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Status |
Public on Feb 20, 2020 |
Title |
TP53-KO-ASTROCYTE-rep1 |
Sample type |
RNA |
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Source name |
TP53-KO-ASTROCYTE
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Organism |
Mus musculus |
Characteristics |
strain background: C57BL/6J genotype/variation: TP53 knocked-out mice tissue: brain cell type: Astrocyte cells
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Extracted molecule |
total RNA |
Extraction protocol |
Total RNA was isolated using TRIzol (Life Technologies) following the manufacturer's recommendations. RNA was quantified using a NanoDrop-2000.
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Label |
Cy3
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Label protocol |
Cyanine-3 (Cy3) labeled cRNA was prepared from 200 ng RNA using Agilent One-Color Microarray-Based Gene Expression Analysis protocol (Agilent Technologies) according to the manufacturer's instructions, followed by RNAeasy column purification (QIAGEN). Dye incorporation and cRNA yield were checked with the NanoDrop 2000 Spectrophotometer.
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Hybridization protocol |
600 ng of Cy3-labelled cRNA (specific activity >10.0 pmol Cy3/ug cRNA) was fragmented at 60°C for 30 minutes in a reaction volume of 25 ul containing 1x Agilent fragmentation buffer and 2x Agilent blocking agent following the manufacturers instructions. On completion of the fragmentation reaction, 25 ul of 2x Agilent hybridization buffer was added to the fragmentation mixture and hybridized to SurePrint G3 Mouse GE 8×60K array slides (G4858A, Agilent Technologies) at 65 °C with rotation at 10 rpm for 17 hours in a rotating Agilent hybridization oven. After hybridization, microarrays were washed 1 minute at room temperature with GE Wash Buffer 1 (Agilent Technologies) and 1 minute with 37°C GE Wash buffer 2 (Agilent Technologies), then dried immediately by brief centrifugation.
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Scan protocol |
Slides were scanned immediately after washing on the Agilent Microarray Scanner (G2565BA, Agilent Technologies) using one color scan setting for 8x60k array slides.
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Description |
TP53-KO-ASTROCYTE#1
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Data processing |
The scanned images were analyzed with Feature Extraction Software 10.7.3.1 (Agilent) using default parameters (protocol GE1_107_Sep09 and Grid: 039494_D_F_20150612) to obtain background subtracted and spatially detrended Processed Signal intensities. Features flagged in Feature Extraction as Feature Non-uniform outliers were excluded.
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Submission date |
Feb 21, 2018 |
Last update date |
Feb 22, 2020 |
Contact name |
Keisuke Katsushima |
E-mail(s) |
kkeisuke@med.nagoya-cu.ac.jp
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Phone |
+81-052-853-8196
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Organization name |
Nagoya City University Graduate School of Medical Sciences
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Department |
Department of Epigenomics
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Lab |
Yutaka Kondo
|
Street address |
1 Kawasumi, Mizuho-cho, Mizuho-ku
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City |
Nagoya |
ZIP/Postal code |
467-8601 |
Country |
Japan |
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Platform ID |
GPL10787 |
Series (2) |
GSE110927 |
Expression analysis of MADM based glioma mouse model and normal brain |
GSE110937 |
ChIP and expression analysis of MADM based glioma mouse model |
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