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Sample GSM2971223 Query DataSets for GSM2971223
Status Public on Feb 01, 2019
Title E18.5_1
Sample type SRA
 
Source name Lung epithelial cells
Organism Mus musculus
Characteristics cell type: lineage (CD31, CD45, CD146 and Ter119) negative, Epcam positive epithelial cells
strain: C57BL/6
tissue: lung
developmental stage: Embryonic day 18.5
Extracted molecule polyA RNA
Extraction protocol Epithelial cells from various developmental stages (E13.5, E15.5, E18.5, P14, and P56) were isolated from C57BL/6 mice. Lungs were cut into small pieces and digested enzymatically using collagenase (Wako, Osaka, Japan), Dispase II (Roche, Basel, Switzerland), and DNase I (Sigma-Aldrich, St Louis, MO) for 60 min at 37°C. Single-cell suspended lung cells were first stained with anti-mouse CD31, CD45, Ter119, CD146, and Epcam antibodies. Cells were next stained with Streptavidin-APC, followed by anti-APC microbeads (Miltenyi). Labeled cells were magnetically depleted. Finally, lineage (CD31, CD45, Ter119, and CD146)– propidium iodide– Epcam+ live epithelial cells were sorted with a MoFlo Astrios flow cytometer (Beckman Coulter).
polyA RNAs were isolated and amplified from donor epithelial cells according to the protocol described in (Huang H, Nucleic Actd Res, 2014) with some modifications. 3'SAGE-seq libraries were constructed according to the protocol described in (Matsumura H, Methods Mol Biol, 2012) with some modifications. Sequencing was performed by using Ion Hi-Q Chef Kit, Ion PI v3 Chip Kit and Ion Proton Sequencer (Thermo Fisher Scientific) according to the manufacture’s instructions except the input library concentration (100 pM) and cycle number (200).
 
Library strategy RNA-Seq
Library source transcriptomic
Library selection cDNA
Instrument model Ion Torrent Proton
 
Data processing Adapter trimming and quality filtering of sequencing data were performed by Trimommatic-0.33 and PRINSEQ-0.20.4. Trimmed reads were mapped to the Refseq mm10 by using Bowtie2-2.2.5. Reads that were not mapped to NlaIII sites were removed, and quantified tag numbers of each gene as the expression level of each gene.
Total tag number was adjusted to 1 million tags, and between-sample normalization was performed by using R-3.4.0 and TCC package.
Genome_build: mm10
Supplementary_files_format_and_content: tab-delimited text files include normalized tag-count values for each sample
 
Submission date Jan 30, 2018
Last update date Feb 01, 2019
Contact name Kazushige Shiraishi
Organization name The University of Tokyo
Department Graduate School of Medicine
Lab Molecular Preventive Medicine
Street address 7-3-1, Hongo, Bunkyo-ku
City Tokyo
ZIP/Postal code 113-0033
Country Japan
 
Platform ID GPL18635
Series (1)
GSE109847 Transcriptome analysis of lung epithelial cells from various developmental stages (E13.5, E15.5, E18.5, P14, and P56)
Relations
BioSample SAMN08437203
SRA SRX3630280

Supplementary file Size Download File type/resource
GSM2971223_E18.5_1.txt.gz 114.2 Kb (ftp)(http) TXT
SRA Run SelectorHelp
Raw data are available in SRA
Processed data provided as supplementary file

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