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Sample GSM2133240 Query DataSets for GSM2133240
Status Public on Aug 10, 2016
Title Tumor RNA_2
Sample type SRA
 
Source name Primary tumor
Organism Mus musculus
Characteristics strain/background: C57BL/6
inoculated tumor: Lewis lung carcinoma (LLC)
tissue: Primary tumor
Treatment protocol We subcutaneously inoculated C57BL/6 mice with Lewis lung carcinoma (LLC). Three weeks later, tumor tissues were cut.
To purify tumor-derived exosomes, primary tumors were cut into small pieces and cultured in RPMI media without FBS for 12h. Then, the media was collected and centrifuged at 800g for 10 min, followed by a centrifugation step of 20,000g for 20 min to remove cellular debris. Next, the supernatant was filtered using a 0.2-μm filter (Pall Corporation). The collected media was then ultracentrifuged at 100,000g for 90 min at 4ºC. Then, the supernatant was discarded. Exosomes used for RNA and protein extraction were isolated using Exosome Precipitation Solution (ExoQuick-TC, System Biosciences) without ultracentrifugation.
Extracted molecule total RNA
Extraction protocol Total RNA was extracted and purified using miRNeasy Mini Kit (Cat # 217004, QIAGEN, GmbH, Germany) following the manufacturer’s instructions and checked for a RIN number to inspect RNA integrity by an Agilent Bioanalyzer 2100 (Agilent Technologies, Santa Clara, CA, US).
The 5' and 3' adaptors were ligated sequentially to the RNAs and amplified by RT-PCR. The amplification products were excised from 6% TBE urea gel (Invitrogen), and purified DNA fragments were clustered and sequenced by Illumina HiSeq 2500 following the manufacturer's protocols.
 
Library strategy RNA-Seq
Library source transcriptomic
Library selection cDNA
Instrument model Illumina HiSeq 2500
 
Description Tu-2
Data processing The exosome RNA sequence reads were pre-processed using FASTXToolkit (fastx_toolkit-0.0.13.2; http://hannonlab.cshl.edu/fastx_toolkit/commandline.html ), excluding low-quality reads (ambiguous N, quality < 10 nt, and length <18 nt) as well as adapter.
RNA sequencing reads that passed this quality control measure were then mapped to the mouse reference genome (GRCm38.p4) using TopHat (version:2.0.9).
Cufflinks v2.1.1 was used for the differential expression analysis: parameters of Cuffdiff were set to allow normalization of the number of fragments mapping to individual loci to improve the strength of differential expression analysis.
Cuffdiff labeled genes as significant or not significant based on whether the p-value of statistical test for differential expression was greater than the false discovery rate (FDR) after Benjamini-Hochberg correction for multiple testing. Genes with a q-value or FDR-adjusted p-value of < 0.05 and a fold change greater than 2 were considered statistically significant.
Genome_build: GRCm38.p4
Supplementary_files_format_and_content: Tab-delimited text files include normalized FPKM values and raw fragment counts for each Sample.
 
Submission date Apr 26, 2016
Last update date May 15, 2019
Contact name Liu Yanfang
E-mail(s) liuyanfang00215@163.com
Phone 0086-13918386805
Organization name Second Military Medical University
Department Immunology
Street address 800 Xiangyin Road
City Shanghai
ZIP/Postal code 200433
Country China
 
Platform ID GPL17021
Series (1)
GSE80678 Gene expression signatures for primary tumor and tumor-derived exosomes
Relations
BioSample SAMN04902345
SRA SRX1727325

Supplementary file Size Download File type/resource
GSM2133240_Tu-2.txt.gz 199.4 Kb (ftp)(http) TXT
SRA Run SelectorHelp
Raw data are available in SRA
Processed data provided as supplementary file
Processed data are available on Series record

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