|
| Status |
Public on Mar 29, 2019 |
| Title |
CNOT6L WT hepatocytes for IP, biological rep2 |
| Sample type |
RNA |
| |
|
| Source name |
Immunoprecipitated RNA of primary hepatocytes from CNOT6L WT mice
|
| Organism |
Mus musculus |
| Characteristics |
gender: Male
strain: C57BL/6J
genotype: Wild-type
tissue: Isolated primary hepatocytes
|
| Growth protocol |
Primary hepatocytes were isolated from 8-week-old wild-type and Cnot6l KO mice.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
RNA were extracted with Trizol Reagent and GlycoBlue Coprecipitant (Invitrogen) and purified with an RNeasy Kit (Qiagen) according to the manufacturer’s instructions. The quality of RNA was determined using the Bioanalyzer nano chip (Agilent Technologies, Inc.).
|
| Label |
Biotin
|
| Label protocol |
Biotinylated cRNA were prepared according to the standard Affymetrix protocol from 10 ug total RNA.
|
| |
|
| Hybridization protocol |
Following fragmentation, 20 ug of cRNA were hybridized for 16hr at 45C on Gene Chip Mouse Genome Array. GeneChips were washed and stained in the Affymetrix Fluidics Station 400.
|
| Scan protocol |
GeneChips were scanned using the Hewlett-Packard GeneArray Scanner.
|
| Description |
Immunoprecipitated RNA data from wild-type primary hepatocytes
|
| Data processing |
For analysis of microarray data, we extracted and normalized data using robust multi-array averaging implemented in the R package “affy” (www.r-project.org). We used updated probe set definitions because these provide improved precision and accuracy (Sandberg and Larsson, 2007). To identify immunoprecipitated mRNAs, we used analysis of partial variance (APV) and applied the random variance model (RVM) as implemented in the anota R package (Larsson et al., 2010, 2011). In anota we applied the following settings (as defined in the anotaPlotSigGenes function) for gene selection: slopeP=0.01; maxSlope=2; minSlope=(-2); selDeltaP=0.5; selDeltaPT=0.5 and maxRvmP=0.01. This identified genes showing enrichment in immunoprecipitated mRNA from WT cells independent of steady-state mRNA level. The resulting genes were further filtered to identify those enriched in immunoprecipitated mRNA from WT cells (i.e. removed those that were depleted).
|
| |
|
| Submission date |
Oct 15, 2014 |
| Last update date |
Mar 29, 2019 |
| Contact name |
Masahiro Morita |
| Organization name |
University of Texas Health Science Center at San Antonio
|
| Department |
Department of Molecular Medicine
|
| Lab |
Morita lab
|
| Street address |
4939 Charles Katz Drive
|
| City |
San Antonio |
| State/province |
Texas |
| ZIP/Postal code |
78229 |
| Country |
USA |
| |
|
| Platform ID |
GPL7546 |
| Series (1) |
| GSE62365 |
RIP-CHIP data from CNOT6L WT and KO hepatocytes |
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