|
| Status |
Public on Feb 01, 2012 |
| Title |
Primary and Secondary Motor Cortex_Stressed_Vinclozolin_biological rep2 |
| Sample type |
RNA |
| |
|
| Source name |
Primary and secondary motor cortex from F3-generation Vinclozolin lineage stressed rat brain
|
| Organism |
Rattus norvegicus |
| Characteristics |
rna prep date: 7-16-10
scan date: 7-23-10
vin/con rats dyad group: 5
strain: Sprague-Dawley rat
gender: Male
tissue: Primary and secondary motor cortex
developmental stage: Rat brain at day PND 120
|
| Treatment protocol |
Half of the F3-Vinclozolin/Control rats dyads were randomly chosen to be administered a chronic stress treatment. This stress paradigm entails 6 hours of daily restraint stress for 21 consecutive days (PND 23-43) in an apparatus consisted of a 25.4 square cm metal wire mesh folded in half and bound with a plastic mould. After the initial 21 days of stressing, all animals were left in the housing room constantly and were only removed for handling, weighing, and behavioural testing on scheduled days. All rats were sacrificed at age of 4 months (PND 120) and certain brain areas samples were collected as 2 mm tissue punches and put into Trizol™ reagent (Invitrogen, USA).
|
| Growth protocol |
Gestating outbred Sprague-Dawley mother rats were given intraperitoneal injections of Vinclozolin (100mg/kg/day) or vehicle (DMSO, Control) from embryonic day 8-14 (E8-E14) of gestation (i.e. F0 generation) [Cupp AS, et al., 2003]. Male rats of the F3 generation of Vinclozolin and Control lineages were selected out of litters from untreated F2 generation. One animal from each lineage (Control and Vinclozolin) were pair-housed (one control and one vinclozolin animal) and remained in these dyads throughout the duration of the study.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
RNA was isolated from tissue homogenates in 800 ul Trizol™ reagent (Invitrogen, USA), according to manufacturer’s instructions. 100 ul Trizol homogenates from 4 randomly chosen individuals within the same tissue/treatment group were pooled together for one RNA sample (one microarray biological replica). Samples from the same set of 4 dyads were pooled for one Vinclozolin or Control replica for all 4 brain areas, stressed or non-stressed. For microarray analysis, 3 biological replicas were prepared as above for each brain area/treatment group.
|
| Label |
biotin
|
| Label protocol |
Biotin-labeled ssDNA were prepared according to the standard Affymetrix protocol from at least 300 ng total RNA (GeneChip® Whole Transcript (WT) Sense Target Labeling Assay Manual, 2005-2209, Affymetrix).
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| |
|
| Hybridization protocol |
Following fragmentation, ssDNA were hybridized on Affymetrix Rat Gene 1.0 ST Array according to standard Affymetrix protocol. Chips were washed and stained in the Affymetrix Fluidics Station 450.
|
| Scan protocol |
GeneChips were scanned using Affymetrix GeneChip® Scanner 3000.
|
| Description |
Primary and secondary motor cortex from 120 days old F3-generation Vinclozolin lineage rat stressed at PND 23-43
|
| Data processing |
The data for each of the four studied brain areas (BLA, CRTX, CA1, and CA3) were pre-processed and analyzed as a separate group (4 groups, 12 chips per group) with Partek Genomic Suite 6.5 beta software (Partek Inc., St. Louis, MO) in August-October, 2010, using RMA, GC-content adjusted algorithm background correction, quintile normalization, median polish methods for probesets summarization, and log values of probes signals using base 2.
Logarithm of signal values, base 2, pre-processed with RMA, GC-content adjusted algorithm - each of the 4 bran areas separately. Scan date batch effect was removed if applicable.
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| |
|
| Submission date |
Jan 19, 2011 |
| Last update date |
Feb 01, 2012 |
| Contact name |
Michael K Skinner |
| E-mail(s) |
skinner@mail.wsu.edu
|
| Organization name |
WSU
|
| Department |
SBS
|
| Street address |
Abelson 507
|
| City |
Pullman |
| State/province |
WA |
| ZIP/Postal code |
99163 |
| Country |
USA |
| |
|
| Platform ID |
GPL6247 |
| Series (1) |
| GSE26737 |
Epigenetic Transgenerational Alterations to Stress Response in Brain Gene Networks and Behaviour |
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