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Sample GSM2860261 Query DataSets for GSM2860261
Status Public on Jan 01, 2018
Title Total tumor rep1
Sample type SRA
Source name Lewis lung carcinomas
Organism Mus musculus
Characteristics cell type: Total tumor
Treatment protocol Alexa 647-collagen was injected into Lewis lung carcinomas formed by subcutaneous inoculation of dTomato-expressing Lewis lung carcinoma cells into transgenic Col1a1-GFP mice. 24 h later, single-cell suspensions were prepared, FC receptor-blocked, and stained with Zombie aqua and anti-F4/80 antibody. Cells were sorted into dTomato+ cells (Lewis lung carcinoma cells), GFP+ cells (fibroblasts), and dTomato-;GFP-;F4/80+;collagen+ cells (M2-TAMs).
Growth protocol For generation of fluorescently tagged Lewis Lung carcinoma cells , cultured cells were transfected with a dTomato-containing pCEFL expression vector and subjected to FACS-sorting for dTomato-positive cells. The sorted cells were subsequently maintained in culture media supplemented with 2 mg/ml G418. Cells (5×10^5 per injection) were injected subcutaneously in both flanks of mice.
Extracted molecule total RNA
Extraction protocol Cells were sorted into cold media, and RNA was isolated by cell lysis with Trizol (Invitrogen), phase separation with chloroform, and subsequent purification of RNA from the aqueous phase using an RNA micro purification kit (Qiagen).
RNA quality was assessed using the Agilent 2100 Bioanalyzer. Total RNA (200 ng) was prepared for sequencing using polydT-mediated cDNA synthesis according to the manufacturer´s (Illumina) instructions. Subsequent library preparation was performed using the NEBNext RNA library preparation kit for Illumina. Library quality was assessed using the Fragment Analyzer (AATI) followed by library quantification (Illumina library quantification kit). Sequencing was carried out on a HiSeq1500 platform (Illumina) with a read length of 50 bp.
Library strategy RNA-Seq
Library source transcriptomic
Library selection cDNA
Instrument model Illumina HiSeq 1500
Data processing Sequenced reads were aligned to the mm9 mouse genome assembly using STAR
Uniquely aligned reads were quantified at exons of annotated genes and normalized to sequence depth and gene length using HOMER. The number of reads per kilobase per million mapped (RPKM) for all RefSeq annotated genes were calculated
Genome_build: mm9
Supplementary_files_format_and_content: tab-delimited text file include RPKM values for all annotated genes for each sample
Submission date Nov 17, 2017
Last update date May 15, 2019
Contact name Lars Grøntved
Phone +45 24 60 14 06
Organization name University of Southern Denmark
Department Biochemistry and Molecular Biology
Street address Campusvej 55
City Odense
ZIP/Postal code 5230
Country Denmark
Platform ID GPL18480
Series (1)
GSE107053 Tumor-associated macrophages derived from circulating inflammatory monocytes degrade collagen through cellular uptake
SRA SRX3403559
BioSample SAMN08044601

Supplementary data files not provided
SRA Run SelectorHelp
Raw data are available in SRA
Processed data are available on Series record

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