title: Control_replicate_3

hyb_protocol: A total of 600 ng Cy3-labeled cRNA was fragmented at 60°C for 30 minutes in a reaction volume of 25 microL containing 0.5x Agilent fragmentation buffer and 2x Agilent gene expression blocking agent following the manufacturers instructions.  On completion of the fragmentation reaction, 25 microL of 2× Hi-RPM Hybridization Buffer was added to the fragmentation mixture and hybridized to Agilent SurePrint GE3 Rat 8x60k v2 gene expression microarrays 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), and dried by slowly removing from Wash buffer 2.

organism: Rattus norvegicus

source_name: Control

growth_protocol: Primary hippocampal neuron cultures were prepared from embryonic day 18 Sprague-Dawley rat brains as described previously. Dissociated neurons were plated at a density of 40,000 neurons per well (96-well plate) or 1 x 106 cells per well (6-well plate) on polyethyleneimine-coated plastic dishes in MEM medium supplemented with 10%  fetal bovine serum. Once the neurons attached to the plates, the cell plating medium was replaced with Neurobasal (NB) medium with 5% B27 supplement, 1% glutamax, and 1% antibiotic-antimycotic (Invitrogen, Waltham, MA) for 6-7 days. Prior to the addition of HU (5 µM unless otherwise specified; H8627, Sigma-Aldrich, St. Louis, MO), the neurons were washed with unsupplemented NB medium to remove traces of antioxidants and the medium was replaced with NB medium supplemented with 5% B27 minus antioxidants (NB + B27 – AOX; Invitrogen), 1% glutamax, and 1% antibiotic-antimycotic. Experiments were performed between days in vitro (DIV) 6 and 8.

treatment_protocol: Rat hippocampal neurons were treated with HU (5 µM) for 2, 6, 12, or 24 hours or left untreated (UT), with three replicates (A, B, C) for each condition.

tag: cell type

value: primary neurons

value: 0 hours


title: Liver_CTRL_replicate1

status: Public on Sep 10 2018

type: RNA

city: Baltimore

first: Supriyo

last: De

state: Maryland

address: 251 Bayview Blvd

department: Laboratory of Genetics and Genomics

country: USA

institute: NIA-IRP, NIH

hyb_protocol: A total of 600 ng Cy3-labeled cRNA was fragmented at 60°C for 30 minutes in a reaction volume of 25 microL containing 0.5x Agilent fragmentation buffer and 2x Agilent gene expression blocking agent following the manufacturers instructions.  On completion of the fragmentation reaction, 25 microL of 2× Hi-RPM Hybridization Buffer was added to the fragmentation mixture and hybridized to Agilent SurePrint GE3 Mouse 8x60k v1 gene expression microarrays (G4852-60510) 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), and dried by slowly removing from Wash buffer 2.

scan_protocol: Following posthybridization rinses, arrays were scanned using an Agilent SureScan microarray Scanner at 5 micron, and hybridization intensity data extracted from the scanned images using Agilent’s Feature Extraction Software 9.1 (Agilent) using default parameters to obtain background subtracted and spatially detrended Processed Signal intensities.

data_processing: Raw microarray hybridization intensity data from four separate experiments were log-transformed.  Raw microarray data were log transformed to yield z-scores. The z-ratio was calculated as the difference between the observed gene z-scores for the experimental and the control comparisons, and dividing by the standard deviation associated with the distribution of these differences. Z-ratio values of ± 1.5 were used as cut-off values and calculated using a 5% false discovery rate (FDR) threshold. The complete set was tested for gene set enrichment using parametric analysis of gene set enrichment. For each pathway z-score, a p-value was computed using JMP 6.0 software to test for the statistical significance.

molecule: total RNA

organism: Mus musculus

label_protocol: Cyanine-3 (Cy3) labeled cRNA was prepared from 200 ng total RNA using the Low-Input Quick Amp Labeling Kit (Agilent, Santa Clara, CA) according to the manufacturer's instructions, followed by RNeasy column purification (QIAGEN, Valencia, CA).  Dye incorporation and cRNA yield were evaluated with the NanoDrop ND-1000 Spectrophotometer (Wilmington, DE).

growth_protocol: Thirty-five 6 month old male C57 BL/6J mice (Jackson Laboratories, Bar Harbor, ME) were housed individually with ad libitum access to water.  Animal rooms were maintained at 20-22°C with 30%-70% relative humidity and a 12 hour light/12 hour dark cycle. Blood samples were collected via eye bleeds and a VO2max test was performed prior to the initiation of dietary and exercise interventions.

extract_protocol: RNA extraction was performed using Qiagen RNeasy Mini Kit (Qiagen, Inc., Valencia, CA, USA), as per the manufacturers instructions, using the fibrous tissue protocol for soleus muscle.

treatment_protocol: Mice were randomly assigned to one of four groups: sedentary ad libitum fed control (CTRL; n=9); sedentary alternate day fasting (AF; n=8); daily treadmill exercise fed ad libitum (EX; n=9); and daily treadmill exercise alternate-day fasting (EXAF; n=9). All other mice completed all tests in the study, except for the maximum endurance test in which one mouse in the CTRL group and one mouse in the EXAF group refused to run on the treadmill.  For AF and EXAF groups, food (standard mouse chow: NIH-07 diet from Harlan-Teklad, Indianapolis, IN) was alternatively provided or removed every day at 17:00. Body weights were recorded weekly.  Average weekly food/energy consumption was calculated (the weight of food provided minus the weight of food remaining uneaten) x kcal/gram of food.  Blood glucose measurements were made at baseline, and every two weeks on consecutive feeding and fasting days, before and after exercise using a Freestyle Lite® Abbott meter. On the fourth week of training blood ketone levels were measured on consecutive feeding and fasting days, before and after exercise using a StatSite® M B-HB meter. After 8 weeks of training the animals were anesthetized and decapitated. AF and EXAF mice were euthanized on a fasting day and EX mice were sacrificed 18 hours after the last exercise session.  All mice were sacrificed between the hours of 9:00 and 12:00. Trunk blood, liver and soleus muscle tissues were collected and immediately flash frozen and stored at -80 C.

tag: treatment

value: Control

tag: tissue

value: Liver

tag: developmental stage

value: 6 month old

tag: Sex

value: male

tag: strain


title: Hydroxyurea_12 hours_replicate_3

data_processing: Raw microarray hybridization intensity data were log-transformed.  Raw microarray data were log transformed to yield z-scores. The z-ratio was calculated as the difference between the observed gene z-scores for the experimental and the control comparisons, and dividing by the standard deviation associated with the distribution of these differences. Z-ratio values of ± 1.5 were used as cut-off values and calculated using a 5% false discovery rate (FDR) threshold. The complete set was tested for gene set enrichment using parametric analysis of gene set enrichment. For each pathway z-score, a p-value was computed using JMP 6.0 software to test for the statistical significance.

source_name: Hydroxyurea_12 hours

extract_protocol: RNA was purified from hippocampal tissue using Trizol (Thermo Fisher Scientific) according to the manufacturer's instructions.

value: Brain

tag: region

value: Hippocampus

tag: age

value: hydroxyurea (5 microM)

tag: treatment time

value: 12 hours


title: Liver_CTRL_replicate3


title: Hydroxyurea_24 hours_replicate_2

source_name: Hydroxyurea_24 hours

value: 24 hours


title: Control_replicate_1


title: Liver_CTRL_replicate6


title: Hydroxyurea_6 hours_replicate_2

source_name: Hydroxyurea_6 hours

value: 6 hours


title: Hydroxyurea_6 hours_replicate_3


title: Hydroxyurea_2 hours_replicate_3

source_name: Hydroxyurea_2 hours

value: 2 hours


title: Liver_CTRL_replicate7


title: Hydroxyurea_2 hours_replicate_1


title: Hydroxyurea_24 hours_replicate_1


title: Liver_CTRL_replicate2


title: WT_replicate_2

status: Public on Aug 29 2018

hyb_protocol: A total of 825 ng Cy3-labeled cRNA (specific activity >6 pmol Cy3/ug cRNA) was fragmented at 60°C for 30 minutes in a reaction volume of 55 microL containing Agilent fragmentation buffer and Agilent gene expression blocking agent following the manufacturers instructions.  On completion of the fragmentation reaction, 55 microL of 2× Hi-RPM Hybridization Buffer was added to the fragmentation mixture and hybridized to Agilent D. melanogaster 4×44K Gene Expression Microarray for 17 hours at 65°C in a rotating Agilent hybridization oven.  After hybridization, microarrays were rinsed 1 minute at room temperature with GE Wash Buffer 1 (Agilent) and 1 minute at 37°C with GE Wash buffer 2 (Agilent), and dried by slowly removing from Wash buffer 2.

data_processing: Raw microarray hybridization intensity data were log-transformed. Raw microarray data were log transformed to yield z-scores. The z-ratio was calculated as the difference between the observed gene z-scores for the experimental and the control comparisons, and dividing by the standard deviation associated with the distribution of these differences. Z-ratio values of ± 1.5 were used as cut-off values and calculated using a 5% false discovery rate (FDR) threshold. The complete set was tested for gene set enrichment using parametric analysis of gene set enrichment. For each pathway z-score, a p-value was computed using JMP 6.0 software to test for the statistical significance.

organism: Drosophila melanogaster

source_name: WT

growth_protocol: Drosophila melanogaster (D. melanogaster) flies were cultured on corn syrup-soy recipe food from Archon Scientific at 25±1°C and 60±5% humidity, under a 12h/12hr light/dark cycle. Wild-type W1118 flies were purchased from Bloomington Drosophila Stock Center, while Top3b p-element remobilized mutants Top3b26 was a generous gift from T.S. Hsieh.

extract_protocol: A total 200 microL of Drosophila heads/per sample were collected by fast freezing and sieving. Total RNA was extracted using TRIzol reagent according to the manufacturer's protocol (ThermoFisher, Waltham, MA USA). Three sets of independently prepared RNA samples employed, and the quality and quantity of RNA was assessed by NanoDrop (ThermoFisher) and by Agilent Bioanalyzer RNA 6000 Chip (Agilent, Santa Clara, CA) analysis.

tag: strain background

tag: genotype/variation

value: TOP3B wildtype

value: Adult

value: head


title: Liver_CTRL_replicate4


title: Hydroxyurea_12 hours_replicate_1


title: TOP3BETA26_replicate_2

source_name: TOP3BETA mutant

value: TOP3B mutant (TOP3B26)


title: WT_replicate_3


title: Control_replicate_2

value: primary neurons


title: WT_replicate_1


title: TOP3BETA26_replicate_3


title: Hydroxyurea_2 hours_replicate_2


title: Control_replicate_1_repeat

value: 0 hours


title: Hydroxyurea_12 hours_replicate_2


title: TOP3BETA26_replicate_1


title: Liver_WT_replicate_4

status: Public on Jul 31 2018

hyb_protocol: A total of 300 ng Cy3-labeled cRNA was fragmented at 60°C for 30 minutes in a reaction volume of 25 microL containing 0.5x Agilent fragmentation buffer and 2x Agilent gene expression blocking agent following the manufacturers instructions.  On completion of the fragmentation reaction, 25 microL of 2× Hi-RPM Hybridization Buffer was added to the fragmentation mixture and hybridized to Agilent SurePrint GE3 Mouse 8x60k v1 gene expression microarrays (G4852-60510) 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), and dried by slowly removing from Wash buffer 2.

growth_protocol: NQO1-transgenic mice overexpressing the rat Nqo1 gene (SI Appendix, SI text and Fig. S4) and CYB5R3-Tg mice that overexpress the rat Cyb5r3 gene (Martin-Montalvo et al. 2016) were employed to generate RedTg mice. Group-housed mice were maintained in a room at constant temperature (20°-22°C) with 30-70% relative humidity and 12-h light:dark cycles. Mice were fed ad libitum on a standard diet [SD - (#110700 - Research Diets, Bethlehem, PA)].  All animal protocols were approved by the ACUC of the National Institute on Aging, an AAALAC accredited institution (protocol #277-TGB-2019). Animals were inspected daily for health issues, and deaths were recorded for each animal and a gross necropsy assessment was performed.

extract_protocol: Total RNA was extracted from frozen liver tissue using Trizol® reagent (Thermo Fisher Scientific, Waltham, MA) as per the manufacturers instructions. RNA concentration and quality was measured by Nanodrop (ThermoFisher, Waltham, MA USA) and the Agilent Bioanalyzer RNA 6000 Chip (Agilent, Santa Clara, CA).

treatment_protocol: Fourteen month old male wildtype and RedTg mice were euthanized, and liver tissue was quickly frozen, and stored at -80oC until processing.

