|
| Status |
Public on Aug 11, 2017 |
| Title |
MCF-7 cell line Anti-miR-29a transfected replicate 2 |
| Sample type |
SRA |
| |
|
| Source name |
MCF-7 cell line Anti-miR-29a transfected
|
| Organism |
Homo sapiens |
| Characteristics |
cell line: MCF-7
transfection: Anti-miR-29a
|
| Treatment protocol |
Cells were transfected with anti-miR-29a (Anti-miRTMs, Ambion), pre-miR-29a and pre-miR-29b-1 (Pre-miRTMs, Ambion), using Lipofectamine RNAiMAX (Invitrogen) for 48 h.
|
| Growth protocol |
MCF-7 and LCC9 cells were grown in triplicates in phenol red IMEM supplemented with 5% dextran-coated charcoal-stripped fetal bovine serum (FBS) and 1% penicillin/streptomycin (P/S).
|
| Extracted molecule |
total RNA |
| Extraction protocol |
RNA was isolated using the Exiqon miRCURYTM RNA Isolation kit and its concentration and quality assessed using a NanoDrop spectrophotometer. Quantitative real-time PCR confirmed overexpression of miR-29b-1, miR-29a, or knockdown of miR-29b-1 and miR-29a in respective samples.
The Truseq Stranded mRNA kit was used to prepare mRNA libraries from 2ug 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. 75-76 cycle single read sequencing was performed with the 500 High-output v2 (75 cycle) sequencing kit on the Illumina NextSeq500 instrument.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina NextSeq 500 |
| |
|
| Description |
MCF-7_2B
|
| Data processing |
Illumina FASTQ Generation version 1.0.0 used for basecalling
Quality control (QC) of the raw sequence data was performed using FastQC. The FastQC results indicated that all sequences were of high quality.
Sequences were directly aligned to the Homo sapiens reference genome assembly hs_ref_GRCh37 downloaded from NCBI. The annotation file Homo_sapiens.GRCh37.73.gtf Downloaded from Ensembl August 26, 2015 was used with all records annotated as mitochondrial or ribosomal RNA removed.
The analysis was performed using The Illumina Basesapce RNA-Seq Alignment App version 1.0.0 with Tophat2 and Bowtie2, generating alignment files in bam format.
Differentially expressed genes (DEGS) were identified using Cuffdiff (v2.2.1).
Genome_build: hg19
Supplementary_files_format_and_content: [.txt] tab-delimited cuffnorm normalized FPKM file
|
| |
|
| Submission date |
May 19, 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) |
| GSE81620 |
Next Generation Sequencing Facilitates Quantitative Analysis of miR-29b-1 and miR-29a targets in tamoxifen-sensitive and tamoxifen-resistant human breast cancer cells |
|
| Relations |
| BioSample |
SAMN05017686 |
| SRA |
SRX1774626 |
