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Sample GSM6209177

Query DataSets for GSM6209177

Status

Public on Jun 03, 2022

Title

SAM17356760

Sample type

SRA

Source name

skin

Organism

Mus musculus

Characteristics

tissue: skin
cell line: Genetically engineered mouse model (GEMM) tumor
background: C57BL/6
animal id: 2658
biopsy type: initial
treatment: na

Extracted molecule

total RNA

Extraction protocol

Total RNA was extracted from cells using the Qiagen All Prep nucleic isolation kit (Qiagen, Inc.; catalog number 80204, Valencia, CA) as per the manufacturer’s protocol, including the on-column DNase digestion. Quality control of samples was done to determine RNA quantity and quality before their processing by RNA sequencing (RNA-seq). The concentration and the integrity of total RNA samples were determined using NanoDrop 8000 (ThermoFisher Scientific, Waltham, MA) and 2200 TapeStation (Agilent Technologies, Santa Clara, CA), respectively.
One microgram of total RNA was used as an input material for library preparation using TruSeq RNA Sample Preparation Kit v2 (Illumina, San Diego, CA). Library size was confirmed using the 2200 TapeStation and high-sensitivity D1K screen tape (Agilent Technologies, Santa Clara, CA), and their concentration was determined by quantitative PCR-based method using Library quantification kit (Kapa Biosystems, Wilmington, MA). The libraries were multiplexed and then sequenced on HiSeq2500 (Illumina, San Diego, CA) to generate 30 M of single-end 50 base pair reads.

Library strategy

RNA-Seq

Library source

transcriptomic

Library selection

cDNA

Instrument model

Illumina HiSeq 2500

Description

These are biopsy samples derived from the Braf;PTEN melanoma GEMM. An initial biopsy from before treatment with vemurafenib and another biopsy sample after resistance on treatment was collected.

Data processing

Sequences were trimmed to 75 bp and filtered for sequencing quality and ribosomal RNA. Reads for which 30% or more of the nucleotides had a Phred quality score of 23 or lower were discarded. The remaining reads were aligned to the mouse genome using the Genomic Short-read Nucleotide Alignment Program (GSNAP), with default settings and the following parameters: -M 2 -n 10 -B 2 -i 1 -N 1 -w 200000 -E 1 --pairmax-rna = 200000 --clip-overlap. Further trimming was not performed. Multimapping reads were discarded. PCR duplications were filtered using PicardTools. We used established gene models from the National Center for Biotechnology Information (NCBI) database and extended those with internal Genomic Mapping and Alignment Program for mRNA and EST sequences (GMAP) alignments for additional genes not in NCBI database.
Assembly: mm9
Supplementary files format and content: Raw counts are provided in a gene x sample format. Entrez gene ids are given.

Submission date

Jun 01, 2022

Last update date

Jun 03, 2022

Contact name

Dorothee Nickles

E-mail(s)

nicklesd@gene.com

Organization name

Genentech

Street address

1 DNA Way