|
| Status |
Public on Apr 14, 2020 |
| Title |
Human_fetal_da_cell_3_RNASeq |
| Sample type |
SRA |
| |
|
| Source name |
Immortalized fetal DA cell rep 1
|
| Organism |
Homo sapiens |
| Characteristics |
passage: 2
cell type: Immortalized fetal DA cell
|
| Growth protocol |
Primary human and mouse myometrial cells, as well as PHM1 cells were cultured in DMEM (Corning MT17205CV) supplemented with 10% fetal bovine serum (FBS), 2mM L-Glutamine, 100 U/ml penicillin-streptomycin. hTERT-HM cells were cultured in DMEM-F12 (Gibco 11-039-021) supplemented with 10% FBS and 100 U/ml penicillin-streptomycin. Primary adult aorta cells were cultured in vascular cell basal medium (ATCC® PCS-100-030™). Fetal aorta and DA cells were cultured in DMEM (Gibco 11965-092) supplemented with 10% FBS and 100 U/ml penicillin-streptomycin.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Flash frozen tissues stored in -80oC were homogenized in TRIzolTM reagent (Invitrogen) using a bead homogenizer (Fisherbrand™ Bead Mill 24) for two 20sec cycles at 5000 rpm. Near-confluent cells were lysed using 1mL TRIzolTM reagent. Total RNA was isolated per the Trizol’s manufacturer instructions, DNase-treated (DNA-free kit, Ambion) and then sent to the Vanderbilt Technologies for Advanced Genomics (VANTAGE) core for quality control and cDNA library preparation. The RNA was analyzed with an Agilent Bioanalyzer 2100, which showed that all samples had an RIN>8.
The coding RNA was enriched using the NEBNext® Poly(A) mRNA Magnetic Isolation Module (E7490) and paired end libraries were generated using the NEBNext Ultra II Directional RNA Library Kit for Illumina (7760).
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina NovaSeq 6000 |
| |
|
| Data processing |
Quality control on raw reads was performed using FastQC.
STAR v2.5.3a used for reads mapping.
FeatureCounts v1.15.2 was used to count the number of mapped reads to each gene.
BiomaRt package was used to map mouse genes to human genes, and only uniquely mapped genes between human and mouse were kept.
Significantly differentially expressed genes with FDR-adjusted p-value≤0.01 and absolute fold-change≥2.0 were detected by DESeq2 (v1.18.1).
Genome Ontology and Kyoto Encyclopedia of Genes and Genome (KEGG) pathway over-representation analysis was performed on differentially expressed genes using the WebGestaltR package.
Gene set enrichment analysis was performed using the GSEA package.
Genome_build: mm10 and hg38
Supplementary_files_format_and_content: tab-delimited text files include count values for each Sample.
|
| |
|
| Submission date |
Jan 26, 2019 |
| Last update date |
Apr 14, 2020 |
| Contact name |
Dhivya Sudhan |
| E-mail(s) |
Dhivya.Sudhan@UTSouthwestern.edu
|
| Organization name |
Vanderbilt University
|
| Department |
Hematology/Oncology Division
|
| Street address |
777 Preston Research Building
|
| City |
Nashville |
| State/province |
TN |
| ZIP/Postal code |
37232-6307 |
| Country |
USA |
| |
|
| Platform ID |
GPL24676 |
| Series (1) |
| GSE125701 |
Comparative transcriptome profiling myometrial and vascular smooth muscle cells |
|
| Relations |
| BioSample |
SAMN10823619 |
| SRA |
SRX5295029 |
