene id and number of mapped reads

source_name: Muscle

extract_protocol:
RNA extraction was performed using RNeasy Mini Kit (Qiagen, Valencia, CA). RNA quantiy and quality were determined using Nanodrop™ (2000/2000c Spectrophotometers, Thermo Scientific). All samples had an A260/280 ratio > 1.8.
All the RNA-sequencing procedures were performed by Novogene Bioinformatics Technology Co., Ltd (Beijing, China). Sequencing libraries were prepared using a Poly-A tail capture method and sequenced with NovaSeq 6000.

value: Muscle

tag: breed

value: Holstein


title: D39_naso_2

source_name: Nasopharynx


title: AN10: patient 25

value: Severe


title: AF2

description: circRNA


title: M7


title: T4_PK_8_6_14_U (2)

description: same as GSM2293985

value: Streptococcus pneumoniae cells in vitro


title: T4_kidney_1

source_name: Kidney


title: T4_kidney_3


title: CF3

description: circRNA

value: calf

value: 6 months old


title: M6


title: mut_litter_1452_m2

value: Gmnn f/f; Prx-Cre

value: Forelimb bud


title: cont_litter_1424_m4

source_name: control_forelimb bud

value: Gmnn +/f; Prx-Cre


title: M5


title: D39_naso_1

value: Nasopharynx


title: mut_litter_1452_m8


title: cont_litter_1420_m2


title: CF1

description: circRNA


title: D39_lung_F2

source_name: Lung

value: Lung


title: M4

value: Control


title: cont_litter_1425_m1


title: EC109 EZH2-ChIPseq

city: San Francisco

first: Wei

last: Wu

address: 600 16th street

department: Department of Medicine

institute: University of California, San Franciso

data_processing:
Illumina Casava1.7 software used for basecalling.
ChIP-seq reads were aligned to the hg19 genome assembly using BWA aligner.
Peaks were called using MACS2 algorithm (version 2.2.6), followed by ChIPseeker package (version 1.22.1) in R
Genome_build: hg19

extract_protocol:
Lysates were clarified from sonicated nuclei and histone-DNA complexes were isolated with antibody.
Libraries were prepared according to Illumina's instructions accompanying the DNA Sample Kit (Part# 0801-0303). Briefly, DNA was end-repaired using a combination of T4 DNA polymerase, E. coli DNA Pol I large fragment (Klenow polymerase) and T4 polynucleotide kinase. The blunt, phosphorylated ends were treated with Klenow fragment (32 to 52 exo minus) and dATP to yield a protruding 3- 'A' base for ligation of Illumina's adapters which have a single 'T' base overhang at the 3’ end. After adapter ligation DNA was PCR amplified with Illumina primers for 15 cycles and library fragments of ~250 bp (insert plus adaptor and PCR primer sequences) were band isolated from an agarose gel. The purified DNA was captured on an Illumina flow cell for cluster generation. Libraries were sequenced on the Genome Analyzer following the manufacturer's protocols.

value: Esophageal squamous carcinoma

tag: chip antibody


title: cont_litter_1452_m3


title: Het-1A EZH2-ChIPseq

value: Immortalized normal esophageal epithelium


title: M2


title: D39_kidney_3

value: Kidney


title: mtrr-1(ww56_rep2

city: Worcestor

first: Gabrielle

last: Giese

address: 368 Plantation St.

institute: University of Massachusetts Medical School

description:
grown on NGM + 20 nM vitamin B12 + E. coli OP50
ww56-2

data_processing:
BGI internal software "SOAPnuke" was used to remove adaptors and low quality reads with parameters: -n 0.001 -l 20 -q 0.4 -A 0.25. Low quality reads are defined as having greater than 40% bases whose quality is less than 20.
Reads were aligned to the reference genome using Bowtie2 with default parameters.
RSEM was used for estimation of read counts with default parameters
The Deseq2 package in R 3.6.3 was used for differential expression analysis.
Genome_build: c_elegans.PRJNA13758 (WormBase WS271)
Supplementary_files_format_and_content: tab delimeted file containing TPM matrix for all samples and all genes

source_name: whole animal

extract_protocol:
A minumum of 200 staged animals were collected and washed thrice in M9 buffer. Total RNA was extracted via Trizol and BCP followed by Dnase I treatment. Zymo Direct-zol RNA miniprep kit was used for clean up.
BGI standard protocol

treatment_protocol: Synchronized animals were grown on NGM-agar containing 20 nM vitamin B12 and seeded with E. coli OP50.

