|
| Status |
Public on Apr 17, 2017 |
| Title |
MCF10A_R3 |
| Sample type |
SRA |
| |
|
| Source name |
mammary gland/breast epithelial cell
|
| Organism |
Homo sapiens |
| Characteristics |
tissue: breast
cell type: luminal ductal cells
neoplasia type: fibrocystic disease
atcc id: ATCC CRL-10317
cell line: MCF10A
|
| Treatment protocol |
Cells were seeded at 1.5x106 cells per 100 mm dish and grown to 80% confluence. Cells were washed twice with phosphate buffered saline (PBS) and stored at -80C in TRIzol.
|
| Growth protocol |
MCF10a cells were grown in DMEM: F12 (Hyclone-SH30271), 5% (v/v) horse serum (Gibco #16050 lot #1075876) + 10ug/ml human insulin (Sigma I-1882)+ 20ng/ml recombinant hEGF (Peprotech AF-100-15) + 100ng/ml Cholera toxin (Sigma C-8052) + 0.5 ug/ml Hydrocortisone (Sigma H-0888) Pen/Strep (Life Technologies) and Glutamine (Life Technologies). MCF10A cells without growth factors were grown in DMEM: F12 (Hyclone-SH30271), 5% (v/v) horse serum (Gibco #16050 lot #1075876) + Pen/Strep (Life Technologies) and Glutamine (Life Technologies).
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was isolated from Trizol using Direct-zol RNA mini-prep kit (Zymo Research #R2050) with DNaseI digestion.
RNA libraries were prepared for sequencing with the TruSeq Stranded Total RNA with Ribo-Zero Gold kit using standard Illumina protocols with the exception that library amplifcation was performed using KAPA HiFi™ Real-Time PCR Library Amplification Kit (Kapa Biosystems #KK2701).
Barcoded libraries (Illumina TruSeq) were then loaded onto an Illumina HiSeq 1500 and single-end 100-base (SE100) sequencing was performed at the Advanced Genome Technologies Core Massively Parallel Sequencing Facility at UVM.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 1500 |
| |
|
| Description |
growth factor+
MCF10A_RNA-Seq_R3
|
| Data processing |
Remove adapter reads (Cutadapt v1.6). Trim low quality base calls from both ends, Min score >= 20, window of 10 step size of 1. (FASTQ Quality Trimmer 1.0.0).
The reads were aligned to hg38 transciptome with Tophat2 (v0.6)
Reads were quantified using HTSeq-count (v0.6) with gencode annotation (v22). Genes with very low expression (<3 counts) were removed from the analysis.
Differential gene expression was calculated by using the Deseq2 version 1.8.1 package in R 3.2.0.
Genome_build: hg38
Supplementary_files_format_and_content: Tables showing the counts from the HTSeq-count quantification.
|
| |
|
| Submission date |
Aug 19, 2016 |
| Last update date |
May 15, 2019 |
| Contact name |
Jonathan AR Gordon |
| E-mail(s) |
Jonathan.A.Gordon@uvm.edu
|
| Organization name |
University of Vermont
|
| Department |
Biochemistry
|
| Street address |
89 Beaumont Ave Given E209
|
| City |
Burlington |
| State/province |
VT |
| ZIP/Postal code |
05405 |
| Country |
USA |
| |
|
| Platform ID |
GPL18460 |
| Series (1) |
| GSE85857 |
Runx1 stabilizes the mammary epithelial cell phenotype and prevents epithelial to mesenchymal transition |
|
| Relations |
| BioSample |
SAMN05596816 |
| SRA |
SRX2035765 |