value: 14 months

value: liver


title: Hydroxyurea_24 hours_replicate_3


title: Hydroxyurea_6 hours_replicate_1


title: G10S04Y: Young 13

status: Public on Jun 04 2018

type: SRA

library_source: transcriptomic

data_processing:
Sequences were cleaned and aligned against the human genome version 19 (Ensembl human transcript annotation v72) using STAR aligner (2.4.0j)
Read counts were obtained using subread featureCounts software
“Fusion reads” were parsed and circRNAs were identified from these “fusion reads” using CircExplorer (v1.1)
miRNAs were identified using mirDeep2 software
Differential miRNA and gene expression between young and old was determined using BRB-ArrayTools (DESeq2 R package)
Genome_build: hg19
Supplementary_files_format_and_content: Text files with read counts and Ensembl IDs

organism: Homo sapiens

source_name: serum

extract_protocol:
Total RNA was isolated from 200 μL of serum from 13 young (30.9 ± 0.60 yrs) HANDLS participants and 10 old (81.8 ± 1.87 yrs) BLSA participants using TRIzol LS (Life Technologies, Waltham, MA) according to the manufacturer’s instructions and with the addition of a second phenol/chloroform extraction prior to RNA precipitation as previously described (Noren Hooten et al. 2013). RNA samples were resuspended in 20 μL of RNAse-free water and frozen at -80°C until further use. RNA quality was assessed using an Agilent BioAnalyzer (Agilent Technologies, Santa Clara, CA). Total RNA was fragmented to <200 bp using the Bioruptor NGS (Diagenode, Denville, NJ). Briefly, RNA was diluted in EB buffer to a final volume of 100 µl and subjected to bioruption for 30 min (Power: H position) in an ice-cold DI water bath using 30”/30” ON/OFF cycles with cold water bath replacement every 7.5 min. Sample RNA was then dry vacuumed to 20 μL and tested for fragmentation using the Agilent BioAnalyzer Small RNA chip.
Total RNA isolated as described above was prepared for sequencing using the Ion Total RNA-seq Kit v2 (Life Technologies) with multiplexing according to the manufacturer’s instructions.  The Life Technologies Ion RNA Adaptors were added to the fragmented RNA at 65oC for 10 min and then allowed to hybridize at 30oC for 5 min.  A ligation reaction was done at 30oC for 1 hr.  The Life Technologies reverse transcription primer was incubated with the ligated samples at 70oC followed by snap-cooling on ice for hybridization. A reverse transcription reaction was done using Superscript III at 42oC for 30 min. Clean-up of the cDNA was done using the supplied magnetic bead cleanup module and 100% ethanol. Samples were eluted from the beads using 37oC nuclease free water. Amplification and addition of sample specific indexes was performed using 16 cycles of PCR to enrich the samples for fragments that contained adaptors on both ends, followed by another Magnetic bead clean up.  These enriched samples were tested using the Agilent Bio-Analyzer High Sensitivity DNA chips. Barcoded samples were pooled, and each library template was clonally amplified on Ion Sphere™ Particles in the Ion One Touch II for sequencing on the Ion Proton™ System.

value: serum

value: adult

tag: age (years)


title: G11S05O: Old 10


title: G08S01Y: Young 11


title: G08S02Y: Young 12


title: G08BLSA6159O: Old 9


title: Liver_WT_replicate_1

value: male


title: G08BLSA5887O: Old 8


title: G07S04O: Old 7


title: G07S03Y: Young 10


title: Liver_WT_replicate_2


title: G07S01Y: Young 9


title: Edar-HaCat with RelB siRNA_replicate_3

status: Public on Jul 24 2018

hyb_protocol: Standard Illumina protocol. In short, a total of 0.75 ug of biotin-labeled cRNA was hybridized at 58°C for 16 hours to Illumina's Sentrix Human-HT12 v4 Expression BeadChips (Illumina, San Diego, CA). Each BeadChip has more than 47,000 probes derived from the National Center for Biotechnology Information Reference Sequence (NCBI) RefSeq Release 38 (November 7, 2009) and other sources. The arrays were washed, blocked and the biotin labeled cRNA was detected by staining with streptavidin-Cy3.

scan_protocol: Arrays were scanned at a resolution of 0.53 um using the Illumina iScan scanner.

data_processing: Data was extracted using the Illumina GenomeStudio software V2011.1, Gene expression module 1.9.0. Any spots at or below the background were filtered out using an Illumina detection Pvalue of 0.02 and above. The natural log of all remaining scores were used to find the avg and std of each array and the z-score normalization was calculated and presented below. Z-score = (raw value - avg)/std. Complete data including detection scores is supplied.

source_name: Edar-HaCat with RelB siRNA

label_protocol: Standard Illumina protocol using Illumina TotalPrep RNA Amplification Kit (Ambion; Austin, TX, cat # IL1791) In short, 0.5ug of total RNA was first converted into single-stranded cDNA with reverse transcriptase using an oligo-dT primer containing the T7 RNA polymerase promoter site and then copied to produce double-stranded cDNA molecules. The double stranded cDNA was cleaned and concentrated with the supplied columns and used in an overnight in-vitro transcription reaction where single-stranded RNA (cRNA) was generated and labeled by incorporation of biotin-16-UTP.

extract_protocol: Total RNA was extracted using the Qiagen RNeasy kit as per the manufacturer's instruction.

treatment_protocol: HaCat cells (ATCC) were cultured in DMEM plus 10% FBS and transfected with scrambled siRNA or with SMARTpooled siRNAs (Dharmacon) against hRelB or hBRG1 for 48 h before RNA extraction.

tag: cell line

value: Edar-HaCaT

value: keratinocyte cell line

tag: edar

value: Stable Edar expression

tag: sirna

value: RelB


title: G06S04Y: Young 8


title: Liver_RedTg_replicate_1

value: RedTg


title: Empty-HaCat with control siRNA_replicate_2

source_name: Empty-HaCat with control siRNA

value: HaCaT

value: No Edar expression

value: scrambled


title: Empty-HaCat with control siRNA_replicate_1


title: Liver_RedTg_replicate_4


title: G06S03Y: Young 7


title: Liver_RedTg_replicate_3


title: Liver_RedTg_replicate_2


title: Edar-HaCat with control siRNA_replicate_1

source_name: Edar-HaCat with control siRNA


title: Edar-HaCat with control siRNA_replicate_2


title: Edar-HaCat with RelB siRNA_replicate_1


title: Liver_WT_replicate_3


title: Edar-HaCat with BRG1 siRNA_replicate_2

source_name: Edar-HaCat with BRG1 siRNA


title: Edar-HaCat with control siRNA_replicate_3


title: Empty-HaCat with control siRNA_replicate_3


title: Edar-HaCat with RelB siRNA_replicate_2


title: Edar-HaCat with BRG1 siRNA_replicate_1


title: G05S04Y: Young 6


title: G06S01O: Old 5


title: G06S02O: Old 6


title: G05S02O: Old 4


title: G05S03Y: Young 5


title: G05S01O: Old 3


title: G01S02Y: Young 2


title: G03S02Y: Young 4


title: G03S04O: Old 2


title: G01S01Y: Young 1


title: G03S01Y: Young 3


title: G01S04O: Old 1


title: NSG 7

status: Public on Jun 01 2018

city: Gothenburg

first: Mattias

last: Andersson

address: Medicinaregatan 1F

department: Sahlgrenska Cancer Center

country: Sweden

institute: University of Gothenburg

hyb_protocol: Hybridization to Affymetrix Human Gene 1.0 ST arrays was performed using standard Affymetrix protocols.

scan_protocol: Arrays were scanned using a GeneChip scanner 3000 and image analysis was performed using the GeneChip Operating Software.

data_processing:
Probe summarization and data normalization were performed using Robust Multi-array Analysis (RMA) with the Expression Console Software v1.4.1.
affymetrix-algorithm-name = rma-gene-default
affymetrix-algorithm-version = 1.0
program-name = Expression Console
program-version = 1.4.1.46

label: Biotin

source_name: NSG

label_protocol: Biotin labled cRNA was syntesized from 200 ng total RNA according to standard Affymetrix protocols.

extract_protocol: Total RNA was isolated using the Trizol reagent (Life Technologies) followed by the RNeasy Mini kit (Qiagen).

value: salivary gland

tag: cells

value: normal salivary gland

value: epithelial


title: ACC case 8

source_name: ACC case

value: submandibular gland

value: adenoid cystic carcinoma cells


title: ACC case 14

value: parotid gland


title: ACC case 13


title: NSG 3


title: ACC case 10

value: auditory meatus


title: NSG 2


title: NSG 6


title: ACC case 1

value: lacrimal gland


title: ACC case 6


title: NSG 5


title: NSG 4


title: ACC case 11


title: Cerebellum_Bhlhe40KO_replicate_4

status: Public on May 03 2018

description: Sample name: KO-14100-CER

hyb_protocol: Standard Illumina protocol. In short, a total of 0.75 μg of biotin-labeled cRNA was hybridized at 58°C for 16 hours to Illumina's HumanHT-12 V4.0 Expression Bead Chips (Illumina, San Diego, CA). Each BeadChip has more than 47,000 probes derived from the National Center for Biotechnology Information Reference Sequence (NCBI) RefSeq Release 38 (November 7, 2009) and other sources. The arrays were washed, blocked and the biotin labeled cRNA was detected by staining with streptavidin-Cy3.

scan_protocol: Arrays were scanned at a resolution of 0.53 μm using the Illumina iScan.

data_processing: Data was extracted using the Illumina GenomeStudio software V2011.1, Gene expression module 1.9.0. Any spots at or below the background were filtered out using an Illumina detection Pvalue of 0.02 and above. The natural log of all remaining scores were used to find the avg and std of each array and the z-score normalization was calculated and presented below.  Z-score = (raw value - avg)/std.  Complete data including detection scores is supplied. Detection_Pval = Detection Pvalue from Illumina GenomeStudio software V2011.1, Gene expression module 1.9.0.

label: Biotin, Cy3

label_protocol: Standard Illumina protocol using Illumina TotalPrep RNA Amplification Kit (Ambion; Austin, TX, cat # IL1791) In short, IP RNA was first converted into single-stranded cDNA with reverse transcriptase using an oligo-dT primer containing the T7 RNA polymerase promoter site and then copied to produce double-stranded cDNA molecules. The double stranded cDNA was cleaned and concentrated with the supplied columns and used in an overnight in-vitro transcription reaction where single-stranded RNA (cRNA) was generated and labeled by incorporation of biotin-16-UTP.

extract_protocol: RNA was extracted from brain tissue samples using an electronic tissue homogenizer and 1mL Qiazol followed by spin column isolation according to manufacturer protocols (Qiagen; Cat#: 74104).

treatment_protocol: Animals were fasted overnight prior to isoflurane anesthesia, decapitation, and the brain harvested from naïve 3-5 month old WT (n=4) and Bhlhe40 KO (n=4) between 10:30 am and 1 pm.  The hippocampus, cortex and cerebellum were dissected  on ice, snap frozen on dry ice, and stored at -80˚C until use.

value: Cerebellum

tag: background strain

tag: genotype

value: Bhlhe40 KO

tag: gender

value: Male


title: ACC case 4

tag: tissue

tag: cell type


title: ACC case 5


title: ACC case 9

value: nasal mucosa


title: ACC case 3

value: pelvic bone


title: ACC case 12


title: Cerebellum_Bhlhe40KO_replicate_1

description: Sample name: KO-14097-CER


title: ACC case 7

value: buccal mucosa


title: NSG 1


title: Cerebellum_Bhlhe40KO_replicate_3

description: Sample name: KO-14099-CER


title: Cortex_Bhlhe40KO_replicate_4

description: Sample name: KO-14100-CTX

value: Cortex


title: Hippocampus_Bhlhe40KO_replicate_4

description: Sample name: KO-14100-HIPP

value: Hippocampus


title: Cortex_Bhlhe40KO_replicate_2

description: Sample name: KO-14098-CTX


title: Cortex_Bhlhe40KO_replicate_3

description: Sample name: KO-14099-CTX


title: Cerebellum_Bhlhe40KO_replicate_2

description: Sample name: KO-14098-CER


title: Cortex_Bhlhe40KO_replicate_1

description: Sample name: KO-14097-CTX


title: Hippocampus_Bhlhe40KO_replicate_3

description: Sample name: KO-14099-HIPP


title: primary_tumor_UPPL1695

status: Public on Apr 12 2018

city: Chapel Hill

first: William

middle: Y.

last: Kim

state: NC

address: 450 West Dr.

