|
| Status |
Public on Dec 01, 2011 |
| Title |
WT vs Hypomethylated DRG Gene Expression |
| Sample type |
RNA |
| |
|
| Channel 1 |
| Source name |
DRG of E13.5 Wnt1-cre Dnmt1 mutant mice
|
| Organism |
Mus musculus |
| Characteristics |
genetic background: C57BL/6
age: Embryonic day 13.5
tissue: Dorsal root ganglion
genotype: Wnt1-cre Dnmt1 mutant
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Trizol Extracted and cleaned with Qiagen RNeasy minElute column and tested the quality of the RNA on a NanoChip (Agilent).
|
| Label |
Cy3
|
| Label protocol |
We converted the RNA into cDNA and then the cDNA into cRNA using the Agilent Low RNA Input Linear Amplification Kit (Agilent). We using a Nanodrop (Nanodrop) to quantify the labeled cRNA. Cy3 Labelled with Agilent Low Input Linear Amplification Kit
|
| |
|
| Channel 2 |
| Source name |
DRG of E13.5 WT mice
|
| Organism |
Mus musculus |
| Characteristics |
genetic background: C57BL/6
age: Embryonic day 13.5
tissue: Dorsal root ganglion
genotype: wild type
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Trizol Extracted and cleaned with Qiagen RNeasy minElute column and tested the quality of the RNA on a NanoChip (Agilent).
|
| Label |
Cy5
|
| Label protocol |
We converted the RNA into cDNA and then the cDNA into cRNA using the Agilent Low RNA Input Linear Amplification Kit (Agilent). We using a Nanodrop (Nanodrop) to quantify the labeled cRNA. Cy3 Labelled with Agilent Low Input Linear Amplification Kit.
|
| |
|
| |
| Hybridization protocol |
1ug of RNA for each channel was labelled using Agilent Low Input Linear Amplification protocol. Labelled cRNA mixed with Agilent 10x Control targets and Agilent 25x Fragmentation Buffer. This was incubated at 37C for 30 min and the reaction was stopped by the addition of 2x Agilent hybridization buffer. This mix was hybridized to arrays for 17 hours at 65C at 4rpms. Arrays were washed as per Agilent protocol and scanned immediately to prevent ozone degradation.
|
| Scan protocol |
The arrays were scanned with Agilent DNA microarray scanner (G2565BA), and probe features were extracted using Feature Extraction Software (version 8.5)
|
| Description |
Using the cre-loxP binary gene deletion strategy, we crossed female mice homozygous for the Dnmt1 conditional allele (Dnmt12lox (Fan et al., 2001; Jackson-Grusby et al., 2001)) with male mice carrying the Wnt1-cre insertion (Wnt1-cre) to generate both control, heterozygous, and mutant offspring in expected Mendelian ratios.
RNA samples were extracted from the DRG of WT and mutant mice at the embryonic day (E) 13.5.
|
| Data processing |
Arrays were processed using Agilent Feature Extraction (Version 8.5). Log 10 ratio were calculated from processed Cy3/ processed Cy5 signals. Replicates 1 - 4 were labeled - mutant mutant with Cy3 and WT with Cy5.
|
| |
|
| Submission date |
Jul 10, 2010 |
| Last update date |
Nov 15, 2012 |
| Contact name |
Masakazu Namihira |
| Organization name |
NAIST
|
| Department |
Graduate School of Biological Sciences
|
| Lab |
Laboratory of Molecular Neuroscience
|
| Street address |
8916-5 Takayama, Ikoma
|
| City |
Nara |
| ZIP/Postal code |
630-0101 |
| Country |
Japan |
| |
|
| Platform ID |
GPL7202 |
| Series (1) |
| GSE23530 |
WT vs Hypomethylated DRG Gene Expression |
|
