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Sample GSM3937552 Query DataSets for GSM3937552
Status Public on Jul 10, 2021
Title 2048-1_heart_OGT_TG
Sample type SRA
Source name whole heart
Organism Mus musculus
Characteristics strain: C57BL/6J
genotype: OGT_TG
Sex: female
Treatment protocol No treatment
Extracted molecule total RNA
Extraction protocol To obtain high quality total RNA, the hearts were processed first in the Trizol reagent (ThermoFisher Scientific, Catalog # 15596018), and then by RNeasy mini columns (Qiagen, catalog # 74104) as follows. To avoid sampling bias, the entire hearts were homogenized in Trizol reagent at a weight (mg) to volume (µL) ratio of 1:15. Then, 1 mL of the homogenate was processed following the manufacturer's protocol until the step of phase separation. 0.5 mL of the aqueous phase was then mixed with an equal volume of 70% ethanol for subsequent RNA purification by the RNeasy mini kit. An on-column DNAse (RNase-Free DNase Set, Qiagen catalog # 79254) treatment step was included. The concentration of the RNA was determined by Qubit fluorometric quantitation (Qubit RNA BR Assay Kit, ThermoFisher Scientific, catalog # Q10210). The integrity of the total RNA was determined by a Fragment Analyzer (Advanced Analytical Technologies, Inc). The average RNA Quality Number (RQN) was 8.77 ± 0.07 (Mean ± SEM, n=24).
1 µg of total RNA from each sample was used for RNA sequencing library preparation with the TruSeq® Stranded mRNA Library Prep kit (Illumina, catalog # 20020594). The libraries were barcoded by TruSeq® RNA Unique Dual Indexes (Illumina, catalog # 20022371) and quantified by qPCR on a Bio-Rad CFX Connect Real-Time PCR detection system with the NEBNext Library Quant Kit for Illumina (New England Biolabs, catalog # E7630L). The libraries were normalized to 10 nM, pooled and sequenced.
RNA-seq. RNA sequencing was carried out at Johns Hopkins School of Medicine Genetic Resources Core Facility on a NovaSeq 6000 sequencing system (Illumina) with a S1 flow cell for 200 cycles, generating 100 bp paired-end reads.
Library strategy RNA-Seq
Library source transcriptomic
Library selection cDNA
Instrument model Illumina NovaSeq 6000
Description cardiac specific OGT transgenic
Data processing #Quality check: fastqc
#Trimming: (tests were done to decide that trimming was not necessary.)
Reads alignment was done with Tophat using mm10_refseq_nopseudo transcriptome index
ClassifyReads: #counts of how many reads are exonic, intronic, intergenic and overlaping a splice junction
Stringtie: assembly the reads into transcripts
Stringtie --merge : # merging transcripts form all samples in one nonredundant file. Automatically filters a number of transfrags that are probably artfifacts. A reference annotation file was used in order to merge novel isoforms and known isoforms and maximize overall assembly quality. # This will be used further on, in the analysis (cuffdiff);
Cuffdiff v2.2.1: # used to find significant changes in transcript expression.
Samtools: samtools index file.bam
IGV: # vizualize alignments and assemblies
Genome_build: Genome version : mm10
Supplementary_files_format_and_content: .xlsx file of FPKM values. One biological sample per column.
Submission date Jul 10, 2019
Last update date Jul 10, 2021
Contact name Mark E Anderson
Organization name Johns Hopkins School of Medicine
Department Department of Medicine
Street address Rangos 471, 855 N. Wolfe Street
City Baltimore
State/province MD
ZIP/Postal code 21205
Country USA
Platform ID GPL24247
Series (1)
GSE134121 Excessive O-GlcNAcylation causes heart failure and sudden death
BioSample SAMN12251649
SRA SRX6429325

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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