|
| Status |
Public on Feb 01, 2017 |
| Title |
MDA MB-231 cell line treated with anacardic acid replicate 2 |
| Sample type |
SRA |
| |
|
| Source name |
MDA MB-231 cell line
|
| Organism |
Homo sapiens |
| Characteristics |
cell line: MDA MB-231
|
| Treatment protocol |
Prior to treatment, normal growth medium is replaced with phenol red-free starvation medium containg 5% dextran coated charcoal stripped fetal bovine serum (DCC-FBS) for 48 hours. Treatment includes addition of 13.5uM or 35uM Anacardic Acid dissolved in EtOH for MCF-7 or MDA-MB-231, respectively.
|
| Growth protocol |
MCF-7 and MDA-MB-231 are maintained in a base growth medium Improved Minimum Essential Medium (IMEM) containing 5% Fetal Bovine Serum (FBS) and 1% Penecillin/Streptomycin. Cells are kept in an incuabtor at 37°C containing 5% CO2 for optimal growth.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
After treatment medium is removed and a total RNA isolation kit from Exiqon was used to extract both mRNA and miRNA. The kit itseld includes the binding of RNA (mRNA and miRNA) to a spin column, followed by a series of washes for purification and finally elution.
RNA libraries were prepared for sequencing using standard Illumina protocols. The Truseq Stranded mRNA kit was used to prepare mRNA libraries from 1ug total RNA. Libraries were confirmed on the Agilent 2100 Bioanalyzer and quantitated using the Illumina Library Quantification Kit, ABI Prism qPCR Mix from Kapa Biosystems and the ABI7900HT real-time PCR instrument. 76 cycle single read sequencing was performed with the 500 High-output v2 (75cycle) sequencing kit on the Illumina NextSeq500 instrument.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina NextSeq 500 |
| |
|
| Data processing |
Illumina BaseSpace FastQ v1.0.0 used for basecalling
Raw sequences were checked for quality values using fastQC v0.10.1. Quality trimming was not deemed necessary.
Raw sequences were mapped to the hg19 genome assembly using tophat2 v2.0.13 generating alignments in bam format, using an Ensembl gene and transcript gtf as a guide.
Mapped reads were assembled according to the Ensembl hg19.gtf file using cufflinks v2.2.1.
Cufflinks assembles were merged using cuffmerge.
Differentially expressed genes were compared in four pairwise comparisions using cuffdiff v2.2.1. The four comparisons include: MCF-7 AnActrt treatment vs. control; MDA MB-231 AnActrt treatment vs. control; MCF-7 AnActrt treatment and MDA MB-231 AnActrt treatment vs. MCF-7 control and MDA MB-231 control; and MCF-7 AnActrt treatment and MCF-7 control vs. MDA MB-231 AnActrt treatment and MDA MB-231 control.
The resulting mapped files from cufflinks were normalized and concatenated into a single tab-delimited gene matrix file using cuffnorm.
Genome_build: hg19
Supplementary_files_format_and_content: tab-delimited text file containing cuffnorm normalized FPKM for each sample
|
| |
|
| Submission date |
Feb 17, 2016 |
| Last update date |
May 15, 2019 |
| Contact name |
Eric Christian Rouchka |
| E-mail(s) |
eric.rouchka@louisville.edu
|
| Organization name |
University of Louisville
|
| Department |
Biochemistry and Molecular Genetics
|
| Lab |
KY INBRE Bioinformatics Core
|
| Street address |
522 East Gray Street
|
| City |
Louisville |
| State/province |
Kentucky |
| ZIP/Postal code |
40292 |
| Country |
USA |
| |
|
| Platform ID |
GPL18573 |
| Series (1) |
| GSE78011 |
RNA-Seq Analysis of Anacardic Acid Treated MCF7 and MDA-MB-231 Breast Cancer Cell Lines |
|
| Relations |
| SRA |
SRX1589792 |
| BioSample |
SAMN04503808 |
