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Sample GSM1686477

Query DataSets for GSM1686477

Status

Public on Dec 04, 2015

Title

P1_tetoff_10619-3

Sample type

SRA

Source name

P1_tetoff_heart_whole

Organism

Mus musculus

Characteristics

strain background: FVB/N
genotype/variation: tet-off control (tetoff)
age: P1
tissue: heart, whole

Extracted molecule

polyA RNA

Extraction protocol

Hearts were removed and ventricular apexes were flash frozen on dry ice, and RNA was harvested using Trizol reagent.
Polyadenylated RNA was isolated using oligo(dT) binding, followed by chemical fragmentation and preparation of non-strand-specific, single-end RNA-Seq libraries, as per Methods Mol Biol. 2015;1299:27-49, PMID 25836573.
non-strand-specific, single-end RNA-Seq

Library strategy

RNA-Seq

Library source

transcriptomic

Library selection

cDNA

Instrument model

Illumina HiSeq 2500

Data processing

Illumina Casava1.6 and Casava1.8 software was used for basecalling. All 50 nt reads use phred33 quality scores.
Sequence reads were mapped to the transcriptome, defined by annotation based on the UCSC mm10 genes.gtf but with mitochondrial RNAs and human-Mfn2 cDNA added, and associated bowtie2 index provided by Illumina iGenomes (incorporating the additional mitochondrial RNAs and human-Mfn2 cDNA), using tophat v2.0.10.
htseq-count (authored by Simon Anders) was used to assign tophat-aligned reads to gene entries in the supplied gtf, based on UCSC mm10 genes.gtf but with mitochondrial RNAs and human-Mfn2 cDNA added.
htseq-count data allows each user maximum flexibility in performing sample (library) depth normalization and differential gene expression using software such as DESeq or edgeR.
Additionally, we used Cufflinks v2.1 to calculate FPKM for each gene entry in each library. However, this was merely done for later graph display. Actual calculations of differential gene expression used htseq-count data as input to DESeq.
Genome_build: mm10
Supplementary_files_format_and_content: single tab-delimited text files include htseq-count values for each gene, or FPKM values for each gene.

Submission date

May 15, 2015

Last update date

May 15, 2019

Contact name

Scot J Matkovich

E-mail(s)

sjmatkovich@lilly.com

Organization name

Eli Lilly and Company

Street address

Lilly Corporate Center