|
| Status |
Public on Apr 01, 2012 |
| Title |
effect of GC-fed after SDS treatment rep4 |
| Sample type |
RNA |
| |
|
| Channel 1 |
| Source name |
two times SDS treated, dorsal skin of GC-fed mouse
|
| Organism |
Mus musculus |
| Characteristics |
tissue: skin
age: 5 weeks
strain: HR-1
gender: male
|
| Treatment protocol |
The dorsal skin on the right side of each mouse was treated with 10% SDS for five minutes daily for two days.
|
| Growth protocol |
5 weeks old male HR-1 mice were fed with GC-containing / non-containing AIN-93G (Oriental Yeast, Tokyo, Japan) rodent diet for 14 days.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA extracted using Isogen. They were further purified using a RNeasy Fibrous Tissue Mini Kit (QIAGEN, Valencia, CA) following manufacturer's instructions.
|
| Label |
Cyanine-5
|
| Label protocol |
500 ng of total RNA was reverse-transcribed to double-stranded cDNA using a poly dT-T7 promoter primer and MMLV-RT enzyme. cDNA products were used as templates for in vitro transcription to generate fluorescent cRNA using T7 RNA polymerase and Cy5-labeled or Cy3-labeled CTP. Labeled cRNAs were purified using QIAGEN RNeasy mini spin columns and eluted in nuclease-free water. cRNA quantity and cyanine incorporation were determined using a Nanodrop ND-1000 with Bioanalyzer.
|
| |
|
| Channel 2 |
| Source name |
two times SDS treated, dorsal skin of control-fed mouse
|
| Organism |
Mus musculus |
| Characteristics |
tissue: skin
age: 5 weeks
strain: HR-1
gender: male
|
| Treatment protocol |
The dorsal skin on the right side of each mouse was treated with 10% SDS for five minutes daily for two days.
|
| Growth protocol |
5 weeks old male HR-1 mice were fed with GC-containing / non-containing AIN-93G (Oriental Yeast, Tokyo, Japan) rodent diet for 14 days.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA extracted using Isogen. They were further purified using a RNeasy Fibrous Tissue Mini Kit (QIAGEN, Valencia, CA) following manufacturer's instructions.
|
| Label |
Cyanine-3
|
| Label protocol |
500 ng of total RNA was reverse-transcribed to double-stranded cDNA using a poly dT-T7 promoter primer and MMLV-RT enzyme. cDNA products were used as templates for in vitro transcription to generate fluorescent cRNA using T7 RNA polymerase and Cy5-labeled or Cy3-labeled CTP. Labeled cRNAs were purified using QIAGEN RNeasy mini spin columns and eluted in nuclease-free water. cRNA quantity and cyanine incorporation were determined using a Nanodrop ND-1000 with Bioanalyzer.
|
| |
|
| |
| Hybridization protocol |
Two labeled cDNA samples, one from GC-fed mouse and the other from control-fed mouse were combined. For each hybridization, 825 ng of labeled cRNAs were mixed, fragmented, and hybridized at 65°C for 17 hours to an Agilent 4x44K Whole Mouse Genome Microarray (Agilent 14868) in Agilent's G2545A hybridization oven.
|
| Scan protocol |
Scanned on an Agilent G2505B scanner.
|
| Description |
Biological replicate 4 of 4. 14 days after feeding with Gccontaine/non-GC diet, 2days after the begging of SDS treatment, treated two times
|
| Data processing |
Feature extraction software version 9.1.3.1 (Agilent Technologies) was used to assess fluorescent hybridization signals and to normalize signals using linear regression and a Lowess curve-fit technique.
Genespring (Agilent) was used further data processing. Feature extraction software version 9.1.3.1 (Agilent Technologies) was used to assess fluorescent hybridization signals and to normalize signals using linear regression and a Lowess curve-fit technique.Flagged signals were eliminated. Since some genes have multiple probes on the Agilent platform, gene-level expression values were calculated by using the "Gene-level experiment" function in Genespring. Expression values below 200 were floored to 200. Processed signal values of GC-fed mice were divided by processed signal values of references, and these values were used as expression ratio data. Log base 2 of expression ratio (log ratio) was used for further analysis. Grubb's method was used to eliminate outliers.
|
| |
|
| Submission date |
Mar 22, 2011 |
| Last update date |
Apr 01, 2012 |
| Contact name |
Ritsuro Ideta |
| E-mail(s) |
ritsuro.ideta@to.shiseido.co.jp
|
| Phone |
+81-45-788-7269
|
| Fax |
+81-45-788-7284
|
| Organization name |
Shiseido Research Center
|
| Department |
Functional Food Research & Development Center
|
| Lab |
Functional Food Appleide Research Group
|
| Street address |
2-12-1 Fukuura, Kanazawa-ku
|
| City |
Yokohama |
| ZIP/Postal code |
236-8643 |
| Country |
Japan |
| |
|
| Platform ID |
GPL7202 |
| Series (1) |
| GSE28086 |
Glucosylceramide fed mice skin before SDS treatment (0d), after SDS treatment (2d) |
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