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Sample GSM5278865 Query DataSets for GSM5278865
Status Public on Jun 27, 2022
Title CM_Hypoxia_2
Sample type SRA
 
Source name Cardiomyocytes
Organism Rattus norvegicus
Characteristics cell type: Neonatal Rat Primary Ventricular Cardiomyocytes
strain: Sprague Dawley
treatment: Hypoxia with glucose and serum free medium for 5 hours
sample type: Cells
Treatment protocol For glucose and serum deprivation (GSD) treatment, HEK and Bewo cells were cultured in glucose free DMEM basal medium for 24 hours; CF and CM cells were exposed to glucose free DEME basal medium for 5 hours. For hypoxia treatment, HEK and Bewo cells were fed with complete medium and maintained in hypoxia chamber with 0.2% oxygen for 24 hours; CF and CM cells were fed with glucose free DMEM basal medium and maintained in hypoxia chamber with 0.2% oxygen for 5 hours. For oxidative stress, HEK and Bewo cells were treated with 0.7mM H2O2 and 1.7 mM H2O2 in complete medium for 24 hours; CF and CM cells were exposed to hypoxia treatment for 5 hours and then fed with complete medium in normal incubator for additional 24 hours. FBS and horse serum used here were extracellular vesicle-depleted by 24 hours of ultracentrifugation.
Growth protocol HEK293 cells and cardiac fibroblasts (CFs) were grown in DMEM medium containing 10% FBS and 1% Pen/Strep; Bewo cells were maintained in F12K medium containing 10% FBS and 1% Pen/Strep; cardiomyocytes (CMs) were cultured in DMEM meidum containing 10% horse serum, 5% FBS and 1% Pen/Strep.
Extracted molecule total RNA
Extraction protocol Cellular total RNAs were isolated by using TRIzol™ and then small RNAs were extracted by using mirVana™ miRNA Isolation Kit; extracellular RNAs were extracted by using exoRNeasy Midi kit.
Small RNAs were sequentially treated with DNase I, T4 PNK, and His-AlkB and then subjected to NEBNext® Multiplex Small RNA Library Prep Set for Illumina®.
 
Library strategy OTHER
Library source transcriptomic
Library selection other
Instrument model Illumina HiSeq 2500
 
Description small RNAs
Data processing Library strategy: ARM-seq
Adapters were removed and reads were merged with SeqPrep [John st. John unpub] using a minimum read length of 15 and adapters AGATCGGAAGAGCACACGTC and GATCGTCGGACTGTAGAACTC. Reads that could not be merged were discarded before mapping.
Data were analyzed with tRAX (http://trna.ucsc.edu/tRAX/), tRAX maps reads using bowtie2 v2.3.5.1 with parameters --very-sensitive --ignore-quals --np 5 -k 100, to a custom database containing mature tRNA transcripts and the genome sequence.
ENSEMBL (release 96) small ncRNA annotations were used to count the number of small RNA transcripts using only primary alignments to prevent double counting. Transcripts with 0 reads across all samples were removed.
tDRnamer (http://trna.ucsc.edu/tDRnamer/) was used to generate unique identifiers for each of the reads which mapped to a tRNA transcript based on the isodecoder(s) they mapped to and positions of any misincorporations.
DESeq2 v1.30.1 was used to generate normalized read counts for each of the transcripts across each sample.
Genome_build: Homo Sapiens GRCh38/hg38 and Rattus Norvegicus rn6
Supplementary_files_format_and_content: Processed data files are comma-separated files containing DESeq2 normalized read counts.
 
Submission date May 04, 2021
Last update date Jun 27, 2022
Contact name Guoping Li
E-mail(s) gli21@mgh.harvard.edu
Organization name Massachusetts General Hospital
Department CVRC
Lab Dr. Saumya Das
Street address Cardiovascular Research Center, 185 Cambridge St, Simches 3.5000
City Boston
State/province MA
ZIP/Postal code 02114
Country USA
 
Platform ID GPL18694
Series (1)
GSE173806 Cellular and extracellular tRNA-derived fragments demonstrate distinct signatures in cellular stress
Relations
BioSample SAMN19006662
SRA SRX10767899

Supplementary data files not provided
SRA Run SelectorHelp
Raw data are available in SRA
Processed data are available on Series record

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