|
| Status |
Public on May 20, 2009 |
| Title |
histone H4 acetylation, saline treatment, replicate 1 |
| Sample type |
genomic |
| |
|
| Channel 1 |
| Source name |
AcH4 immunoprecipitation, saline
|
| Organism |
Mus musculus |
| Characteristics |
antibody: anti-acetylated K5/K8/K12/K16 H4
strain: C57/BL6
gender: male
age: 10-12 weeks
tissue: brain
brain region: nucleus accumbens
treatment: saline
|
| Treatment protocol |
7 days of saline or cocaine (20mg/kg).
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
Fresh nucleus accumbens (NAc) punches were cross-linked and lysed, and the chromatin was fragmented by sonication to an average length of 700 bp. For each sample, bilateral NAc punches were pooled from ~8 mice. Aliquots of sonicated chromatin were incubated with 4 µg of the following polyclonal antibodies for 8 hr at 4°C: anti-acetylated K9/K14 H3 (cat. #06-599, Upstate/Millipore, Billerica, MA,) or anti-acetylated K5/K8/K12/K16 H4 (cat. #06-866, Upstate). Immunprecipitates were reverse cross-linked, and both the immunoprecipitated (enriched) and total (input or non-enriched) DNA from each treatment group was amplified and labeled with a fluorescent dye (Cy5 and Cy3) with the use of ligation-mediated-polymerase chain reaction (Sikder et al., JBC 2006).
|
| Label |
Cy5
|
| Label protocol |
NimbleGen standard protocol.
|
| |
|
| Channel 2 |
| Source name |
input
|
| Organism |
Mus musculus |
| Characteristics |
strain: C57/BL6
gender: male
age: 10-12 weeks
tissue: brain
brain region: nucleus accumbens
treatment: saline
|
| Treatment protocol |
7 days of saline or cocaine (20mg/kg).
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
Fresh nucleus accumbens (NAc) punches were cross-linked and lysed, and the chromatin was fragmented by sonication to an average length of 700 bp. For each sample, bilateral NAc punches were pooled from ~8 mice. Aliquots of sonicated chromatin were incubated with 4 µg of the following polyclonal antibodies for 8 hr at 4°C: anti-acetylated K9/K14 H3 (cat. #06-599, Upstate/Millipore, Billerica, MA,) or anti-acetylated K5/K8/K12/K16 H4 (cat. #06-866, Upstate). Immunprecipitates were reverse cross-linked, and both the immunoprecipitated (enriched) and total (input or non-enriched) DNA from each treatment group was amplified and labeled with a fluorescent dye (Cy5 and Cy3) with the use of ligation-mediated-polymerase chain reaction (Sikder et al., JBC 2006).
|
| Label |
Cy3
|
| Label protocol |
NimbleGen standard protocol.
|
| |
|
| |
| Hybridization protocol |
NimbleGen standard protocol. Samples of Cy5-labeled immunoprecipitated DNA and Cy3-labeled input DNA were mixed and hybridized to NimbleGen (Madison, WI) MM5 mouse promoter microarrays.
|
| Scan protocol |
NimbleGen standard protocol.
|
| Description |
H4S1
|
| Data processing |
The log2 ratios of signal intensities between the immunoprecipitated samples and control genomic DNA (input).These ratios were scaled by Tukey's Bi-weight method.
|
| |
|
| Submission date |
May 20, 2009 |
| Last update date |
May 21, 2009 |
| Contact name |
guanghua xiao |
| E-mail(s) |
guanghua.xiao@utsouthwestern.edu
|
| Phone |
(214)6484553
|
| Organization name |
University of Texas Southwestern Medical Center
|
| Department |
Psychiatry
|
| Lab |
Eric Nestler
|
| Street address |
5323 Harry Hines Blvd
|
| City |
Dallas |
| State/province |
TX |
| ZIP/Postal code |
75390 |
| Country |
USA |
| |
|
| Platform ID |
GPL7496 |
| Series (2) |
| GSE15968 |
Genome-Wide Promoter Analysis of Epigenetic Regulation by Cocaine |
| GSE16183 |
Genome-Wide Promoter Analysis of Epigenetic Regulation by Cocaine (MM5 data) |
|
