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Sample GSM673575 Query DataSets for GSM673575
Status Public on Apr 17, 2011
Title GFP-pos_P8_B_rep3
Sample type RNA
 
Source name GFP-positive cells from P8 retina in homozygous Tie2-GFP transgenic mice
Organism Mus musculus
Characteristics strain: FVB/N
genotype/variation: Tie2-GFP transgenic
age: postnatal day 8
tissue: retina
cell type: endothelial cell (GFP-positive)
Treatment protocol Eyes from homozygous Tie2-GFP transgenic mice were enucleated, rinsed with PBS, and kept in staining solution (PBS containing 2mM EDTA, 0.01%, NaN3, 5% fetal calf serum, 50U/ml penicillin, and 0.05mg/ml streptomycin). The retinas were dissected under stereomicroscope, pooled in staining solution, and minced into small pieces. Collected retinal pieces were rinsed twice in PBS and digested in 10 ml of papain/DNase solution (papain (33U/ml; Sigma-Aldrich, St. Louis, MI) in Dulbecco’s PBS (Invitrogen) containing L-cysteine (0.4mg/ml; Nacalai tesque, Kyoto, Japan), EDTA (0.50mM; Sigma), and DNase I (125U/ml; Sigma)) for 60 min at 37°C with gentle agitation. Following digestion, cells were pelleted and treated two times with DNase/ovomucoid solution (a solution containing ovomucoid (2mg/ml; Sigma), DNase I (125 U/ml; Sigma), and BSA (1mg/ml; Wako, Osaka, Japan)) in Dulbecco’s PBS for 60 min at 37°C with gentle agitation. Digested retinal cells were filtered through nylon mesh, centrifuged at 300x g for 5 min to pellet cells, resuspended again with 10ml of DNase/ovomucoid solution incubated at 37°C for 60 min. Cells were rinsed twice in staining solution, pelleted, resuspended with staining solution containing 5µg/ml of propidium iodide, and filtered through nylon mesh. These cells were analyzed and sorted by FACS Aria (Becton Dickinson, San Jose, CA).
Extracted molecule total RNA
Extraction protocol Total RNA was isolated using RNeasy Micro-Kit (QIAGEN) and was amplified using MessageAmpII aRNA Kit (Ambion, Austin, TX) to obtain aRNA (>60μg).
Label biotin
Label protocol Biotin-labeled cRNA was synthesized from the obtained RNA sample using the BioArray RNA Transcript labeling Kit (Affymetrix, Santa Clara, CA).
 
Hybridization protocol Biotin-labeled cRNA was fragmented in a 40-μl reaction mixture containing 40 mM Tris-acetate (pH 8.1), 100 mM potassium acetate, and 30mM magnesium acetate, and incubated at 94°C for 35 min, and then hybridized onto the MGU74v2 series of arrays.
Scan protocol GeneChips were scanned with a GeneArray Scanner (Hewlett-Packard, Santa Clara, CA), controlled by MicroSuite 5.1 software (Affymetrix).
Description Gene expression data from GFP-positive cells from P8 retina in homozygous Tie2-GFP transgenic mice
Data processing The dChip software (version 1.3) was used to normalize the CEL files at probe level and compute model-based expression values using the PM only model.
 
Submission date Feb 11, 2011
Last update date Apr 17, 2011
Contact name Sentaro Kusuhara
E-mail(s) kusu@med.kobe-u.ac.jp
Phone +81-78-382-6048
Organization name Kobe University Graduate School of Medicine
Department Department of Surgery
Lab Division of Ophthalmology
Street address 7-5-2 Kusunoki-cho, Chuo-ku
City Kobe
State/province Hyogo
ZIP/Postal code 650-0017
Country Japan
 
Platform ID GPL82
Series (1)
GSE27238 FACS-array profiling in retinal endothelial cells from living mouse retinas

Data table header descriptions
ID_REF
VALUE dChip normalized signal intensity

Data table
ID_REF VALUE
105967_at 107.18
105972_r_at 9.24
105975_at 130.17
105977_f_at 29.72
105978_at 20.95
105981_at 27.5
105991_at 30.33
105993_at 57.22
106004_at 92.85
106005_at 145.82
106006_at 45.67
106007_at 212.39
106008_at 191.17
106009_at 241.26
106010_at 31.83
106011_at 83.63
106012_at 148.97
106015_at 150.08
106016_at 5.25
106017_at 405.72

Total number of rows: 12411

Table truncated, full table size 198 Kbytes.




Supplementary file Size Download File type/resource
GSM673575.CEL.gz 2.6 Mb (ftp)(http) CEL
Processed data included within Sample table

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