|
| Status |
Public on Jun 28, 2021 |
| Title |
Control_E13.5OFT_replicate2 [SL203086] |
| Sample type |
SRA |
| |
|
| Source name |
Control_E13.5_Outlfow tract
|
| Organism |
Mus musculus |
| Characteristics |
developmental stage: E13.5
genotype: wildtype
tissue: Embryonic cardiac outflow tract
|
| Extracted molecule |
total RNA |
| Extraction protocol |
E13.5 embryos were collected in an RNase-free environment. A blunt cut at the base of the OFT, below the conal cushions, was used to collect the OFT tissues from each embryo. The OFT were snap frozen until genotyping had been completed using tissue from the remaining embryo. Following genotyping of the embryos, three OFTs were pooled in 1 ml Trizol to form one biological replicate. Three biological replicates of the mutant embryos and three biological replicates of the wild type embryos were used for the final RNA-Seq analysis (i.e. 3 OFTs per sample submitted, 18 total OFTs). Total RNA was collected following tissue homogenization using TissueLyser II (Qiagen) and chloroform-isopropanol extraction and purification and isolated using the Total RNA Purification kit (Norgen Biotek Corp, 17200).
RNA-Seq was performed at OceanRidge Biosciences as previously described. RNA TruSeq Stranded Total RNA LT with Ribo-Zero Gold Set A kit (Illumina, RS-122-2301) was used to generate libraries.
Libraries were sequenced using Illumina HiSeq 2500 to generate paired-end 50 bp reads.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 2500 |
| |
|
| Description |
C9CUFANXX_s7_*_TruseqHTDual_D705-TruseqHTDual_D503_SL203087
|
| Data processing |
The raw FASTQ files were split into files containing 4,000,000 reads and checked for quality using the FASTX-Toolkit1 (version 0.0.14, http://hannonlab.cshl.edu/fastx_toolkit/). The reads were filtered (removing sequences that did not pass Illumina's quality filter) and trimmed based on the quality results (3 nucleotides at the left end of the R1 reads)
Then, sequence alignment was performed using TopHat (v2.1.0) to mouse genome version mm10 (ftp://ftp.ccb.jhu.edu/pub/data/bowtie2_indexes/mm10).
Following the alignment to the mouse genome, BAM files were merged on a per-sample basis. Generation of BAM files was performed by OceanRidge Biosciences.
Aligned bam files are used to count the number of reads mapping to exons in each transcript using the GenomicAlignments (version 1.22.1) package in R to generate a gene-count matrix.
Differential expression was evaluated from this gene-count matrix using the DESeq2 package (version 1.26.0) using the standard differential expression analysis pipeline (without log fold change shrinkage method) in R.
Genome_build: mouse genome version mm10 (ftp://ftp.ccb.jhu.edu/pub/data/bowtie2_indexes/mm10).
Supplementary_files_format_and_content: xlsx, Differential expression analysis output from DESeq2
|
| |
|
| Submission date |
Mar 31, 2021 |
| Last update date |
Jun 28, 2021 |
| Contact name |
Vidu Garg |
| E-mail(s) |
vidu.garg@nationwidechildrens.org
|
| Phone |
614-355-5710
|
| Organization name |
Research Institute at Nationwide Children's Hospital
|
| Department |
Cardiovascular Research
|
| Lab |
Vidu Garg
|
| Street address |
700 Children's Drive WB4239
|
| City |
Columbus |
| State/province |
OH |
| ZIP/Postal code |
43205 |
| Country |
USA |
| |
|
| Platform ID |
GPL17021 |
| Series (2) |
| GSE171237 |
A multi-omics approach using a mouse model of cardiac malformations for prioritization of human congenital heart disease contributing genes [RNA-seq] |
| GSE171239 |
A multi-omics approach using a mouse model of cardiac malformations for prioritization of human congenital heart disease contributing genes |
|
| Relations |
| BioSample |
SAMN18572564 |
| SRA |
SRX10489298 |