department: Medicine and Genetics

institute: University of North Carolina at Chapel Hill

data_processing:
75b paired-end reads were sequenced on a NextSeq 500 Desktop Sequencer using a high output flow cell kit (Illumina), yielding an average of over 28M reads per sample.
QC-passed reads were aligned to the mouse reference genome (mm9) using MapSplice
The alignment profile was determined by Picard Tools v1.64 (http://broadinstitute.github.io/picard/).
Aligned reads were sorted and indexed using SAMtools and translated to transcriptome coordinates then filtered for indels, large inserts, and zero mapping quality using UBU v1.0 (https://github.com/mozack/ubu).
Transcript abundance estimates for each sample were performed using RSEM, an expectation-maximization algorithm using the UCSC knownGene transcript and gene definitions.
Raw RSEM read counts for all RNAseq samples were normalized to the overall upper quartile
Genome_build: mm9
Supplementary_files_format_and_content: Tab-delimited text file includes RSEM values for each sample

source_name: bladder tumor

extract_protocol:
RNA was extracted with Qiagen Rneasy Kit
Libraries were prepared using TruSeq Stranded mRNA Sample Prep Kit (Illumina)

tag: treatment

value: Untreated

tag: abbreviations

value: UPPL

tag: sample type


title: Hippocampus_WT_replicate_1

description: Sample name: WT-14626-HIPP

value: Wildtype


title: Cortex_WT_replicate_2

description: Sample name: WT-14627-CTX


title: Cortex_WT_replicate_1

description: Sample name: WT-14626-CTX


title: Cerebellum_WT_replicate_3

description: Sample name: WT-14629-CER


title: Cortex_WT_replicate_4

description: Sample name: WT-14629-CTX


title: Ikbkap_DRG_CKO_3

status: Public on Apr 25 2018

hyb_protocol: Standard Illumina protocol. In short, a total of 0.75 μg of biotin-labeled cRNA was hybridized at 58°C for 16 hours to Illumina's Sentrix Mouse Red-8 v3 Expression Bead Chips (Illumina, San Diego, CA). Each BeadChip has more than 47,000 probes derived from the National Center for Biotechnology Information Reference Sequence (NCBI) RefSeq Release 38 (November 7, 2009) and other sources. The arrays were washed, blocked and the biotin labeled cRNA was detected by staining with streptavidin-Cy3.

scan_protocol: Arrays were scanned at a resolution of 0.8um using the Illumina Beadstation 500 X

data_processing: Data was extracted using the Illumina GenomeStudio software V2011.1, Gene expression module 1.9.0. Any spots at or below the background were filtered out using an Illumina detection Pvalue of 0.02 and above. The natural log of all remaining scores were used to find the avg and std of each array and the z-score normalization was calculated and presented below.  Z-score = (raw value - avg)/std.  Complete data including detection scores is supplied.

source_name: DRG_E17.5_Ikbkap CKO

label_protocol: Standard Illumina protocol using Illumina TotalPrep RNA Amplification Kit (Ambion; Austin, TX, cat # IL1791) In short, 0.5 μg of total RNA was first converted into single-stranded cDNA with reverse transcriptase using an oligo-dT primer containing the T7 RNA polymerase promoter site and then copied to produce double-stranded cDNA molecules. The double stranded cDNA was cleaned and concentrated with the supplied columns and used in an overnight in-vitro transcription reaction where single-stranded RNA (cRNA) was generated and labeled by incorporation of biotin-16-UTP.

extract_protocol: Total RNA was extracted using the Rneasy Plus Mini kit (QIAGEN, Valencia, CA) as per the manufacturers instructions. RNA quality and quantity was assessed using the RNA 6000 Nano Chip on an Agilent Bioanalyzer (Agilent, Wilmington, DE).

treatment_protocol: Examining the major genetic pathways that require IKAP are hampered by that Ikbkap null mice are embryonic lethal (E10), but can be examined by genome-wide transcriptome analysis of the dorsal root ganglia from Wnt1-Cre;IkbkapLoxP/LoxP  E17.5 fetuses (Ikbkap is deleted from the neural crest), or in the brains in adult Tuba1a-Cre;IkbkapLoxP/LoxP   mice. Deletion of Ikbkap from the neural crest lineage induces a caspase-3, p53-dependent cell loss with the pups dying within postnatal day 1, while mice with a deletion of Ikbkap in postmitotic neurons of the PNS and CNS live on average 6 months. These lines were obtained by crossing mice homozygous for a floxed allele of Ikbkap (exon 4) mice to mice that are heterozygous for this floxed allele, and either (1) a Wnt1-Cre transgene or (2) a Tuba1a-Cre transgene. Dorsal root ganglia were removed from E17-18 Wnt1-Cre;IkbkapLoxP/LoxP embryos and brains (cerebellum and brainstem) removed from adult Tuba1a-cre;IkbkapLoxP/LoxP  mice and stored at -80 until RNA was isolated.

tag: developmental stage

value: Embryonic day 17.5

value: PNS

tag: region

value: Dorsal root ganglia


title: Cerebellum_WT_replicate_1

description: Sample name: WT-14626-CER


title: Cortex_WT_replicate_3

description: Sample name: WT-14628-CTX


title: primary_tumor_UPPL1598


title: primary_tumor_UPPL1694


title: primary_tumor_UPPL1595


title: primary_tumor_UPPL1597


title: primary_tumor_UPPL1612


title: Ikbkap_DRG_CKO_2


title: primary_tumor_UPPL1591


title: Hippocampus_WT_replicate_2

description: Sample name: WT-14627-HIPP


title: Hippocampus_WT_replicate_4

description: Sample name: WT-14629-HIPP


title: primary_tumor_UPPL1544


title: Hippocampus_WT_replicate_3

description: Sample name: WT-14628-HIPP


title: Ikbkap_DRG_CKO_1


title: Ikbkap_DRG_CTR4_REPEAT


title: primary_tumor_UPPL1541


title: Hippocampus_Bhlhe40KO_replicate_1

description: Sample name: KO-14097-HIPP


title: Ikbkap_DRG_CTR1


title: Hippocampus_Bhlhe40KO_replicate_2

description: Sample name: KO-14098-HIPP


title: Ikbkap_DRG_CTR4


title: Cerebellum_WT_replicate_2

description: Sample name: WT-14628-CER


title: Ikbkap_DRG_CTR2


title: primary_tumor_BBN976

value: Ctrl

value: BBN


title: primary_tumor_BBN974


title: primary_tumor_BBN975


title: primary_tumor_BBN968


title: primary_tumor_BBN973


title: primary_tumor_BBN969


title: primary_tumor_BBN972


title: cell_line_UPPL1541

source_name: cell line


title: cell_line_KT1975

value: KT


title: primary_tumor_BBN963


title: cell_line_KT1970


title: Ikbkap_DRG_CTR3


title: primary_tumor_BBN965


title: cell_line_KT1044


title: cell_line_MB49


title: Edar-HaCat with BRG1 siRNA_replicate_3

value: Edar-HaCaT

value: keratinocyte cell line

value: Stable Edar expression


title: primary_tumor_BBN962


title: cell_line_derived_tumor_UPPL1541_3


title: cell_line_derived_tumor_UPPL1541_4


title: cell_line_derived_tumor_UPPL1541_2


title: cell_line_derived_tumor_MB49_4


title: cell_line_derived_tumor_UPPL1541_1


title: cell_line_derived_tumor_MB49_2


title: cell_line_derived_tumor_MB49_3


title: cell_line_derived_tumor_BBN963_2


title: cell_line_derived_tumor_MB49_1


title: cell_line_derived_tumor_BBN963_1


title: primary_tumor_BBN964


title: cell_line_BBN963


title: Cortical_Astrocyte_Klotho 1nM_replicate4

status: Public on Mar 08 2018

growth_protocol: Primary cortical astrocyte cell cultures were prepared from newborn C57BL/6 mice (postnatal days 0-5) by dissecting cortices from newborn pups in ice-cold Hanks' balanced salt solution (HBSS) under a microscope and removing their meninges. Cortices were then cut into small pieces and incubated in a 0.05% trypsin solution (GIBCO) for 20 minutes in a 37oC, 5% CO2 incubator. Cortical tissue was then transferred to a conical tube containing DMEM (GIBCO) + 10% serum (Hyclone FetalClone III Serum, GE Healthcare), and underwent mechanical trituration with a glass pipette. One milion viable cells were plated in T75 flasks and allowed to grow in DMEM + 10% serum until approximately 90% confluent. To obtain astrocyte-pure cell cultures (>98%), flasks were incubated in an orbital shaker at 37oC, 180 rpm, for 15 hours. Primary cortical neuronal cultures were prepared using cortices collected from E16 C57BL/6 embryos.  Cortices were cut into small pieces and incubated in trypsin solution (2 mg/mL) for 20 minutes in a 37oC, 5% CO2 incubator.  After removal of   the trypsin solution, the tissue was washed twice with HBSS, incubated in a trypsin inhibitor solution (2 mg/mL) for one minute, and rinsed with HBSS.Tissue was dissociated in HBSS containing 0.1 mg/mL DNAse by mechanical trituration with a glass pipette. Cells were counted and plated (one hundred thousand cells/cm2) in polyethyleneimine (Sigma-Aldrich) pre-coated dishes. Neurons were maintained for two weeks in Neurobasal (GIBCO) supplemented with B27 (GIBCO), 2 mM glutamine, 100 U/mL penicillin, 100 µg/mL streptomycin and 0.25 µg/mL amphotericin B. This research was approved by the National Institute on Aging Animal Care and Use Committee, and was performed according to guidelines in the NIH Guide for the Care and Use of Laboratory Animals.

extract_protocol: RNA extraction was performed using Qiagen RNeasy Mini Kit (Qiagen, Inc., Valencia, CA, USA), as per the manufacturers instructions.

treatment_protocol: Primary cortical astrocytic cell cultures were treated for 24 hours with 1xPBS vehicle (HGC samples, n=4) or 1nM  recombinant secreted Klotho (HGK samples, n=4) before harvesting and RNA preparation. Primary C57BL/6  cortical neuronal cultures were treated for 24 hrs at DIV 14 with either 1xPBS vehicle (CTL, n=4) or 1 nM recombinant secreted Klotho (KLO, n=4).

tag: strain


title: Cortical_Astrocyte_control_replicate1

value: Primary cortical astrocytic cell cultures


title: Cortical_Neuron_control_replicate2

value: Primary  cortical neuronal cultures


title: Cortical_Neuron_Klotho 1nM_replicate2


title: Islets_CB1_KO_replicate2-2

status: Public on Mar 07 2018

growth_protocol: All animal procedures and care followed US National Institute of Health guidelines and were approved by the National Institute on Aging Animal Care and Use Committee. Mice were housed in groups of 4 using 12 hrs dark/light cycles, provided with water and fed ad libitum. Male mice were fed standard diet (SD; 16.7% kcal fat and 12.4% kcal sugar). Global CB1 receptor knockout (CB1KO) mice backcrossed to a C57Bl/6J background.

extract_protocol: Total RNA was isolated from islets using the TRIzol reagent according to the manufacturers instructions, and RNA concentration and quality was evaluated by Nanodrop (ThermoFisher, Waltham, MA USA) and the Agilent Bioanalyzer RNA 6000 Chip (Agilent, Santa Clara, CA).

treatment_protocol: Islets were isolated by perfusing the pancreas with 0.7 mg/ml collagenase P containing DNAse I in HBSS without calcium and magnesium immediately following sacrifice of the mice. Pancreata were incubated 9 min at 37°C with gentle agitatio, and the reaction was blocked with cold HBSS containing 1% horse serum. Disaggregated pancreata were spun and washed with cold HBSS, and islets were then handpicked.

value: islets

tag: genotype/variation

value: CB1 KO


title: Cortical_Astrocyte_Klotho 1nM_replicate2


title: Cortical_Astrocyte_control_replicate2


title: Cortical_Neuron_control_replicate4


title: Cortical_Astrocyte_control_replicate3


title: Cortical_Astrocyte_Klotho 1nM_replicate1


title: Islets_CB1_KO_replicate2-1

source_name: KO-ISL6 REPEAT


title: Islets_WT_replicate4

value: CB1 WT


title: Cortical_Neuron_Klotho 1nM_replicate1


title: Cortical_Neuron_Klotho 1nM_replicate4


title: Cortical_Neuron_Klotho 1nM_replicate3


title: Cortical_Neuron_control_replicate1


title: Cortical_Astrocyte_Klotho 1nM_replicate3


title: Cortical_Astrocyte_control_replicate4


title: Islets_CB1_KO_replicate3


title: Cortical_Neuron_control_replicate3


title: Hippocampus_ADP_NR_replicate_4

status: Public on Feb 26 2018

growth_protocol: All mice were maintained on a standard NIH diet in a 12 h light/dark cycle.  The animals were group housed if possible.  All mice were housed in the National Institute on Aging (Baltimore, MD, USA).  All animal experiments were performed using protocols approved by the appropriate institutional animal care and use committee of the National Institute on Aging.  The original AD line was generated as described previously.  The breeding methods of Polb and AD/Polb strains were described previously.  Both male and female adult mice were used in the experiments with littermate controls used throughout.  All experiments were performed blinded using only mouse identification numbers.