value: whole animal

tag: genotype


title: cont_litter_1452_m1


title: D39_kidney_1


title: AT8

source_name: Adipose

value: Adipose


title: M1

value: Muscle


title: metr-1(ww52)_rep2

description:
grown on NGM + 20 nM vitamin B12 + E. coli OP50
ww52-2

tag: Stage


title: D39_BIP_2

source_name: Blood

value: Blood


title: CF2

description: circRNA

value: adipose tissue


title: mtrr-1(ww56)_rep1

description:
grown on NGM + 20 nM vitamin B12 + E. coli OP50
ww56-1


title: HGC-27 SOS

city: shanghai

first: yang

last: tang

address: Yueyang Road 320

institute: Shanghai Institute of Biochemistry and Cell Biology

data_processing:
Illumina platform used for base-calling, fastx_toolkit_0.0.14, SeqPrep, and Sickle softwares used for quality control of sequence reads.
Clean data was assambled for mapped data with Hisat2 software, then mapped to GRCh38.p10 whole genome using Cufflinks software.
Reads Per Kilobase of exon per Megabase of library size (RPKM) were calculated with RSEM software. Exons from all isoforms of a gene were merged to create one meta-transcript considering paired-end reads, reads length, and fragment length.
Genes were considered significantly differentially expressed under the following criteria: p-adjust < 0.05 & |log2FC| >= 1.
Genome_build: GRCh38.p10
Supplementary_files_format_and_content: xls file includes RPKM values for each Sample.

source_name: HGC-27 cells

growth_protocol: Cells were incubated with RPMI 1640 medium with 10% FBS in CO2 incubators at 37℃.

extract_protocol:
Total mRNA was extracted using TRIZOL Reagent (Cat#15596-018) following the manufacturer’s instructions and checked for a RIN number to inspect RNA integrity by an Agilent Bioanalyzer 2100 (Agilent technologies). Illumina Hiseq 4000 SBS Kit (Cat#FC-410-1003) was used for the construction of sequencing libraries.
RNA libraries were prepared for sequencing using standard Illumina protocols.

treatment_protocol: Cells were treat with SOS or control peptide using PDPA-drug delivery micelles for 48 hours

value: gastric cancer cell line

tag: differentiated level

value: undifferented

tag: cell line


title: metr-1(ww52)_rep1

description:
grown on NGM + 20 nM vitamin B12 + E. coli OP50
ww52-1


title: HGC-27 Ctrl


title: HGC-27 siSTRN3_3

treatment_protocol: Cells were treat with siSTRN3 mixture or negative control siRNA using RNAIMAX (Thermo) reagent for 72 hours


title: D39_BIP_1


title: AT6

value: Holstein


title: Tumor-infiltrating MAIT cells from 3 CRC patients and PBMC from healthy control

city: Seattle

first: Evan

last: Newell

address: 1100 Fairview Ave N

department: VIDD

institute: Fred Hhutchinson Cancer research center

description: Tumor-infiltrating MAIT cells from 3 CRC patients (MAIT_CRC_2128,MAIT_CRC_2170, MAIT_CRC_22267) and control PBMC (MAIT_PBMC_231117cone3) from 1 healthy donor were obtained by FACS sorting using tet-MR1

data_processing: FASTQ files containing sequenced data were analyzed using the Seven Bridges platform provided by BD

growth_protocol: no growth

extract_protocol:
Tumor-infiltrating MAIT cells from 3 CRC patients and control PBMC from 1 healthy donor were obtained by FACS sorting using tet-MR1
Single cells were isolated using Single cell capture and cDNA synthesis with BD Rhapsody express single-cell analysis system.
Parallel RNA and BD AbSeq sequencing libraries were generated uing BD Rhapsody targeted mRNA and AbSeq amplification kits.

treatment_protocol: Tumor samples were digested using collagenase + Dnase and PBMC were obtained by Ficoll separation.

tag: cell type


title: 6A10_naso_2

tag: tissue


title: AT7

tag: treatment


title: M3


title: D39_lung_F1


title: HGC-27 siSTRN3_2


title: sams-1(ww51)_rep1

description:
grown on NGM + 20 nM vitamin B12 + E. coli OP50
ww51-1


title: D39_kidney_2


title: mthf-1(ww50)_rep2

description:
grown on NGM + 20 nM vitamin B12 + E. coli OP50
ww50-2


title: HGC-27 siSTRN3_1


title: HGC-27 siNC_3


title: mthf-1(ww50)_rep1

description:
grown on NGM + 20 nM vitamin B12 + E. coli OP50
ww50-1


title: 6A10_lung_F4


title: 6A10_naso_1


title: 6A10_lung_F3


title: ∆nhr-10_rep2

description:
grown on NGM + 20 nM vitamin B12 + E. coli OP50
nhr10-2


title: HGC-27 siNC_1


title: HGC-27 siNC_2


title: 6A10_BIP_7


title: AT2


title: ∆nhr-10_rep1

description:
grown on NGM + 20 nM vitamin B12 + E. coli OP50
nhr10-1


title: AT3


title: AT5


title: HCT116_spike-in_40

status: Public on Jun 04 2020

city: Wuhan

first: Dong

last: Chen

state: Hubei

address: 388 GaoXin 2nd Road, East Lake Hi-Tech Development Zone

department: Center for Genome Analysis

institute: ABLife, Inc.

