|
| Status |
Public on Jan 01, 2016 |
| Title |
J1014_untreated |
| Sample type |
SRA |
| |
|
| Source name |
A375 melanoma bearing mouse
|
| Organism |
Homo sapiens |
| Characteristics |
tissue: A375 tumor
phenotype: vemurafenib-sensitive
host mouse treatment: none
|
| Treatment protocol |
A375 bearing mice were treated with 120 mg/kg vemurafenib until day 39 (n=5) when tumors became resistant to vemurfenib treatment. For comparison control untreated A375 tumors (n=4) were taken and frozen (liquid nitrogen). For RNA extraction see extract protocol.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Five slides (5 µm each) from frozen xenografts were used for total RNA extraction using TRIzol reagent (Invitrogen). RNA was purified using a miRNeasy kit (Qiagen) according to the manufacturer's instructions. RNA-seq transcriptome libraries were prepared following the TruSeq RNA Sample Preparation v2 (Illumina, Part# 15026495 Rev. A), using 2 μg of total RNA. PolyA+ RNA was isolated using Sera-Mag oligo(dT) beads (Thermo Scientific) and fragmented with the Ambion Fragmentation Reagents kit. cDNA synthesis, end repair, A-base addition and ligation of the Illumina-indexed adaptors were performed according to Illumina's protocol. Libraries were then PCR amplified using Phusion DNA polymerase (NEB) for 14 PCR cycles before quantification and multiplexing. 10 pmol of multiplexed (n=4) paired-end libraries were sequenced with the Illumina HiSeq1000 (2 × 50-nt read length). Reads that passed the chastity filter of Illumina BaseCall software were used for subsequent analysis.
RNA libraries were prepared for sequencing using standard Illumina protocols
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 1000 |
| |
|
| Description |
control sample
|
| Data processing |
Read sequence alignment using STAR aligner v2.3.0.1 against hybrid human(GRCh37/hg19)/mouse genome(GRCm38/mm10)
Retaining only reads that map uniquely to human chromosomes
Realignment of retained reads using GNAP aligner version 2012-12-20 to human reference genome (GRCh37/hg19)
HTSeq version 0.5.3 on Ensembl v70 annotation
GC-content correction using EDASeq
Differential gene expression analysis using DESeq version 1.18.0
Genome_build: GRCh37/hg19
Supplementary_files_format_and_content: tab-delimited format for differential gene expression analysis results; matrix for raw read counts after GSNAP alignment
|
| |
|
| Submission date |
Nov 05, 2015 |
| Last update date |
May 15, 2019 |
| Contact name |
Andreas Schlattl |
| E-mail(s) |
andreas.schlattl@boehringer-ingelheim.com
|
| Organization name |
Boehringer Ingelheim RCV GmbH & Co KG
|
| Street address |
Dr. Boehringer-Gasse 5-11
|
| City |
Vienna |
| ZIP/Postal code |
1121 |
| Country |
Austria |
| |
|
| Platform ID |
GPL15433 |
| Series (1) |
| GSE74729 |
A novel RAF kinase inhibitor with DFG-out binding mode: high efficacy in BRAF-mutant tumor xenograft models in the absence of normal tissue hyperproliferation |
|
| Relations |
| BioSample |
SAMN04244276 |
| SRA |
SRX1420826 |