extract_protocol: RNA was extracted from cortex and hippocampus using Nucleospin RNA isolation kit (Macherey-Nagel, #740955.250) following the manufacturer's protocol.  RNA concentration and quality was evaluated using Nanodrop (ThermoFisher, Waltham, MA USA) and the Agilent Bioanalyzer RNA 6000 Chip (Agilent, Santa Clara, CA).

treatment_protocol: 16-18 months old WT, Polb, AD and AD/Polb mice were given 12 mM NR in their drinking water ad libitum while the control groups received drinking water without NR.  Dosing with NR continued for 3-months prior to the start of and throughout behavioral tests.  Water bottles were changed twice a week.

value: Brain

value: Hippocampus

value: nicotinamide riboside _12 mM_6 mo

tag: age

value: 23 months

tag: Sex

value: M


title: Islets_WT_replicate3


title: Hippocampus_ADP_NR_replicate_1

value: 22 months

value: F


title: Islets_CB1_KO_replicate1


title: Islets_WT_replicate1


title: Hippocampus_ADP_Vehicle_replicate_3

value: Control

value: 24 months


title: Hippocampus_ADP_NR_replicate_3


title: Hippocampus_ADP_NR_replicate_2


title: Islets_WT_replicate2


title: Hippocampus_ADP_Vehicle_replicate_4


title: Hippocampus_PolB_NR_replicate_1

value: Polb+/-


title: Hippocampus_WT_vehicle_replicate_3

value: WT


title: Hippocampus_WT_NR_replicate_3


title: Hippocampus_PolB_NR_replicate_3


title: Hippocampus_PolB_NR_replicate_4


title: Hippocampus_AD_vehicle_replicate_4


title: Cortex_ADP_NR_replicate_4

value: Cortex


title: Hippocampus_ADP_Vehicle_replicate_1


title: Hippocampus_AD_vehicle_replicate_3


title: Hippocampus_PolB_Vehicle_replicate_4


title: Hippocampus_AD_NR_replicate_3


title: Cortex_ADP_NR_replicate_3


title: Hippocampus_WT_vehicle_replicate_4


title: Hippocampus_PolB_Vehicle_replicate_1


title: Hippocampus_AD_NR_replicate_4


title: Hippocampus_WT_NR_replicate_4


title: Hippocampus_ADP_Vehicle_replicate_2


title: Hippocampus_PolB_NR_replicate_2


title: Hippocampus_PolB_Vehicle_replicate_3


title: Hippocampus_WT_vehicle_replicate_1


title: Hippocampus_AD_vehicle_replicate_1


title: Hippocampus_PolB_Vehicle_replicate_2


title: Hippocampus_AD_NR_replicate_1


title: Hippocampus_AD_NR_replicate_2


title: Hippocampus_WT_NR_replicate_1


title: Hippocampus_AD_vehicle_replicate_2


title: Cortex_ADP_NR_replicate_2


title: Hippocampus_WT_NR_replicate_2


title: Hippocampus_WT_vehicle_replicate_2


title: Cortex_ADP_NR_replicate_1


title: Cortex_ADP_Vehicle_replicate_1


title: Cortex_ADP_Vehicle_replicate_3


title: Cortex_AD_NR_replicate_3


title: Cortex_AD_NR_replicate_2


title: Cortex_AD_vehicle_replicate_2


title: Cortex_PolB_Vehicle_replicate_1


title: Cortex_ADP_Vehicle_replicate_2


title: Cortex_ADP_Vehicle_replicate_4


title: Cortex_AD_NR_replicate_1


title: Cortex_PolB_Vehicle_replicate_2


title: Cortex_PolB_NR_replicate_1


title: Cortex_PolB_Vehicle_replicate_3


title: Cortex_PolB_Vehicle_replicate_4


title: Cortex_PolB_NR_replicate_3


title: Cortex_AD_vehicle_replicate_1


title: Cortex_WT_vehicle_replicate_3


title: Cortex_PolB_NR_replicate_2


title: Cortex_PolB_NR_replicate_4


title: Cortex_AD_NR_replicate_4


title: Cortex_AD_vehicle_replicate_4


title: Cortex_AD_vehicle_replicate_3


title: Cortex_WT_NR_replicate_2


title: Cortex_WT_vehicle_replicate_2


title: Cortex_WT_NR_replicate_1


title: Cortex_WT_vehicle_replicate_4


title: Cortex_WT_NR_replicate_3


title: Liver_SD+1%metformin_two consecutive weeks per month_replicate_4

status: Public on Feb 23 2018

hyb_protocol: Standard Illumina protocol. In short, a total of 0.75 μg of biotin-labeled cRNA was hybridized at 58°C for 16 hours to Illumina Mouse Ref-8 v2 Expression Bead Chip (Illumina, San Diego, CA). The arrays were rinsed, blocked, and the biotin-labeled cRNA was detected by staining with streptavidin-Cy3.

scan_protocol: Arrays were scanned at a resolution of 0.53 μm using the Illumina iScan scanner.

label_protocol: Standard Illumina protocol using Illumina TotalPrep RNA Amplification Kit (Ambion; Austin, TX, cat # IL1791) In short, 500 ng total RNA was converted into single-stranded cDNA with reverse transcriptase using an oligo-dT primer containing the T7 RNA polymerase promoter site and then copied to produce double-stranded cDNA molecules. The double stranded cDNA was cleaned and concentrated with the supplied columns and used in an overnight in-vitro transcription reaction where single-stranded RNA (cRNA) was generated and labeled by incorporation of biotin-16-UTP.

extract_protocol: Total RNA was extracted from frozen tissue using Trizol® Reagent (Thermo Fisher Scientific, Waltham, MA). Briefly, 50-100mg of liver was homogenized in 1 mL Trizol® Reagent and phase separation was carried out with 0.2 mL of chloroform. The aqueous phase was collected, RNA precipitated with 100% isopropanol. After centrifugation, the resultant pellet was rinsed twice with 75% ethanol and briefly airdried before being resuspended in RNAse-free water.  RNA quality and  quantity was evaluated using bith a NanoDrop 1000 spectrophotometer (Thermo Fisher Scientific, Waltham, MA), and the Agilent RNA 6000 Nano kit (Agilent, Santa Clara, CA).

treatment_protocol:
Animal procedures, housing and diets were in accordance with the guidelines issued by the Intramural Research Program of the National Institutes of Health protocol number 444TGB2016. Male C57BL/6 mice at 104 weeks of age were purchased from the National Institute on Aging Aged Rodent Colony from Charles River. The C57BL/6 mice maintained on a standard purified mouse diet (AIN-93G) for a month were randomly distributed in 3 experimental groups. The mice were on a light:dark 12:12-h schedule and maintained between 20–22°C with 30–70% relative humidity according to animal protocols and NIH guidelines. Food intake and body weight were measured on a biweekly basis for the duration of the study for control group and weekly for metformin fed mice. Survival curves were plotted using the Kaplan–Meier method, which includes all available animals at each time point. The criteria for euthanasia was based on an independent assessment by a veterinarian, according to AAALAC guidelines and only cases, where the condition of the animal was considered incompatible with continued survival, are represented in the curves. Animals removed at sacrifice for experimental procedures were considered as censored deaths.
Control group was kept on standard diet (SD; n=68) until their death while the other groups of mice were fed SD plus 1% metformin following different on-and-off feeding protocols. Feedings protocol consist of every other week (EOW; n=64) and two consecutive weeks each month (2WM; n=67) with AIN-93G plus 1% metformin until their death. Pure metformin was obtained from Farmhispania (Farmhispania S.A., Barcelona, Spain) and mixed to homogeneity during manufacturing of the diets (Dyets Inc., Bethlehem, PA). The chow was produced every 3 months during the length of the study, never was permitted to exceed 50°C and was kept away from light whenever possible to ensure the stability of metformin (the light/dark cycle in the mouse facility was not altered).

value: Liver

tag: mouse strain

tag: diet

value: Male


title: Cortex_WT_NR_replicate_4


title: Cortex_WT_vehicle_replicate_1


title: CBE-/- (1)/C1-/-C2-/- VDJ-Seq replicate 2

status: Public on Feb 21 2018

description: Deletion of C1, C2 site and mutation of two CTCF binding sites within intergenic control region 1 (IGCR1)

library_source: genomic

data_processing:
Paired-end reads were generated by Illumina HiSeq 2500 sequencer.
Samples were separated using barcodes present in Read-1.
Adapters, if present were removed by cutadapt and bad quality bases (< Q33) were trimmed from Read-2 keeping a minimum length of 50 bases.
The reads (Read-2 only) were aligned to 600 bases of different V-regions using bowtie2
Reads which aligned to different V-regions with an alighnment quality > 20 were counted.
Genome_build: mm9 IgH V-regions
Supplementary_files_format_and_content: The processed data file contain the read counts of the different V-regions in a tab-delimited TXT format.

molecule: genomic DNA

source_name: Pro-B cell line

growth_protocol: Pro-B cell lines were grown with complete 1640 RPMI medium (10% fetal bovine serum and 0.05mM 2-Mercaptoethanol)

extract_protocol:
DNA were extracted with  DNeasy Blood & Tissue Kit (#69506, Qiagen)
libraries were prepared for sequencing using standard Illumina protocols.
VH utilization Seq. Total reads in each cell line were aligned to 128 VH genes in 129 strain mice

treatment_protocol: All cell lines were infected with RAG2-expressing lentivirus to assay recombination.  After 10 days selection with puromycin, genomic DNA was prepared to analyze VH utilization by deep sequencing .

value: Pro-B cell line

value: Bone marrow


title: CBE-/- (1)/C1-/-C2-/- VDJ-Seq replicate 1


title: CBE-/- (1) VDJ-Seq replicate 3

description: Mutation of two CTCF binding sites within intergenic control region 1 (IGCR1)


title: CBE-/- (1) VDJ-Seq replicate 2


title: CBE-/- (1) VDJ-Seq replicate 1


title: Liver_SD+1%metformin_two consecutive weeks per month_replicate_3


title: CBE-/- (2) VDJ-Seq replicate 2


title: Liver_SD+1%metformin_two consecutive weeks per month_replicate_2


title: Liver_SD+1%metformin_two consecutive weeks per month_replicate_1


title: CBE-/- (2) VDJ-Seq replicate 1


title: Liver_SD+1%metformin_every other week_replicate_5

source_name: Liver_SD+1%metformin_every other week

value: SD+1%metformin_every other week


title: Liver_SD+1%metformin_every other week_replicate_4


title: Liver_SD+1%metformin_every other week_replicate_1


title: Liver_SD+1%metformin_every other week_replicate_3


title: CBE-/- (1)/C2-/- VDJ-Seq replicate 1

description: Deletion of C2 site and mutation of two CTCF binding sites within intergenic control region 1 (IGCR1)


title: Liver_SD+1%metformin_two consecutive weeks per month_1 Technical replicate


title: CBE-/- (1)/C1-/- VDJ-Seq replicate 2

description: Deletion of C1 site and mutation of two CTCF binding sites within intergenic control region 1 (IGCR1)


title: Liver_Standard diet (SD)_replicate_4

source_name: Liver_Standard diet

value: Standard AIN-93G diet


title: Liver_Standard diet (SD)_replicate_1_Technical replicate


title: Liver_Standard diet (SD)_replicate_3


title: Liver_Standard diet (SD)_replicate_1


title: Liver_SD+1%metformin_every other week_replicate_2


title: CBE-/- (1)/C2-/- VDJ-Seq replicate 2


title: CBE-/- (1)/C1-/- VDJ-Seq replicate 1


title: Liver_SD+1%metformin_every other week_replicate_1 Technical replicate


title: L2_hypoxia_replicate_1

status: Public on Jan 23 2018

scan_protocol: Arrays were scanned at a resolution of 0.54 μm using the Illumina iScan scanner.

growth_protocol: SUDHL-2 cells (L2) were obtained from American type culture collection (ATCC), while HLY-1 cells (HLY) were a generous gift from Dr. Lisa Rimsza (University of Arizona, AZ), and Dr. Ari Melnick (Weill Cornell, NY), and were grown in RPMI-1640 media supplemented with 100 units/ml penicillin and 0.1mg/ml streptomycin and 10% FBS.

extract_protocol: RNA was extracted using Trizol (Trizol Reagent, Invitrogen) according to manufacturer’s instructions. RNA was evaluated by Nanodrop and RNA concentration and integrity was assessed by Agilent Bioanalyzer RNA 6000 Chip (Agilent, Santa Clara, CA).

treatment_protocol: Cells were cultured under normoxia (21% oxygen) and hypoxia (1% oxygen) in the presence of 5% CO2.