description: processed data file: length_statistic.txt

data_processing:
NextSeq Control Software 1.4 was used for basecalls.
Reads were trimmed for polyG sequence, and polyA sequence shorter than 11nt were discarded. PolyA lengths were counted from basecalls.
Sequenced reads were trimmed for polyA and adaptor sequence, and reads longer than 20nt were mapped to GRCH38 whole genome using tophat2 with parameters -a 8 -m 0 -g 2 --microexon-search --report-secondary-alignments --read-edit-dist 4 N 2 --b2-N 1
the median of polyA sequence length was calculated of each gene.
Genome_build: GRCH38
Supplementary_files_format_and_content: txt file, the median distribution of polyA length of genes in each sample

source_name: HCT116 cells

extract_protocol:
Total RNA was extracted with Trizol and treated with RQ1 DNase (Promega) to remove DNA
Total RNA was partially fragmented with RNase T1. Polyadenylated mRNAs were purified and concentrated with oligo(dT)-conjugated magnetic beads. 3’ adaptor was added to purified mRNAs, followed by reverse transcription performed with RT primer harboring 5' adaptor sequence and randomized hexamer. The cDNAs were purified and PCR amplified for library construction for Illumina HiSeq X Ten system.


title: HeLa_spike-in_40

source_name: HeLa cells

value: HeLa


title: 6A10_BIP_3

data_processing:
FASTQ files were mapped to their respective genomes using HISAT2 (Kim, Langmead et al. 2015) for Mus musculus and Bowtie (Langmead, Trapnell et al. 2009) for pneumococcal genomes of D39, TIGR4, and 6A10. Reads mapped to each respective genome were tabulated in Supplemental Table 1. Gene counts for all samples were then estimated using HTseq (Anders, Pyl et al. 2015). All sample counts for the mouse data were tabulated in Supplemental Table 2. For bacterial counts, core genes without paralogs were determined for D39, TIGR4, and 6A10 using PanOCT (Fouts, Brinkac et al. 2012). Counts were then tabularized for analysis based on core genes, using TIGR4 locus tags as identifiers in Supplemental Table 2. Counts tables were then imported into Rstudio for analyses and estimation of differentially expressed (DE) genes. For mouse DE gene estimation, infected/colonized tissues were compared to their respective uninfected tissue control samples using DEseq2 (Love, Huber et al. 2014) and filtered using an FDR cutoff of 0.05. For bacterial DE gene estimation, each infected tissue was compared to the nasopharyngeal colonization state, while accounting for strain differences within core genes using DEseq2 and filtered using an FDR cutoff of 0.05.
Genome_build: Streptococcus pneumoniae D39 (NC_008533.1), Streptococcus pneumoniae TIGR4 (NC_003028.3), Streptococcus pneumoniae 6A10 (CP053210), Mus musculus (GCA_000001635.5).

extract_protocol:
For nasopharynx samples, euthanized mice were decapitated, exterior skin removed, brain and cerebral column excised, and the skull defleshed. The skull and oropharynx were subsequently crushed in RNA Protect Bacteria Reagent (Qiagen) and stored at -80°C. For blood, samples were obtained by exsanguination via retro-orbital bleed and mixed with RNAprotect Bacteria Reagent and stored at -80°C. For lungs, heart, and kidneys, organs were collected, rinsed with PBS, minced, and placed into RNAprotect Bacteria Reagent and stored at -80°C. On the day of RNA extractions, samples were thawed and spun down to discard the supernatant. Pellets were then incubated in 100 μL of lysis buffer (10 μL of mutanolysin, 20 μL of proteinase K, 30 μL of lysozyme, 40 μL of TE buffer) for 10 minutes. Pellets were mechanically disrupted in 600 μL RLT buffer (RNeasy Mini Kit) containing 1% beta-mercaptoethanol, using a motorized mortar and pestle for 30 seconds. Samples were then spun through a Qiashredder followed by addition of 590 μL of 80% ethanol, and RNA was captured on the RNeasy Mini Kit columns with DNase treatment on column, as per their protocol, followed by DNase treatment in solution. Extracted RNA was quantitated using a Bioanalyzer. Ribosomal RNA was depleted using the Ribo-Zero rRNA Removal Kit for Gram-positive bacteria and/or for human/mouse/rat. RNA was isolated from the planktonic seed cultures and their respective biofilms. In total, 3 independent biological samples were collected with planktonic and biofilm RNAs from each experiment paired. Total RNA was extracted from each replicate separately using enzymatic lysis of pneumococcal cells (10 μL mutanolysin, 20 μL proteinase K, 15 μL lysozyme, and 55 μL TE) followed by RNA extraction using the same protocol mentioned above.
300 bp-insert RNA-seq Illumina libraries were constructed using 0.01 – 2.0 μg of enriched mRNA that was fragmented then used for synthesis of strand-specific cDNA using the NEBNext Ultra Directional RNA Library Prep Kit for Illumina (NEB-E7420L). The cDNA was purified between enzymatic reactions and the size selection of the library performed with AMPure SpriSelect Beads (Beckman Coulter Genomics). The titer and size of the libraries was assessed on the LabChip GX (Perkin Elmer) and with the Library Quantification Kit (Kapa Biosciences).