|
| Status |
Public on Dec 01, 2023 |
| Title |
MCF10A_WT_rep3_RNA |
| Sample type |
SRA |
| |
|
| Source name |
Breast Epithelium
|
| Organism |
Homo sapiens |
| Characteristics |
tissue: Breast Epithelium
cell line: MCF-10A
genotype: WT
|
| Growth protocol |
All cell lines were grown in 5% CO2 at 37oC with 1% Pen Strep in respective media conditions. MCF-10A parental cell lines were purchased from American Type Culture Collection (ATCC). MCF-10A cell line knock-ins were previously generated as described in Gustin et al. Parental MCF-10A cell lines were cultured in DMEM/F12 (1:1) supplemented with 5% horse serum, 20 ng/ml epidermal growth factor (EGF), 10 µg/ml insulin (Roche), 0.5 µg/mL hydrocortisone (Sigma), and 100 ng/ml cholera toxin (Sigma). Knock-in cell lines were maintained in the same conditions except without EGF. Cells were grown to 80% confluence. Twenty-four hours prior to collection of RNA or nuclei, for RNA-seq or RNA-seq respectively, all MCF-10A cells were transferred to an assay media of phenol red-free DMEM/F12 (1:1) supplemented with 1% charcoal-dextran stripped FBS (Fisher), 0.2 ng/ml EGF, 10 µg/ml insulin, 0.5 µg/mL hydrocortisone, and 100 ng/ml cholera toxin.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
RNA was prepared from all cell lines using the Qiagen RNeasy kit
Libraries were prepared by the Vanderbilt Technologies for Advanced Genomics (VANTAGE) Core using the Illumina Ribo-Zero Plus rRNA Depletion Kit. Each library was sequenced on an Illumina NovaSeq, PE150, at a requested depth of 50 million reads.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina NovaSeq 6000 |
| |
|
| Description |
RNAfeatureCounts_MCF10A_WT_E545K.txt
|
| Data processing |
All sequencing library reads were trimmed of adapters and assessed for quality using the Trim Galore! (version 0.4.0) Wrapper of Cutadapt and FastQC.
Trimmed reads were mapped to the human genome assembly hg38 using the Spliced Transcripts Alignment to a Reference (STAR) aligner (version 2.5.4b)
Mapped reads were sorted and filtered for mapping quality score over 30 using utilities within the SAMtools package (version 1.5)
Reads were counted to gene transcripts using featureCounts (version 2.0.0)
Assembly: hg38
Supplementary files format and content: tab-delimited text file containing raw counts for each sample
|
| |
|
| Submission date |
Nov 15, 2023 |
| Last update date |
Dec 01, 2023 |
| Contact name |
Adam Xavier Miranda |
| E-mail(s) |
adam.x.miranda@vanderbilt.edu
|
| Phone |
7045169891
|
| Organization name |
Vanderbilt University
|
| Street address |
2220 Pierce Avenue
|
| City |
Nashville |
| State/province |
TN |
| ZIP/Postal code |
37013 |
| Country |
USA |
| |
|
| Platform ID |
GPL24676 |
| Series (2) |
| GSE247819 |
A functional genomics process for systematic dissection and mutation-specific target discovery in breast cancer PIK3CA hotspot mutations [RNA-seq] |
| GSE247822 |
A functional genomics process for systematic dissection and mutation-specific target discovery in breast cancer PIK3CA hotspot mutations |
|
| Relations |
| BioSample |
SAMN38265165 |
| SRA |
SRX22533399 |