tag: cell line

value: Human lymphoma cell line SUDHL2

value: Hypoxia


title: Liver_Standard diet (SD)_replicate_2


title: L2_hypoxia_replicate_2


title: HLY_normoxia_replicate_1

value: Human lymphoma cell line HLY-1

value: Normoxia


title: L2_normoxia_replicate_3


title: L2_normoxia_replicate_1


title: HLY_normoxia_replicate_2


title: HLY_normoxia_replicate_3


title: Vastus lateralis_Young_replicate2

status: Public on Jan 22 2018

hyb_protocol: Standard Illumina protocol. In short, a total of 0.75 μg of biotin-labeled cRNA was hybridized at 58°C for 16 hours to Illumina's HumanHT-12 V4.0 Expression Bead Chips (Illumina, San Diego, CA).The arrays were rinsed, blocked, and the biotin-labeled aRNA was visualized by staining with streptavidin-Cy3.

source_name: Vastus lateralis_Young

extract_protocol: Total RNA was extracted with Trizol® (Invitrogen) from vastus lateralis muscle of 24 of the 38 participants (age 25-84 years; mean 67 ± 15) according to manufacturer’s instructions. RNA quantity and integrity was evaluated by Agilent Bioanalyzer RNA 6000 Chip (Agilent, Santa Clara, CA).

treatment_protocol: The Baltimore Longitudinal Study on Aging (BLSA) is a study of human aging established in 1958 and currently supported by the National Institute on Aging (NIA). Adult volunteers age 20 years and older are recruited from the community and healthy at study entry. BLSA participants come in for study visits at age-specific intervals, undergo medical and various physiological testings at each visits, and are followed for life. The BLSA protocol has been approved by the Intramural Research Program of the NIA and the Insitutional Review Board of the National Institute of Environmental health Sciences. The percutaneous muscle biopsy was recently introduced in BLSA. Between August 2013 and June 2016, muscle biopsy specimens were obtained in 41 participants along with measures of mitochondrial respiration by high resolution respirometry immediately after the biopsy. Of these, 38 (21 men), aged 24-91 years (mean age 68.6 ± 16.8 years) had complete data from a physical examination, whole-body aerobic capacity (VO2max), body composition assessment, gait speed in different tasks and muscle function.

value: Young


title: L2_hypoxia_replicate_3


title: HLY_hypoxia_replicate_1


title: Vastus lateralis_Young_replicate3

value: Female


title: HLY_hypoxia_replicate_3


title: Vastus lateralis_Young_replicate4


title: Vastus lateralis_Old_replicate11

source_name: Vastus lateralis_Old

value: Old


title: HLY_hypoxia_replicate_2


title: Vastus lateralis_Old_replicate13


title: L2_normoxia_replicate_2


title: Vastus lateralis_Young_replicate5


title: Vastus lateralis_Old_replicate13_repeat


title: Vastus lateralis_Old_replicate12


title: Vastus lateralis_Old_replicate8


title: Vastus lateralis_Young_replicate1


title: Vastus lateralis_Old_replicate10


title: Vastus lateralis_Old_replicate9


title: Vastus lateralis_Old_replicate7


title: Vastus lateralis_Old_replicate6


title: Vastus lateralis_Old_replicate4


title: Vastus lateralis_Old_replicate3


title: DHTS_HuR_replicate1

status: Public on Dec 20 2017

data_processing:
Data was extracted using the Illumina GenomeStudio software V2011.1, Gene expression module 1.9.0. Any spots at or below the background were filtered out using an Illumina detection Pvalue of 0.02 and above. The natural log of all remaining scores were used to find the avg and std of each array and the z-score normalization was calculated and presented below.  Z-score = (raw value - avg)/std.  Complete data including detection scores is supplied.
Z_VALUE = Z transformation of the natural log of the raw intensity values
Detection_Pval = Detection Pvalue from Illumina GenomeStudio software V2011.1, Gene expression module 1.9.0.

source_name: HeLa_DHTP_HuR IP

label_protocol: Standard Illumina protocol using Illumina TotalPrep RNA Amplification Kit (Ambion; Austin, TX, cat # IL1791), to convert total RNA into single-stranded cDNA with reverse transcriptase using an oligo-dT primer containing the T7 RNA polymerase promoter site and then copied to produce double-stranded cDNA molecules. The double stranded cDNA was cleaned and concentrated with the supplied columns and used in an overnight in-vitro transcription reaction where single-stranded RNA (cRNA) was generated and labeled by incorporation of biotin-16-UTP.

extract_protocol: RNA was extracted using phenol-chloroform and precipitated.

treatment_protocol: HeLa cells were exposed to either 1 micromolar (1 uM) DHTS or DMSO vehicle for 3 hours, followed by immunoprecipitation (IP) of endogenous ribonucleoprotein complexes. HeLa cells were lysed in 20 mM Tris-HCl at pH 7.5, 100 mM KCl, 5 mM MgCl2, and 0.5% NP-40 for 10 min on ice and centrifuged at 15,000 × g for 10 min at 4°C. Supernatants were incubated for 2 h at 4°C with protein G Sepharose beads (GE Healthcare) coated either with anti-HuR or with control IgG antibodies (both from Santa Cruz Biotechnology). The beads were washed with NT2 buffer (50 mM Tris-HCl [pH 7.5], 150 mM NaCl, 1 mM MgCl2, 0.05% NP-40), followed by incubation with 20 units of RNase-free DNase I for 15 min at 37°C to remove the DNA. The samples were then incubated for 15 min at 55°C with 0.1% SDS–0.5 mg/ml proteinase K to digest proteins. The RNA from the IP samples was extracted using phenol-chloroform, precipitated, and used for cDNA microarray analysis or q-RT-PCR validation.

value: HeLa

value: cervical

tag: disease state

value: carcinoma

tag: exposed to

value: 1 uM DHTS for 3 h


title: Vastus lateralis_middle-aged_replicate3

source_name: Vastus lateralis_middle-aged

value: Middle-aged

tag: gender


title: Vastus lateralis_Old_replicate2


title: DHTS_HuR_replicate2

tag: tissue


title: Vehicle_IgG_replicate1

source_name: HeLa_Vehicle_IgG IP

value: 1 uM DMSO vehicle for 3 h


title: Vehicle_HuR_replicate1

source_name: HeLa_Vehicle_HuR IP


title: atm1_Nicotinamide riboside_Day10_Replicate 2

description:
ATM1_NR_DAY10_2
Included in analysis.

hyb_protocol: A total of 1.65 micrograms Cy3-labeled cRNA (specific activity >6 pmol Cy3/ug cRNA) was fragmented at 60°C for 30 minutes in a reaction volume of 55 microL containing 0.5x Agilent fragmentation buffer and 2x Agilent gene expression blocking agent following the manufacturer's instructions. On completion of the fragmentation reaction, 55 microL of 2x Hi-RPM Hybridization Buffer was added to the fragmentation mixture and hybridized to Agilent-020186 C. elegans (V2) 4x44K Gene Expression Microarrays (G2519F-020186) 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), and dried by slowly removing from Wash buffer 2.

scan_protocol: Slides were scanned immediately after washing on the Agilent SureScan Microarray Scanner using one color scan setting for 4x44k array slides (Scan resolution 5 microns, Dye channel: Green and Green PMT: 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.

organism: Caenorhabditis elegans

source_name: atm1 worm, NR-treated, day10

label_protocol: Cyanine-3 (Cy3) labeled cRNA was prepared from 300 ng total RNA using the Low Input Quick Amp Labeling Kit (Agilent, Santa Clara, CA) according to the manufacturer's instructions, followed by RNeasy column purification (QIAGEN, Valencia, CA). Dye incorporation and cRNA yield were evaluated with the NanoDrop ND-1000 Spectrophotometer (Wilmington, DE).

growth_protocol: C. elegans worms from a wild-type strain (N2) and atm1 mutated (atm1) were cultured in the presence of 5-Fluoro-deoxyuridine (5-FdUrd) to prevent development of progeny and produce a synchronized population.

extract_protocol: Worms were washed off the plates with M9 buffer from 4 independent experiments and stored at -80 oC in lysis reagent (Qiagen Cat. no. 75142). Crude extracts were prepared by bead beating packed worm pellets with a Mini-Beadbeater 8 (Biospec Products). Total RNA was isolated using RNeasy Mini Kit (Qiagen, Cat.74106) on the QIAcube according to the manufacturer's protocol. RNA integrity and quantity were evaluated using the Agilent Bioanalyzer RNA 6000 chip (Agilent).

treatment_protocol: L4-stage N2 (wild-type) and atm-1 mutated (atm1) C. elegans worms were treated with NAD+ precursor nicotinamide riboside (NR), SIRT1 inhibitor SRT1720 or PARP1 inhibitor Olaparib for 1 or 10 days.

tag: strain/background

value: nicotinamide riboside

tag: concentration

value: 500 microM

tag: days after treatment


title: Vastus lateralis_Old_replicate1

value: Male


title: atm_Olaparib_Day10_Replicate 1

description:
ATM1_OLAP_DAY10_1
Included in analysis.

source_name: atm1 worm, Olaparib-treated, day10

tag: treatment

value: Olaparib


title: atm_Olaparib_Day10_Replicate 3

description:
ATM1_OLAP_DAY10_3
Included in analysis.


title: Vehicle_IgG_replicate2


title: DHTS_IgG_replicate2

source_name: HeLa_DHTP_IgG IP


title: Vastus lateralis_middle-aged_replicate1


title: atm_Olaparib_Day10_Replicate 2

description:
ATM1_OLAP_DAY10_2
Included in analysis.


title: DHTS_IgG_replicate1


title: Vastus lateralis_middle-aged_replicate4


title: Vehicle_HuR_replicate2


title: Vastus lateralis_middle-aged_replicate2


title: atm_Sirtuin1720_Day10_Replicate 2

description:
ATM1_SRT1720_DAY10_2
Included in analysis.

source_name: atm1 worm, SIRT1720-treated, day10

value: 10 microM


title: atm_Sirtuin1720_Day10_Replicate 3

description:
ATM1_SRT1720_DAY10_3
Included in analysis.


title: Vastus lateralis_middle-aged_replicate5


title: atm_Sirtuin1720_Day10_Replicate 1

description:
ATM1_SRT1720_DAY10_1
Included in analysis.


title: Vastus lateralis_Old_replicate5


title: atm1_Nicotinamide riboside_Day10_Replicate 1

description:
ATM1_NR_DAY10_1
Included in analysis.


title: atm1_CTR_Day10_Replicate 3

description:
ATM1_CON_DAY10_3
Included in analysis.

source_name: atm1 worm, untreated, day10


title: atm1_CTR_Day10_Replicate 2

description:
ATM1_CON_DAY10_2
Included in analysis.


title: WT_Olaparib_Day10_Replicate 2

description:
N2_OLAPARIB_DAY10_3
Excluded from analysis.

source_name: WT worm, Olaparib-treated, day10

value: WT


title: WT_Nicotinamide riboside_Day10_Replicate 2

description:
N2_NR_DAY10_3
Included in analysis.

source_name: WT worm, NR-treated, day10


title: WT_Sirtuin1720_Day10_Replicate 3

description:
N2_SRT1720_DAY10_3
Included in analysis.

source_name: WT worm, SIRT1720-treated, day10


title: atm1_CTR_Day10_Replicate 1

description:
ATM1_CON_DAY10_1
Included in analysis.


title: WT_Nicotinamide riboside_Day10_Replicate 1

description:
N2_NR_DAY10_2
Included in analysis.


title: atm_Olaparib_Day1_Replicate 3

description:
ATM1_OLAP_DAY1_3
Included in analysis.

source_name: atm1 worm, Olaparib-treated, day1


title: atm1_Nicotinamide riboside_Day10_Replicate 3

description:
ATM1_NR_DAY10_3
Included in analysis.


title: atm_Olaparib_Day1_Replicate 2

description:
ATM1_OLAP_DAY1_2
Included in analysis.


title: WT_Sirtuin1720_Day10_Replicate 2

description:
N2_SRT1720_DAY10_2
Included in analysis.


title: atm_Olaparib_Day1_Replicate 1

description:
ATM1_OLAP_DAY1_1
Included in analysis.


title: atm1_Nicotinamide riboside_Day1_Replicate 3

description:
ATM1_NR_DAY1_3
Excluded from analysis.

source_name: atm1 worm, NR-treated, day1


title: atm1_Nicotinamide riboside_Day1_Replicate 1

description:
ATM1_NR_DAY1_1
Included in analysis.


title: atm_Sirtuin1720_Day1_Replicate 1

description:
ATM1_SRT1720_DAY1_1
Included in analysis.

source_name: atm1 worm, SIRT1720-treated, day1


title: WT_CTR_Day10_Replicate 1

description:
N2_CON_DAY10_1
Included in analysis.

source_name: WT worm, untreated, day10


title: WT_Olaparib_Day10_Replicate 1

description:
N2_OLAPARIB_DAY10_2
Included in analysis.


title: atm_Sirtuin1720_Day1_Replicate 2

description:
ATM1_SRT1720_DAY1_2
Included in analysis.


title: atm_Sirtuin1720_Day1_Replicate 3

description:
ATM1_SRT1720_DAY1_3
Included in analysis.


