|
| Status |
Public on Jun 24, 2016 |
| Title |
RWPE1 - Control Vector, CAD1 |
| Sample type |
SRA |
| |
|
| Source name |
RWPE1, control vector
|
| Organism |
Homo sapiens |
| Characteristics |
cell line: RWPE1
expression: Control vector
|
| Treatment protocol |
Human RWPE1 cells were engineered with empty vector (altered pTRIPZ), NKX3.1 wild type over-expression, and NKX3.1 (T164A) mutant over-expression. Cells were snap frozen for subsequent molecular analysis.
|
| Growth protocol |
NA
|
| Extracted molecule |
total RNA |
| Extraction protocol |
RNA was extracted using the MagMAX-96 RNA isolation kit (Ambion). Quality control was performed with Agilent Bioanalyser 2100. We used poly-A pull-down to enrich mRNAs from total RNA samples (200ng-1ug per sample, RIN>8 required).
Library preparation was performed using the Illumina TruSeq RNA prep kit. Libraries were sequenced using Illumina HiSeq 2000 at Columbia Genome Center.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 2000 |
| |
|
| Description |
CAD1
Processed data file: RNAseq_human_RWPE1.txt
|
| Data processing |
Libraries were sequenced using Illumina HiSeq 2000 at Columbia Genome Center. We multiplexed samples in each lane, which yielded targeted number of single-end/paired-end 100bp reads for each sample, as a fraction of 180 million reads for the whole lane.
We used RTA (Illumina) for base calling and CASAVA (version 1.8.2) for converting BCL to fastq format, coupled with adaptor trimming. We mapped the reads to a reference genome (Human: NCBI/build37.2) using TopHat (version 2.0.4) with 4 mismatches (--read-mismatches = 4) and 10 maximum multiple hits (--max-multihits = 10). To tackle the mapping issue of reads that are from exon-exon junctions, TopHat infers novel exon-exon junctions ab initio, and combines them with junctions from known mRNA sequences (refgenes) as the reference annotation. We estimated the relative abundance (aka expression level) of genes and splice isoforms using Cufflinks (version 2.0.2) with default settings. We tested for differentially expressed genes under various conditions using DESeq. It is an R package based on a negative binomial distribution that models the number of reads from RNA-seq experiments and tests for differential expression.
Raw RNA-seq counts were normalized (variance stabilization) using DESeq package in R-studio Version 0.99.484.
Genome_build: GRCh37
Supplementary_files_format_and_content: Tab-delimited text file includes DESeq normalized values for each Sample.
|
| |
|
| Submission date |
May 13, 2016 |
| Last update date |
May 15, 2019 |
| Contact name |
Antonina Mitrofanova |
| E-mail(s) |
mitrofanova.antonina@hotmail.com
|
| Organization name |
Rutgers University
|
| Street address |
65 Bergen Street, Rm 923B
|
| City |
Newark |
| State/province |
NJ |
| ZIP/Postal code |
07107 |
| Country |
USA |
| |
|
| Platform ID |
GPL11154 |
| Series (2) |
| GSE81439 |
Identification of an NKX3.1-G9a-UTY regulatory network that controls prostate differentiation (Human_RWPE1_RNA-Seq) |
| GSE81440 |
Identification of an NKX3.1-G9a-UTY regulatory network that controls prostate differentiation |
|
| Relations |
| BioSample |
SAMN04999845 |
| SRA |
SRX1761105 |
