|
| Status |
Public on Aug 01, 2020 |
| Title |
MCF10A_gc10_R5 |
| Sample type |
SRA |
| |
|
| Source name |
mammary gland/breast epithelial cell
|
| Organism |
Homo sapiens |
| Characteristics |
cell line: MCF10A
cell type: mammary gland/breast epithelial cell
neoplasia type: luminal ductal cells
atcc id: fibrocystic disease;ATCC CRL-10317
treatment: gc10
|
| Treatment protocol |
Prior to treatment cells were plated in serum containing media for 24hrs to establish a monolayer. Media was removed and replaced with 2% charcoal-stripped serum for 48hrs prior to 24hr treatment with each pharmaceutical agent. Cells were harvested for ChIP-seq analysis 24 hrs post-treatment as previously described (Messier, et al. Oncotarget 2016).
|
| Growth protocol |
MCF10A cells were grown in DMEM: F12 (Hyclone-SH30272 without phenol red), 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). MCF7 were grown in DMEM: F12 (Hyclone-SH30272 without phenol red) + 10% (v/v) FBS (Atlanta Biologicals). ) Prior to treatment cells were plated in serum containing media for 24hrs to establish a monolayer. Media was removed and replaced with 2% charcoal-stripped serum for 48hrs prior to 24hr treatment with each pharmaceutical agent. Cells were harvested for ChIP-seq analysis 24 hrs post-treatment as previously described (Messier, et al. Oncotarget 2016).
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was isolated using RNeasy Plus Mini kit (Qiagen #74134). RNA libraries were prepared for sequencing using standard Illumina protocols using the TruSeq Stranded Total RNA with Ribo-Zero Gold kit (Illumina #RS-122-2301). 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.
RNA libraries were prepared for sequencing using standard Illumina protocols using the TruSeq Stranded Total RNA with Ribo-Zero Gold kit (Illumina #RS-122-2301). 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 |
MCF10A_gc10_merged_norm_pos.bw
MCF10A_gc10_merged_norm_neg.bw
|
| Data processing |
Remove adapter reads (Cutadapt v1.6)
Trim 10bp from 5'end. (FASTQ Trimmer 1.0.0)
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 (v21). Genes with very low expression (< 10 counts) were removed from the analysis.
Strand specific read depth normalized bigwigs created by UCSC's genomeCoverageBed (specifying scale 10^6/readcount and strand) and bedGraphToBigWig
Genome_build: hg38
Supplementary_files_format_and_content: Tables showing the normalized counts from the HTSeq-count quantification. Bigwig tracks, strand specific and normalized for read depth (UCSC's genomeCoverageBed -bg -split -scale 10^6/$read_count -strand + or -).
|
| |
|
| Submission date |
Aug 09, 2017 |
| Last update date |
Aug 01, 2020 |
| 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 (2) |
| GSE102469 |
Epigenetic and Expression Profiling of Tissue Selective Estrogen Compounds in Breast Epithelial Cell Lines (RNA-seq) |
| GSE102472 |
Epigenetic and Expression Profiling of Tissue Selective Estrogen Compounds in Breast Epithelial Cell Lines |
|
| Relations |
| BioSample |
SAMN07488183 |
| SRA |
SRX3083312 |