title: WT_Olaparib_Day10_Replicate 3

description:
N2_OLAPARIB_DAY10_4
Included in analysis.


title: WT_CTR_Day10_Replicate 3

description:
N2_CON_DAY10_3
Included in analysis.


title: WT_Nicotinamide riboside_Day10_Replicate 3

description:
N2_NR_DAY10_4
Included in analysis.


title: atm1_CTR_Day1_Replicate 3

description:
ATM1_CON_DAY1_3
Included in analysis.

source_name: atm1 worm, untreated, day1


title: WT_Sirtuin1720_Day10_Replicate 1

description:
N2_SRT1720_DAY10_1
Included in analysis.


title: atm1_CTR_Day1_Replicate 2

description:
ATM1_CON_DAY1_2
Included in analysis.


title: miRNA T. muris exosomes rep 2

status: Public on Dec 13 2017

city: Smithfield

first: Javier

last: Sotillo

address: 14-88 McGregor rd

department: AITHM

country: Australia

institute: James Cook University

description: small RNA

data_processing:
Adapters were  filtered using cutadapt software with Illumina adaptor sequence TGGAATTCTCGGGTGCCAAGG as input
Reads were aligned to the trichuris genome using the mapper module of mirdeep2 which uses bowtie2 with default options that collapse duplicate reads and map reads without any mismatches
mirdeep2 was used to identify known and putative novel miRNA using five closest species to generate comparative mature miRNA sequences (Schmidtea mediterranea, Echinococcus granulosus and Echinococcus multilocularis, Schistosoma japonicum and S. mansoni)
Genome_build: Assembly= TMUE2.2; bioproject= PRJEB126
Supplementary_files_format_and_content: bed files were generated

organism: Trichuris muris

source_name: Extracellular vesicles

growth_protocol: Adult worms were harvested from the caecum of infected mice 5 weeks after infection, washed in PBS containing 5× antibiotic/antimycotic (AA) and cultured in 6-well plates for 5 days in RPMI containing 1× AA, at 37°C and 5% CO2.

extract_protocol:
Two different biological replicates of EVs obtained from two different batches of worms were used. ODG fractions with a density between 1.06 and 1.08 (fractions containing pure EV samples after TRPS analysis) were pooled and excess Optiprep® solution was removed by buffer exchanging. miRNA were extracted using the mirVana miRNA Isolation Kit (ThermoFischer) according to the manufacturer’s instructions.
miRNA was prepared for sequencing using Illumina TruSeq Small RNA-seq library preparation kit according to the manufacturer’s instructions.

treatment_protocol: The media obtained during the first 4 h after parasite culturing was discarded for further analysis. Dead worms were removed and ES products were collected daily, subjected to sequential differential centrifugation at 500 g, 2000 g and 4000 g for 30 min each to remove eggs and parasite debris. For the isolation of ES products, media was concentrated using a 10 kDa spin concentrator and stored at 1.0 mg/ml in PBS at -80°C until required. For the isolation of EVs, the media obtained after differential centrifugation was processed as described previously. Briefly, ES products were concentrated using a 10 kDa spin concentrator, followed by centrifugation for 45 min at 12,000 g to remove larger vesicles. A MLS-50 rotor (Beckman Coulter, USA) was used to ultracentrifuge the supernatant for 3 h at 120,000 g and the resultant pellet was resuspended in 70 μl of PBS and subjected to Optiprep® discontinous gradient (ODG) separation. One mL of 40%, 20%, 10% and 5% iodixanol solutions prepared in 0.25 M sucrose, 10 mM Tris-HCl, pH 7.2, was layered in decreasing density in an ultracentrifuge tube, and the 70 µl containing the resuspended EVs was added to the top layer and ultracentrifuged at 120,000 g for 18 h at 4°C. Seventy (70) µl of PBS was added to the control tube prepared as described above. A total of 12 fractions were recovered from the ODG, and the excess Optiprep® solution was removed by buffer exchanging with 8 ml of PBS containing 1× EDTA-free protease inhibitor cocktail (Santa Cruz, USA) using a 10 kDa spin concentrator.

tag: developmental stage

value: Adult worms

tag: vesicle type

value: Exosome-like vesicles


title: WT_Olaparib_Day1_Replicate 3

description:
N2_OLAPARIB_DAY1_3
Included in analysis.

source_name: WT worm, Olaparib-treated, day1


title: WT_Olaparib_Day1_Replicate 2

description:
N2_OLAPARIB_DAY1_2
Included in analysis.


title: PHH 0d

status: Public on Nov 23 2017

city: Shanghai

first: Xiaoran

last: zhang

address: Yueyang Road 320

department: Shanghai Institutes for Biological Sciences

country: China

institute: Chinese Academy of Sciences

data_processing:
Basecalling using Illumina Casava1.8.2 software
Map to human genome hg19 using Tophat(v 2.0.13) and
Bam to Bedgraph using genomeCoverageBed,Bedgraph to bigwig using UCSC bedGraphToBigWig
Counted the mapping reads and normalized as FPKM values using Cufflinks(v.2.2.1).
Differential expression analysis was carried out using the R (v.3.1.1) DEseq (v.1.18.0)
Genome_build: hg19
Supplementary_files_format_and_content: bigwig file (UCSC bigwig in an indexed binary format. http://genome.ucsc.edu/goldenPath/help/bigWig.html)

source_name: liver tissue

growth_protocol: UCF were maintained in HFM, hiHeps were maintained in HMM, hiPSC-Heps were differentiated from hiPSC

extract_protocol:
RNA was harvested using Trizol reagent.
Libraries were prepared for sequencing using standard Illumina protocols

treatment_protocol: Cells were maintained in their own medium and cultured under 37℃ in the incubator

tag: cell type

value: Hepatocyte

value: liver


title: PHH 2d


title: PHH 1d


title: hiHep rep4

source_name: human induced hepatocyte-like cells

value: human induced hepatocyte-like cells


title: atm1_CTR_Day1_Replicate 1

description:
ATM1_CON_DAY1_1
Included in analysis.

tag: genotype/variation


title: hiHep rep3


title: WT_Sirtuin1720_Day1_Replicate 3

description:
N2_SRT1720_DAY1_3
Included in analysis.

source_name: WT worm, SIRT1720-treated, day1


title: WT_Sirtuin1720_Day1_Replicate 2

description:
N2_SRT1720_DAY1_2
Included in analysis.


title: WT_Olaparib_Day1_Replicate 1

description:
N2_OLAPARIB_DAY1_1
Included in analysis.


title: WT_CTR_Day10_Replicate 2

description:
N2_CON_DAY10_2
Included in analysis.


title: hiHep rep2


title: hiHep rep1


title: WT_Nicotinamide riboside_Day1_Replicate 3

description:
N2_NR_DAY1_3
Included in analysis.

source_name: WT worm, NR-treated, day1


title: WT_Sirtuin1720_Day1_Replicate 1

description:
N2_SRT1720_DAY1_1
Included in analysis.


title: WT_CTR_Day1_Replicate 1

description:
N2_CON_DAY1_1
Included in analysis.

source_name: WT worm, untreated, day1


title: WT_CTR_Day1_Replicate 3

description:
N2_CON_DAY1_3
Included in analysis.


title: WT_CTR_Day1_Replicate 2

description:
N2_CON_DAY1_2
Included in analysis.


title: WT_Nicotinamide riboside_Day1_Replicate 2

description:
N2_NR_DAY1_2
Included in analysis.


title: hiPSC-Hep rep3

source_name: hiPSC derived hepatocyte-like cells

value: hiPSC derived hepatocyte-like cells


title: hiPSC-Hep rep4


title: miRNA T. muris exosomes rep 1

description: small RNA


title: hiPSC-Hep rep2


title: WT_Nicotinamide riboside_Day1_Replicate 1

description:
N2_NR_DAY1_1
Included in analysis.


title: PHH 4d


title: hiPSC-Hep rep1


title: PHCs rep1

status: Public on Oct 17 2017

city: shanghai

first: yan

last: hexin

address: No. 1630 East Road

institute: Renji Hospital Affiliated to Shanghai Jiaotong University School of Medicine

data_processing:
Basecalls performed using CASAVA version 1.8.
Sequenced reads were trimmed for adaptor sequence, and masked for low-complexity or low-quality sequence using Seqtk.
mRNA-seq reads were aligned to the hg38 genome assembly using Hisat2(version:2.0.4).
Gene abundance measurements were generationed and normalized using HTSeq. The normalized standard were Fragments Per Kilobase of exon per Megabase of library size (FPKM).
Differential expression were analysed using edgeR.
Genome_build: hg38
Supplementary_files_format_and_content: tab-delimited text files include normalized FPKM values and raw fragment counts for each Sample

source_name: Donor_1 PHCs

extract_protocol:
Total RNA was isolated using RNeasy mini kit (Qiagen, Germany).
Pairedend libraries were synthesized by using the TruSeq® RNA Sample Preparation Kit (Illumina, USA) following TruSeq® RNA Sample Preparation Guide.

value: Primary hepatocytes

tag: culture condition

value: Uncultured

tag: ethnicity

value: Asian


title: PHCs rep2

source_name: Donor_2 PHCs


title: SAN_AMPK R299Q WT_replicate2 [RS normalized]

status: Public on Sep 30 2017

hyb_protocol: Standard Illumina protocol. In short, a total of 0.75ug of biotin-labeled cRNA was hybridized at 58 degrees C for 16 hours to Illumina's Mouse-Ref8 v2 Expression Bead Chips (Illumina, San Diego, CA). Each BeadChip has ~25,600 well-annotated RefSeq transcripts . The arrays were washed, blocked and the biotin labeled cRNA was detected by staining with streptavidin-Cy3.

scan_protocol: Arrays were scanned at a resolution of 0.8um using the Beadstation 500 X from Illumina.

data_processing: Data was extracted using the Illumina GenomeStudio software V2011.1, Gene expression module 1.9.0. Any spots at or below the background were filtered out using an Illumina detection Pvalue of 0.02 and above. Data was analyzed using two different normalization approaches. For z-score normalized data, the natural log of all remaining scores were used to find the avg and std of each array and the z-score normalization was calculated.  Z-score = (raw value - avg)/std.  For Robust Spline Normalisation, all data were log2-transformed and normalised using the lumi software package in Bioconductor. 18,185 nucleotide probes were filtered from 25,697 total using an Illumina detection P value of α = 0.05. Significant genes were selected by a one-way ANOVA or t-test (FDR = 0.05) for further analysis. GSEA software was used to select differentially expressed (DE) genes by calculating out a ‘score’ using the Kolmogorov-Smirnov (KS) test. The score indicates how a gene relates with genotype, where positive values relate to up-regulation and negative values relate to down-regulation. Only transcripts with a KS score > |0.15| were considered DE. Transcripts demonstrating greater than 1.2-fold change in expression were processed using Ingenuity Pathway Analysis software (Ingenuity Systems, CA, USA) to identify networks and canonical pathways overrepresented in enriched genes.Complete data including detection scores is supplied.

source_name: SAN_AMPK wildtype

extract_protocol: RNA was extracted with an RNeasy Mini Kit (Qiagen) using DNase on-column digestion according to the manufacturer’s protocol, and assessed for quality and quantity using an Agilent 2100 Bioanlyzer with RNA 6000 Nano Chips, (Aligent technologies).

treatment_protocol:
Experimental_protocol:Animal studies were performed in accordance with the 1986 British Home Office Animals Scientific Procedures Act (UK) incorporating European Directive 2010/63/EU, the European Directive (86/609/CEE) on the care and use of laboratory animals, and the Guide for the Care and Use of Laboratory Animals published by the National Institutes of Health (NIH Publication no. 85-23, revised 1996). All experimental protocols were assessed and approved by the local university ethical review committee. Sinoatrial node (SAN) isolation. The heart with lungs was quickly removed and immersed at 4 °C to wash out the blood in an external solution that contained (in mmol/L): NaCl, 137; KCl, 4.9; NaH2PO4, 1.2; glucose,15; HEPES, 20; MgCl2, 1.0; pH 7.4. The heart-lung block was pinned to the tissue bath to excise the right atrium (RA) and SAN under a stereomicroscope. The tissue bath was perfused with the external solution at a rate of 10 mL/min. After removal of both ventricles and the left atrium, the RA was opened to expose the crista terminalis, the intercaval area and the interatrial septum. This preparation was pinned by small stainless steel pins to the chamber with the endocardial side exposed up and trimmed carefully to extract the SAN region correctly. The SAN region was delimited by the borders of the crista terminalis, the interatrial septum, superior and inferior vena cava. All tissues were snap-frozen in liquid nitrogen and stored at -80 °C for subsequent RNA extraction.
AMPK R299Q γ2 knock-in mice  were created through gene-targeting to introduce the R299Q point mutation (equivalent to the human R302Q mutation) into exon 7 of murine Prkag2. Mice heterozygous for the R299Q γ2 knock-in mutation were initially on a mixed C57BL/6/129/Ola genetic background and subsequently backcrossed to C57BL/6 for at least 7 generations. Four mouse SAN tissues were pooled from R299Q γ2 and WT littermate controls and processed together for the extraction of one sample of total RNA for a total of 4 homozygote R299Q γ2, 4 heterozygote R299Q γ2 and 3 WT samples.

