|
| Status |
Public on May 09, 2014 |
| Title |
Ttf+/+T43H_pre_pach_rep3 |
| Sample type |
RNA |
| |
|
| Source name |
Ttf+/+T43H, pre-mid-pachytene spermatocytes
|
| Organism |
Mus musculus |
| Characteristics |
strain: C57BL/10J-Ttf+/+T43H
gender: male
age: adult (2 months)
tissue: pre-mid-pachytene spermatocytes
|
| Treatment protocol |
Mice were euthanized by cervical dislocation. For isolation of spermatogenic populations the single cell suspension from testicular tubules was prepared and the populations were isolated using fluorescence activated cell sorting of testicular single cell suspension according to ref. (Bastos et al. 2005). Testicular RNA from 15 days old animals was isolated from the testes after removal of tunica.
|
| Growth protocol |
Mouse lines used in the experiment were C57BL/6J , C57BL/10J, C57BL/10J -Ttf+/+T43H and C57BL/10J - t12+/+T43. All the male mice used for isolation of spermatogenic populations were two months old. 15 days old males were used for comparison of testicular expression.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA from individual populations was isolated using RNeasy Micro kit (Qiagen) according to manufacturer‘s protocol. Testicular RNA from 15 days old males was isolated using RNeasy Mini isolation kit (Qiagen) according to manufacturer‘s protocol.
|
| Label |
biotin
|
| Label protocol |
Between 20 to 30 ng of total RNA were used to prepare biotinylated cRNA according to the standard Affymetrix Two-Cycle protocol (Expression Analysis Technical Manual, 701021 Rev. 5, Affymetrix).
|
| |
|
| Hybridization protocol |
Following fragmentation, 15 ug of cRNA were hybridized for 16 hr at 45°C on GeneChip Mouse Genome 430 2.0 Array. GeneChips were washed and stained in the Affymetrix Fluidics Station 450.
|
| Scan protocol |
GeneChips were scanned using the GeneChip Scanner 3000 7G.
|
| Data processing |
The data were analyzed independently for spermatogenic populations and testicular samples from 15 days old animals. The analysis was done with Bioconductor and R statistics (version 2.12) using the gcRMA method with default settings and custom chip description file (probes were annotated to Entrez identifiers - NCBI build 37) mm430mmentrezg, created by Microarray Lab at the Molecular and Behavioral Neuroscience Institute at the University of Michigan.
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| |
|
| Submission date |
May 10, 2011 |
| Last update date |
May 09, 2014 |
| Contact name |
David Homolka |
| E-mail(s) |
david.homolka@img.cas.cz
|
| Organization name |
Institute Of Molecular Genetics
|
| Department |
Department of Mouse Molecular Genetics
|
| Street address |
Videnska 1083
|
| City |
Prague 4 |
| ZIP/Postal code |
142 20 |
| Country |
Czech Republic |
| |
|
| Platform ID |
GPL7546 |
| Series (1) |
| GSE29177 |
Meiotic Consequences of Genetic Extension of Unsynapsed Autosomal Chromatin |
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