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Sample GSM1586248 Query DataSets for GSM1586248
Status Public on Jan 04, 2019
Title Whole lung, Control 3
Sample type RNA
 
Source name Whole lung_Control
Organism Mus musculus
Characteristics strain background: C57Bl/6
genotype/variation: wild type
gender: female
age: 8-16 weeks
tissue: Lung
Treatment protocol Healthy wild type C57BL/6 female mice were sacrificed to obtain the small intestine, lungs, liver and peripheral blood (as reference). Organs were disintegrated on a Gentle MACS Octo Dissociator using the Mouse Lamina Propria Dissociation, Mouse Lung Dissociation, and Mouse Tumor Dissociation kits, respectively (all from Miltenyi Biotec), according to the manufacturer’s instructions. Crude lysates were subjected to 40%/80% Percoll gradient centrifugation (Sigma Aldrich). Then, CD8b+ CD103+ Trm cells were isolated using Miltenyi’s AutoMACS Pro magnetic sorter and Multisort MACS kits, performing two subsequent rounds of positive selection. First, cells were labeled with anti-CD8b-biotin (eBioscience) and anti-CD103-PE (BD Biosciences) antibodies. Next, CD8b+ T cells were positively selected with Anti-Biotin Multisort Microbeads (Miltenyi Biotec). Microbeads were released, the cells incubated with Anti-PE Microbeads (Miltenyi Biotec), and subjected to a second round of positive selection to retrieve the CD103+ subset. As a reference, peripheral blood-derived, non-naïve CD8b+ CD62L- T cells were also isolated from the same animals by CD8b+ T cell separation and subsequent depletion of CD62L+ cells with anti-mouse CD62L microbeads (Miltenyi Biotec). As an additional control, whole organ samples were used (n=3), which were not isolated after organ disintegration.
Extracted molecule total RNA
Extraction protocol Total RNA was extracted from MACS-separated T cells using the RNeasy Plus Micro Kit (Qiagen) following the manufacturer's recommendations. RNA yield was measured and integrity checked on an Agilent 2100 Bioanalyzer (Agilent Technologies).
Label Cy3
Label protocol Three hundred pg of total RNA, with an RNA integrity number >8.0 per sample was reverse transcribed, amplified in two rounds, and Cy3 labeled by the Arcturus RiboAmp HS PLUS, Cy3 Kit (Life Technologies) according to the manufacturer's instructions. Dye incorporation and cRNA yield were checked with the NanoDrop ND-1000 Spectrophotometer.
 
Hybridization protocol Amplified samples were hybridized to 4x44k mouse whole genome microarrays using the Gene Expression Hybridization Kit (Agilent)
Scan protocol Samples were scanned on an Agilent Microarray Scanner using one color scan setting for 4x44k array slides.
Description 29
Data processing The scanned images were analyzed with Feature Extraction Software 10.7 (Agilent). Raw data were quantile normalized and further analyzed by Partek GS (Partek). To identify T-cell transcripts, whole organs were also processed in parallel, and used to identify mRNAs enriched in respective T cell samples (ANOVA with FDR Correction, p<0.005, >2 fold change). To assess organ-specific differences in T cell gene expression profiles, these 6411 transcripts were subjected to multidimensional scaling (MDS), ANOVA or t-tests, as appropriate (with FDR Correction, p<0.05), to identify differentially expressed genes (DEGs).
 
Submission date Jan 16, 2015
Last update date Jan 04, 2019
Contact name Nikolett Lupsa
E-mail(s) lupsa.nikolett@gmail.com
Organization name Semmelweis University
Department Genetics, Cell-and Immunbiology
Lab MTA-SE Lendület Research Group
Street address Nagyvárad tér 4
City Budapest
State/province Pest
ZIP/Postal code 1085
Country Hungary
 
Platform ID GPL11202
Series (2)
GSE65044 Characterization of resident CD8+ Trm cell populations using whole-genome biomarker analysis
GSE65045 Comparative analysis of murine CD8+ resident memory and effector T cells in various organ environments

Data table header descriptions
ID_REF
VALUE log2 quantile normalized signal intensity

Data table
ID_REF VALUE
A_55_P1989846 8.27819
A_55_P1991598 8.28377
A_55_P2022211 15.8162
A_55_P1980764 9.90482
A_55_P1964375 11.7861
A_51_P128876 15.2071
A_55_P2121042 8.30764
A_52_P219230 8.31319
A_51_P207591 12.8549
A_55_P2131920 8.59597
A_55_P2404223 8.32629
A_55_P2101944 15.3105
A_52_P358860 10.0649
A_51_P119031 8.53973
A_51_P309854 8.24649
A_51_P343900 12.1754
A_51_P234359 8.34988
A_51_P487813 13.2457
A_52_P613977 11.122
A_55_P1957209 8.35766

Total number of rows: 39429

Table truncated, full table size 827 Kbytes.




Supplementary file Size Download File type/resource
GSM1586248_US22502634_252665515146_S01_GE1_1100_Jul11_1_1.txt.gz 2.3 Mb (ftp)(http) TXT
Processed data included within Sample table

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