tag: strain

value: Sinoatrial node

value: Pool of 4 SAN

tag: age

value: 3 months


title: SAN_AMPK R299Q WT_replicate3 [RS normalized]


title: SAN_AMPK R299Q γ2 heterozygote_replicate1 [z-score normalized]

source_name: SAN_AMPK R299Q γ2 heterozygote

treatment_protocol:
Experimental protocol:Animal studies were performed in accordance with the 1986 British Home Office Animals Scientific Procedures Act (UK) incorporating European Directive 2010/63/EU, the European Directive (86/609/CEE) on the care and use of laboratory animals, and the Guide for the Care and Use of Laboratory Animals published by the National Institutes of Health (NIH Publication no. 85-23, revised 1996). All experimental protocols were assessed and approved by the local university ethical review committee. Sinoatrial node (SAN) isolation. The heart with lungs was quickly removed and immersed at 4 °C to wash out the blood in an external solution that contained (in mmol/L): NaCl, 137; KCl, 4.9; NaH2PO4, 1.2; glucose,15; HEPES, 20; MgCl2, 1.0; pH 7.4. The heart-lung block was pinned to the tissue bath to excise the right atrium (RA) and SAN under a stereomicroscope. The tissue bath was perfused with the external solution at a rate of 10 mL/min. After removal of both ventricles and the left atrium, the RA was opened to expose the crista terminalis, the intercaval area and the interatrial septum. This preparation was pinned by small stainless steel pins to the chamber with the endocardial side exposed up and trimmed carefully to extract the SAN region correctly. The SAN region was delimited by the borders of the crista terminalis, the interatrial septum, superior and inferior vena cava. All tissues were snap-frozen in liquid nitrogen and stored at -80 °C for subsequent RNA extraction.
AMPK R299Q γ2 knock-in mice  were created through gene-targeting to introduce the R299Q point mutation (equivalent to the human R302Q mutation) into exon 7 of murine Prkag2. Mice heterozygous for the R299Q γ2 knock-in mutation were initially on a mixed C57BL/6/129/Ola genetic background and subsequently backcrossed to C57BL/6 for at least 7 generations. Four mouse SAN tissues were pooled from R299Q γ2 and WT littermate controls and processed together for the extraction of one sample of total RNA for a total of 4 homozygote R299Q γ2, 4 heterozygote R299Q γ2 and 3 WT samples.


title: SAN_AMPK R299Q WT_replicate3 [z-score normalized]


title: SAN_AMPK R299Q γ2 homozygote_replicate1 [RS normalized]

source_name: SAN_AMPK R299Q γ2 homozygote


title: SAN_AMPK R299Q γ2 homozygote_replicate1 [z-score normalized]


title: SAN_AMPK R299Q γ2 homozygote_replicate3 [RS normalized]


title: SAN_AMPK R299Q γ2 homozygote_replicate3 [z-score normalized]


title: SAN_AMPK R299Q γ2 heterozygote_replicate3 [RS normalized]


title: SAN_AMPK R299Q γ2 heterozygote_replicate4 [z-score normalized]


title: SAN_AMPK R299Q γ2 heterozygote_replicate3 [z-score normalized]


title: SAN_AMPK R299Q γ2 homozygote_replicate2 [z-score normalized]


title: SAN_AMPK R299Q WT_replicate1 [z-score normalized]


title: SAN_AMPK R299Q γ2 homozygote_replicate4 [RS normalized]


title: SAN_AMPK R299Q γ2 heterozygote_replicate2 [z-score normalized]


title: HeLa_Cyrano-MS2 pulldown_replicate3

status: Public on Sep 25 2017

description:
Cyrano3
IP RNA

source_name: HeLa_Cyrano-MS2 pulldown

growth_protocol: Human cervical carcinoma HeLa cells were grown in Dulbecco’s modified Eagle’s medium (DMEM) containing 10% (v/v) fetal bovine serum (FBS), antibiotics and antimycotics

extract_protocol: RNA was extracted using Trizol (Trizol Reagent, Invitrogen) according to manufacturer’s instructions. RNA integrity was assessed by Agilent Bioanalyzer RNA 6000 Chip (Agilent, Santa Clara, CA).

treatment_protocol: HeLa cells were transfected with siRNA duplexes at 50 nM final concentration using Lipofectamine 2000 (Invitrogen).  Plasmids (2 µg each) were transfected using Lipofectamine 2000.

value: cervical

value: carcinoma

tag: cell line

value: HeLa

value: MS2-pulldown against lncRNA OIP5-AS1/Cyrano-MS2


title: SAN_AMPK R299Q WT_replicate2 [z-score normalized]


title: SAN_AMPK R299Q γ2 heterozygote_replicate4 [RS normalized]


title: SAN_AMPK R299Q γ2 heterozygote_replicate2 [RS normalized]


title: SAN_AMPK R299Q WT_replicate1 [RS normalized]


title: HeLa_Cyrano-MS2 pulldown_replicate4

description:
Cyrano4
IP RNA

tag: disease state


title: HeLa_Cyrano-siRNA transfected_replicate4

description:
Cyrano_si4
total RNA

source_name: HeLa_Cyrano-siRNA transfected

value: siRNA transfection with OIP5-AS1/Cyrano siRNA


title: HeLa_Cyrano-siRNA transfected_replicate2

description:
Cyrano_si2
total RNA


title: HeLa_Cyrano-MS2 pulldown_replicate1

description:
Cyrano1
IP RNA


title: SAN_AMPK R299Q γ2 heterozygote_replicate1 [RS normalized]


title: SAN_AMPK R299Q γ2 homozygote_replicate2 [RS normalized]


title: HeLa_Cyrano-MS2 pulldown_replicate2

description:
Cyrano2
IP RNA


title: HeLa_Cyrano-siRNA transfected_replicate3

description:
Cyrano_si3
total RNA


title: HeLa_MS2 pulldown_replicate1

description:
Control1
IP RNA

source_name: HeLa_MS2 pulldown

value: MS2-pulldown against empty-MS2


title: HeLa_Control-siRNA transfected_replicate4

description:
Ctl_si4
total RNA

source_name: HeLa_Control-siRNA transfected

value: siRNA transfection with control siRNA


title: HeLa_Cyrano-siRNA transfected_replicate1

description:
Cyrano_si1
total RNA


title: Gastrocnemius skeletal muscle_Middle-aged_replicate_2_mouse

status: Public on Sep 20 2017

description: Sample name: MA-2

scan_protocol: Arrays were scanned at a resolution of 0.8 μm using the Illumina BeadArray scanner.

data_processing: Data was extracted using the Illumina GenomeStudio software V2011.1, Gene expression module 1.9.0. Any spots at or below the background were filtered out using an Illumina detection Pvalue of 0.02 and above. The natural log of all remaining scores were used to find the avg and std of each array and the z-score normalization was calculated and presented below.  Z-score = (raw value - avg)/std.  Complete data including detection scores is supplied. ID_REF = Unique identifiers from GPL6885; Z_VALUE = Z transformation of the natural log of the raw intensity values; Detection_Pval = Detection Pvalue from Illumina GenomeStudio software V2011.1, Gene expression module 1.9.0.

growth_protocol: Experimental protocol: Young (5 months, N=5) , middle-aged (17 months, N=5), and old (28 months, N=5) male gastrocnemius skeletal muscle samples from C57BL/6 mice were used in this study. C57BL/6J mice were obtained from the Jackson Laboratory (Bar Harbor, ME, USA) and housed at the Gerontology Research Center (Baltimore, MD, USA). Mice were housed in cages of four with ad libitum access to house chow (Harlan Teklad Global 18% Protein Rodent Diet, Harlan Teklad, Madison, WI) and tap water. Muscle samples were flash frozen in liquid nitrogen and stored at -80oC until assayed.

extract_protocol: RNA was isolated from skeletal muscle using Trizol Reagent (Invitrogen, Carlsbad, CA) following the manufacturer’s instructions

value: Muscle

tag: Sex

value: Male

value: Middle-aged


title: HeLa_MS2 pulldown_replicate4

description:
Control4
IP RNA


title: HeLa_Control-siRNA transfected_replicate2

description:
Ctl_si2
total RNA


title: Gastrocnemius skeletal muscle_Old_replicate_1_mouse

description: Sample name: O-1

source_name: Gastrocnemius skeletal muscle_Old

value: Old


title: HeLa_Control-siRNA transfected_replicate3

description:
Ctl_si3
total RNA


title: Gastrocnemius skeletal muscle_Old_replicate_4_mouse

description: Sample name: O-4


title: Gastrocnemius skeletal muscle_Middle-aged_replicate_3_mouse

description: Sample name: MA-3


title: Gastrocnemius skeletal muscle_Middle-aged_replicate_5_mouse

description: Sample name: MA-5


title: Gastrocnemius skeletal muscle_Old_replicate_2_mouse

description: Sample name: O-2


title: Gastrocnemius skeletal muscle_Middle-aged_replicate_4_mouse

description: Sample name: MA-4


title: Gastrocnemius skeletal muscle_Old_replicate_5_mouse

description: Sample name: O-5


title: Gastrocnemius skeletal muscle_Young_replicate_3_mouse

description: Sample name: Y-3

source_name: Gastrocnemius skeletal muscle_Young

value: Young


title: Gastrocnemius skeletal muscle_Young_replicate_2_mouse

description: Sample name: Y-2


title: HeLa_MS2 pulldown_replicate2

description:
Control2
IP RNA


title: Gastrocnemius skeletal muscle_Middle-aged_replicate_1_mouse

description: Sample name: MA-1


title: HeLa_MS2 pulldown_replicate3

description:
Control3
IP RNA


title: HeLa_Control-siRNA transfected_replicate1

description:
Ctl_si1
total RNA


title: Gastrocnemius skeletal muscle_Young_replicate_1_mouse

description: Sample name: Y-1


title: Gastrocnemius skeletal muscle_Old_replicate_3_mouse

description: Sample name: O-3


title: Gastrocnemius skeletal muscle_Young_replicate_4_mouse

description: Sample name: Y-4


title: Gastrocnemius skeletal muscle_Young_replicate_5_mouse

description: Sample name: Y-5


title: SAN_AMPK R299Q γ2 homozygote_replicate4 [z-score normalized]


title: Vastus lateralis muscle_Old_replicate_6_rhesus

description: Sample name: O-17517

data_processing: Data was extracted using the Illumina GenomeStudio software V2011.1, Gene expression module 1.9.0. Any spots at or below the background were filtered out using an Illumina detection Pvalue of 0.02 and above. The natural log of all remaining scores were used to find the avg and std of each array and the z-score normalization was calculated and presented below.  Z-score = (raw value - avg)/std.  Complete data including detection scores is supplied. ID_REF = Unique identifiers from GPL10558; Z_VALUE = Z transformation of the natural log of the raw intensity values; Detection_Pval = Detection Pvalue from Illumina GenomeStudio software V2011.1, Gene expression module 1.9.0.

organism: Macaca mulatta

source_name: Vastus lateralis muscle_Old

growth_protocol: Experimental protocol: Young (6.00±0.00 y), middle-aged (16.48±0.31 y), and old (26.83±0.22 y) male skeletal muscle samples from rhesus monkeys (Macaca mulatta) were used in this study. Rhesus monkeys were housed continuously at the NIH Animal Center, Poolesville, MD. Monkeys were housed individually in standard nonhuman primate caging on a 12h light/12h dark cycle, room temperature 25.5 ± 0.5 °C, humidity at 60 ± 20%. Monkeys were maintained on standard monkey chow (TestDiet® #5038 Purina Mills, Richmond, IN), and received 2 meals per day at estimated ad libitum levels throughout the study and water was always available. Monkeys were monitored minimally 3 times daily by trained animal care staff. All animal procedures for this study were reviewed and approved by the Animal Care and Use Committee (ACUC) at the Biomedical Research Center (NIA/NIH).

treatment_protocol: Biopsies of the vastus lateralis were obtained from the monkeys from ongoing NIA studies, and flash frozen in liquid nitrogen and stored at -80oC until assayed.

