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Sample GSM673567 Query DataSets for GSM673567
Status Public on Apr 17, 2011
Title GFP-pos_P8_A_rep1
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 GPL81
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
92539_at 176.01
92540_f_at 48.59
92541_at 52.58
92542_at 800.68
92543_at 39.8
92544_f_at 3213.12
92545_f_at 45
92546_r_at 46.9
92547_at 66.76
92548_g_at 83.17
92549_at 82.51
92550_at 29.3
92551_at 114.9
92553_at 235.26
92554_at 97.3
92555_at 549.89
92556_at 21.85
92557_at 17.36
92558_at 175.11
92559_at 39.04

Total number of rows: 12422

Table truncated, full table size 194 Kbytes.




Supplementary file Size Download File type/resource
GSM673567.CEL.gz 2.6 Mb (ftp)(http) CEL
Processed data included within Sample table

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