|
| Status |
Public on Feb 02, 2011 |
| Title |
Abcb4 knockout mouse treated with norUDCA for 3 days 2 |
| Sample type |
RNA |
| |
|
| Source name |
mouse liver RNA
|
| Organism |
Mus musculus |
| Characteristics |
strain: FVB/N
age: 3 months
gender: male
tissue: liver
genotype: Abcb4 KO
treatment: side-chain modified bile acid "norUDCA" for 3 days
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Tissues were immediately snap-frozen in pre-chilled 2-methylbutane after excision and stored in liquid nitrogen. RNA was isolated using TrizolTM reagent according to the manufacturer’s protocol. RNA samples were analyzed using Agilent Bioanalyzer Lab-on-chip Nano 6000 to determine integrity, quality, and concentration of the samples. Only samples fulfilling the quality criteria (Ratio of 28S to 18S rRNA of >2.0) were used for further microarray analysis.
|
| Label |
Applied Biosystems Chemiluminescent RT Labeling Kit
|
| Label protocol |
Each microarray was first pre-hybridized at 55°C for 1 hour in hybridization buffer with blocking reagent. Oligo(dT)-primed, DIG-labeled cDNA targets were fragmented, mixed with internal control target.
|
| |
|
| Hybridization protocol |
Microarrays were hybridized in a volume of 1.5 ml at 55°C for 16 hours. After hybridization, arrays were washed with hybridization wash buffer and chemiluminescence rinse buffer. Enhanced chemiluminescent signals were generated by incubating arrays with alkaline phosphatase conjugated anti-digoxigenin antibody followed by incubation with chemiluminescence enhancing solution and a final addition of chemiluminescence substrate.
|
| Scan protocol |
Four images were collected for each microarray using the ABI 1700 Chemiluminescent Microarray Analyzer.
|
| Data processing |
Images were auto-gridded and the chemiluminescent signals were quantified, corrected for background and spot, and spatially normalized.
Expression values were imported into GeneSpring 6.2.1 (Silicon Genetics, Redwood, CA, USA) and processed according to Applied Biosystems standard procedure ("per chip" and "per gene" normalization). Data were filtered using the Cross-Gene Error Model (CEM); S/N >3 and Flag = 0.
|
| |
|
| Submission date |
Jan 11, 2011 |
| Last update date |
Feb 02, 2011 |
| Contact name |
Karin Wagner |
| E-mail(s) |
kar.wagner@medunigraz.at
|
| Organization name |
Medical University Graz
|
| Department |
Molecular Biology
|
| Street address |
Stiftingtalstrasse 24
|
| City |
Graz |
| State/province |
Styria |
| ZIP/Postal code |
8010 |
| Country |
Austria |
| |
|
| Platform ID |
GPL2995 |
| Series (1) |
| GSE26547 |
Molecular and metabolic effects of norUDCA in Abcb4 knockout mice |
|