tag: gender

value: male


title: Vastus lateralis muscle_Old_replicate_5_rhesus

description: Sample name: O-22383

tag: tissue


title: Vastus lateralis muscle_Old_replicate_4_rhesus

description: Sample name: O-12392


title: Vastus lateralis muscle_Old_replicate_3_rhesus

description: Sample name: O-509


title: Vastus lateralis muscle_Middle-aged_replicate_6_rhesus

description: Sample name: MA-NJ06

source_name: Vastus lateralis muscle_Middle-aged


title: Vastus lateralis muscle_Old_replicate_2_rhesus

description: Sample name: O-19Z


title: Vastus lateralis muscle_Young_replicate_2_rhesus

description: Sample name: Y-2

source_name: Vastus lateralis muscle_Young


title: Vastus lateralis muscle_Middle-aged_replicate_5_rhesus

description: Sample name: MA-NJ24


title: Vastus lateralis muscle_Middle-aged_replicate_1_rhesus

description: Sample name: MA-12237


title: Vastus lateralis muscle_Old_replicate_1_rhesus

description: Sample name: O-65Z


title: Vastus lateralis muscle_Middle-aged_replicate_2_rhesus

description: Sample name: MA-NJ29


title: Vastus lateralis muscle_Young_replicate_4_rhesus

description: Sample name: Y-ZD33


title: Vastus lateralis muscle_Young_replicate_5_rhesus

description: Sample name: Y-ZD64


title: Skeletal muscle_Old_replicate_4_human

description: Sample name: O-604

source_name: Skeletal muscle_Old

extract_protocol:
Experimental protocol: Young (31.2±0.86 y), middle-aged (46.0±0.71 y), and old (74.0±4.38 y) male skeletal muscle samples were obtained from humans who underwent surgery for hip dysplasia. Written informed consent was obtained from those participants, and the study was approved by the medical ethics committee of the Rizzoli Hospital (Italy). Muscle samples were flash frozen in liquid nitrogen and stored at -80oC until assayed.
RNA was isolated from skeletal muscle using Trizol Reagent (Invitrogen, Carlsbad, CA) following the manufacturer’s instructions.


title: Vastus lateralis muscle_Young_replicate_1_rhesus

description: Sample name: Y-1


title: Vastus lateralis muscle_Young_replicate_6_rhesus

description: Sample name: Y-ZC69


title: Vastus lateralis muscle_Middle-aged_replicate_4_rhesus

description: Sample name: MA-NJ18


title: Skeletal muscle_Old_replicate_3_human

description: Sample name: O-398


title: Vastus lateralis muscle_Young_replicate_3_rhesus

description: Sample name: Y-3


title: Skeletal muscle_Young_replicate_5_human

description: Sample name: Y-666

source_name: Skeletal muscle_Young


title: Skeletal muscle_Middle-aged_replicate_3_human

description: Sample name: M-149

source_name: Skeletal muscle_Middle aged


title: Skeletal muscle_Old_replicate_7_human

description: Sample name: O-492


title: Skeletal muscle_Middle-aged_replicate_2_human

description: Sample name: M-187


title: Skeletal muscle_Old_replicate_2_human

description: Sample name: O-297


title: Skeletal muscle_Old_replicate_5_human

description: Sample name: O-468


title: Skeletal muscle_Old_replicate_1_human

description: Sample name: O-344


title: Skeletal muscle_Young_replicate_4_human

description: Sample name: Y-613


title: Skeletal muscle_Middle-aged_replicate_4_human

description: Sample name: M-603


title: Skeletal muscle_Old_replicate_6_human

description: Sample name: O-114


title: Skeletal muscle_Middle-aged_replicate_1_human

description: Sample name: M-262


title: Vastus lateralis muscle_Old_replicate_5_rat

description: Sample name: 10-O_AL

data_processing: Data was extracted using the Illumina GenomeStudio software V2011.1, Gene expression module 1.9.0. Any spots at or below the background were filtered out using an Illumina detection Pvalue of 0.02 and above. The natural log of all remaining scores were used to find the avg and std of each array and the z-score normalization was calculated and presented below.  Z-score = (raw value - avg)/std.  Complete data including detection scores is supplied. ID_REF = Unique identifiers from GPL6101; Z_VALUE = Z transformation of the natural log of the raw intensity values; Detection_Pval = Detection Pvalue from Illumina GenomeStudio software V2011.1, Gene expression module 1.9.0.

growth_protocol: Experimental protocol: Young (N=5, 5 months), middle-aged (N=5, 17 months), and old (N=5, 30 months) male skeletal muscle samples from Fisher-344 rats were used in this study. Rats were fed a standard NIH-31 chow (Harlan Teklad, Indianapolis, IN, USA) and were singly housed in an environmentally controlled vivarium with unlimited access to water. Animal rooms were maintained at 20-22 oC with 30-70% relative humidity and a 12-hour light/dark cycle according to animal protocols and NIH guidelines. Vastus lateralis muscles were harvested from rats. Muscle samples were flash frozen in liquid nitrogen and stored at -80oC until assayed.


title: Skeletal muscle_Young_replicate_3_human

description: Sample name: Y-634


title: Skeletal muscle_Young_replicate_1_human

description: Sample name: Y-958


title: Vastus lateralis muscle_Old_replicate_3_rat

description: Sample name: 8-O_AL


title: Vastus lateralis muscle_Old_replicate_4_rat

description: Sample name: 9-O_AL


title: Vastus lateralis muscle_Middle-aged_replicate_2_rat

description: Sample name: AL7_MA


title: Vastus lateralis muscle_Young_replicate_2_rat

description: Sample name: 7-Y_AL


title: Vastus lateralis muscle_Middle-aged_replicate_3_rat

description: Sample name: AL8_MA


title: Vastus lateralis muscle_Young_replicate_3_rat

description: Sample name: 8-Y_AL


title: Vastus lateralis muscle_Young_replicate_4_rat

description: Sample name: 9-Y_AL


title: Vastus lateralis muscle_Young_replicate_1_rat

description: Sample name: 6-Y_AL


title: Vastus lateralis muscle_Middle-aged_replicate_5_rat

description: Sample name: AL10_MA


title: Vastus lateralis muscle_Old_replicate_2_rat

description: Sample name: 7-O_AL


title: Vastus lateralis muscle_Young_replicate_5_rat

description: Sample name: 10-Y_AL


title: Vastus lateralis muscle_Middle-aged_replicate_4_rat

description: Sample name: AL9_MA


title: Vastus lateralis muscle_Middle-aged_replicate_1_rat

description: Sample name: AL6_MA


title: Vastus lateralis muscle_Old_replicate_1_rat

description: Sample name: 6-O_AL


title: Age 33-35 at donation + high ovarian reserve Sample 1 [D29.1]

status: Public on Aug 29 2017

city: Barcelona

first: Montse

last: BARRAGAN

state: Select One

address: Baldiri Reixac, 15

country: Spain

institute: Clinica EUGIN

description: CELs transformed using GCCN-SST with gain_lock=0.75, then processed using RMA-Sketch. These analysis capabilities are available in Affymetrix Power Tools 1.18.0

hyb_protocol: Affymetrix's standard protocol

scan_protocol: AGCC Standard Protocol / GCS 3000

data_processing: Samples were processed using Affymetrix Power Tools (APT) utilizing GCCN-SST RMA with gain lock percent set to 0.75

label: biotin

source_name: Mature MII oocytes

label_protocol: 8 µg cDNA (3.2 µg from the first round and 4.8 µg from the second) were subsequently fragmented by DNAseI and biotinylated by terminal transferase (GeneChip Mapping 250K Nsp assy kit; Affymetrix, CA, USA).

extract_protocol: The zona pellucida was removed by Pronase (Roche Diagnostics, Spain) and oocytes were immediately and individually placed in 45 µl of a Proteinase K-based lysis buffer (20 mM DTT, 0.5% SDS, 1 μg/μl proteinase K, 10 mM Tris.HCl, pH 7.4), incubated at 65ºC for 15 minutes, and stored at -80ºC until use (González-Roca, 2010).   Total RNA extraction and amplification were based on the Pico-Profiling protocol and successive improvements for single oocyte use (Gonzalez-Roca et al, 2010; Vassena et al, 2011). Pico-profiling faithfully amplifies the total RNA from as little as 10 somatic cells, or 1 MII oocyte, with a level of false positive signal in downstream analysis that is equivalent to that obtained without an amplification step. Briefly, RNA was purified individual oocytes using RNA Clean XP bead suspension (Agencourt Bioscience, La Jolla, CA, USA) and eluted in 20 µl water. The RNA preparation was used for library preparation and amplification by Whole Transcriptome Amplification (WTA2, Sigma-Aldrich, Spain). SYBRGreen (Sigma-Aldrich, Spain) was added to the amplification reaction, which was performed in a CFX Real-time instrument (Bio-Rad, CA, USA) to monitor amplification yield. The first amplification reaction was stopped when the SYBRGreen signal reached a plateau, and cDNA was purified and quantified on a Nanodrop ND-1000 spectrophotometer (Thermo-Fischer, MA, USA). A second round of amplification was performed from 10 ng of cDNA.

treatment_protocol: Mature MII oocytes were vitrified on the day of retrieval using an open method and were warmed for the study according to standard procedure (Cryotop®, Kitazato®, BioPharma Co., Ltd; Japan) in five randomized groups, on different days during a period of 14 days, to minimize bias due to the handling process. After warming, oocytes were left for 2 hours at 37ºC and 6% CO2, to allow for reassembly of the metaphase plate (Bromfield et al, 2009).

value: Age 33-35 at donation

tag: ovarian reserve

value: high ovarian reserve


title: HR_3

status: Public on Aug 21 2017

first: Suvarna

last: Nadendla

state: MD

address: 670 W.Baltimore Street

department: Institute for Genome Sciences

institute: University of Maryland School of Medicine

description: Mus musculus cells

data_processing:
Illumina reads were mapped to the Mus musculus GRCm38 genome using TopHat v. 1.4.0
The mapped reads were counted using HTSeq and normalized and then the differential expression of each gene for one sample compared to another was determined using DESeq v. 1.5.24, and statistical analyses were performed using R-2.15.2, and was represented as the log2 of the fold-change values.
RPKMs for each gene were calculated for each sample.
Genome_build: Mus musculus GRCm38
Supplementary_files_format_and_content: Tab-delimited text files include RPKMs

source_name: Uninfected mouse heart controls

growth_protocol: 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). Blood was collected from anesthetized mice retro-orbitally and transferred to heparin-coated collection tubes. Following euthanasia, hearts were surgically excised and washed in PBS to remove blood. Isolated hearts were homogenized in 5ml of PBS followed by filtration of the homogenate through a 40µm cell strainer. Paired blood (BIP) and strained heart (HIP) samples were flash-frozen at -80° C in working aliquots with 10% glycerol.

extract_protocol:
For HIP-RNA, hearts were excised from HIP-infected mice (n=3) when deemed moribund. Hearts were rinsed, diced, fragments washed with ice cold PBS, and the fragments homogenized in RNAprotect bacteria reagent (Qiagen). For BIP-RNA, 10 mice pre-depleted for neutrophils using anti-Ly6G antibody (BioXCell, clone RB6-8C5), were infected with TIGR4 and blood was collected in RNAprotect bacteria reagent (Qiagen) when the mice were deemed moribund such that blood from 5 mice were pooled as one sample (n=2 BIP samples). These heart homogenates were stored at -80°C. On the day of total RNA isolation from heart homogenates, the samples were thawed and spun down to discard the supernatant. The pellets were further homogenized in 600 μL RLT with B-ME buffer using a motorized mortar for 30 seconds. The re-homogenized samples were then disaggregated in a Qiashredder followed by RNA extraction with the RNeasy Micro Kit (Qiagen) with DNase treatment on column and in solution. The isolated RNA was quantitated using Nanodrop and Bioanalyzer. Samples were then depleted of rRNAs using the Ribo-Zero rRNA Removal Kit for Gram-positive bacteria and human/mouse/rat (Illumina, San Diego, CA). 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. 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. Blood samples were isolated in a similar manner as biofilm and planktonic samples, including enzymatic lysis, except that cells were first disaggregated with the Qiashredder like for the heart samples.
Strand specific RNA libraries were prepared for sequencing using standard Illumina protocols

tag: sample type

value: uninfected mouse heart


title: Age 33-35 at donation + high ovarian reserve Sample 1 [D26.2]


title: Age 33-35 at donation + high ovarian reserve Sample 1 [D27.1]


title: Age 20-22 at donation + low ovarian reserve Sample 1 [D13.2]

value: Age 20-22 at donation

value: low ovarian reserve


title: Age 33-35 at donation + high ovarian reserve Sample 1 [D25.1]


title: Age 33-35 at donation + high ovarian reserve Sample 1 [D24.1]


title: Age 33-35 at donation + low ovarian reserve Sample 1 [D1.1]


title: HR_2

description: Mus musculus cells


title: HR_1

description: Mus musculus cells


title: Age 20-22 at donation + low ovarian reserve Sample 1 [D12.1]


title: Age 33-35 at donation + high ovarian reserve Sample 1 [D30.1]


title: Age 20-22 at donation + high ovarian reserve Sample 1 [D19.1]


title: Age 20-22 at donation + high ovarian reserve Sample 1 [D17.2]