characteristics.text: ERC000011
characteristics.characteristic: ENA-CHECKLIST
characteristics.text: 2017-10-15T17:01:20Z
characteristics.characteristic: ENA-FIRST-PUBLIC
characteristics.text: 2017-09-04T10:55:22Z
characteristics.characteristic: ENA-LAST-UPDATE
characteristics.text: SAMEA104279665
characteristics.characteristic: External Id
characteristics.text: Institute of Experimental Immunology, University of Zurich
characteristics.characteristic: INSDC center name
characteristics.text: 2017-10-15T17:01:20Z
characteristics.characteristic: INSDC first public
characteristics.text: 2017-09-04T10:55:22Z
characteristics.characteristic: INSDC last update
characteristics.text: public
characteristics.characteristic: INSDC status
characteristics.text: ERS1904643
characteristics.characteristic: SRA accession
characteristics.text: E-MTAB-6028:Sample 4
characteristics.characteristic: Submitter Id
characteristics.text: 3
characteristics.unit: day
characteristics.characteristic: age
characteristics.text: ArrayExpress
characteristics.ontologyTerms: http://www.ebi.ac.uk/efo/EFO_0009737
characteristics.characteristic: broker name
characteristics.text: alveolar macrophage
characteristics.characteristic: cell type
characteristics.text: house mouse
characteristics.characteristic: common name
characteristics.text: Protocols: Mice were sacrificed by CO2 inhalation and intracardially perfused with approximately 20 ml PBS. The lung was carefully removed and cut into small pieces using scissors, followed by a tissue digestion in 0.4 mg/ml collagenase type IV (from Clostridium histolyticum) in HBSS (containing Ca2+ / Mg2+) supplemented with 10 % FCS for 45 min at 37C while agitating. Digested tissues were passed through an 18 G syringe and filtered through a 70m cell strainer to obtain a homogeneous cell suspension. After PBS washing and centrifugation at 4C for 5 min at 1500rpm, the cell pellet was resuspended in red blood cell lysis buffer and incubated on ice for 8 mins. After further PBS washing and centrifugation, the cell pellet was resuspended in antibody mix and stained at 4C for 25 mins. After PBS washing and centrifugation, resuspend the cell pellet in FACS buffer (2mM EDTA, 2%FCS in PBS) and ready for FACS sorting. Sorted cells were lysed in RLT plus buffer (Qiagen) supplemented with 1 % mercaptoethanol. RNA isolation was performed using the RNeasy Micro Plus Kit (Qiagen) according to the manufacturers instructions. The quantity and quality of the isolated RNA was determined with a Qubit® (1.0) Fluorometer (Life Technologies, California, USA) and a Bioanalyzer 2100 (Agilent,Waldbronn, Germany). The TruSeq Stranded mRNA Sample Prep Kit (Illumina, Inc,California, USA) was used in the succeeding steps. Briefly, total RNA samples (100 ng) were poly-A selected and then reverse-transcribed into double-stranded cDNA with Actinomycin added during first-strand synthesis. The cDNA samples was fragmented, end-repaired and adenylated before ligation of TruSeq adapters. The adapters contain the index for multiplexing. Fragments containing TruSeq adapters on both ends were selectively enriched with PCR. The quality and quantity of the enriched libraries were validated using Qubit® (1.0) Fluorometer and the Bioanalyzer 2100 (Agilent, Waldbronn, Germany). The product is a smear with an average fragment size of approximately 360 bp. The libraries were normalized to 10nM in Tris-Cl 10 mM, pH8.5 with 0.1% Tween 20.
characteristics.characteristic: description
characteristics.text: normal
characteristics.ontologyTerms: http://purl.obolibrary.org/obo/PATO_0000461
characteristics.characteristic: disease
characteristics.text: CD11cCre/+ Tgfbr2fl/fl
characteristics.characteristic: genotype
characteristics.text: Mus musculus
characteristics.characteristic: organism
characteristics.text: Mus musculus
characteristics.ontologyTerms: http://purl.obolibrary.org/obo/NCBITaxon_10090
characteristics.characteristic: organism
characteristics.text: lung
characteristics.ontologyTerms: http://purl.obolibrary.org/obo/UBERON_0002048
characteristics.characteristic: organism part
characteristics.text: Tgfbr2 knockout in CD11c positive cells
characteristics.characteristic: phenotype
characteristics.text: Mus musculus
characteristics.characteristic: scientific_name
characteristics.text: mixed
characteristics.ontologyTerms: http://www.ebi.ac.uk/efo/EFO_0001271
characteristics.characteristic: sex
characteristics.text: C57BL/6
characteristics.characteristic: strain
characteristics.text: Sample 4
characteristics.characteristic: title