|
| Status |
Public on Jan 12, 2012 |
| Title |
Intestine_KO_biological rep2 |
| Sample type |
RNA |
| |
|
| Source name |
Mature proximal small intestine with intestinal epithelium-specific Pdx1 inactivation, first 5 cm immediately adjacent to the pylorus
|
| Organism |
Mus musculus |
| Characteristics |
gender: male
age: 6 months old
genotype: Pdx1flox/flox;VilCre (KO)
litter: brother of the same litter
|
| Treatment protocol |
Mice with intestinal epithelium-specific Pdx1 inactivation (Pdx1flox/flox;VilCre) were generated by intercross mating between VilCre and Pdx1flox/flox mouse strains. The VilCre mouse strain carries a Villin-Cre transgene and expresses Cre recombinase under the control of a 12.4 kb mouse villin 1 gene promoter fragment. Pdx1 exon 2 encoding the DNA-binding Homeodomain is flanked by loxP target sites in Pdx1flox/flox mice. Pdx1 expression is absent in the epithelium of mature Pdx1flox/flox;VilCre proximal small intestine.
|
| Growth protocol |
Mice were fed a standard diet (4.5 % fat, Prolab RMH 3000, LabDiet) and maintained under normal, non-stressed condition.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
The small intestine tissue was preserved in RNAlater (Qiagen, Valencia, CA) during harvest and before processing for RNA isolation. Total RNA was then extracted and treated with DNase I using the RNeasy Mini kit (Qiagen, Valencia, CA) according to manufacturer’s protocol.
|
| Label |
biotin
|
| Label protocol |
The total RNA samples were reverse transcribed into double-stranded cDNA, followed by an in vitro transcription (IVT) reaction that produced amplified amounts of biotin-labeled antisense mRNA (cRNA) using GeneChip 3' IVT Express Kit (Affymetrix, Santa Clara, CA), according to manufacturer’s recommendation.
|
| |
|
| Hybridization protocol |
The labeled cRNA prepared from each mouse was fragmented and hybridized to a single GeneChip Mouse Genome 430 2.0 Array (Affymetrix), according to manufacturer’s recommendation. The microarrays were washed, stained with fluorescent molecule SAPE that binds to biotin in combination of biotinylated anti-streptavidin antibody on GeneChip Fluidics Station 450 (Affymetrix).
|
| Scan protocol |
The arrays were scanned with GeneChip Scanner 3000 7G (Affymetrix).
|
| Description |
Mice were fed a standard diet (4.5 % fat, Prolab RMH 3000, LabDiet) and maintained under normal, non-stressed condition.
|
| Data processing |
The fluorescence intensity image files obtained from scanning each of the arrays were reviewed and processed initially by the Stanford PAN Facility with GeneChip Operating Software (Affymetrix). The resulting .CEL files were imported into Partek Genomics Suite (version 6.4, Partek Inc., St. Louis, MO) for probe set summarization and statistical analysis.
|
| |
|
| Submission date |
May 04, 2011 |
| Last update date |
Jan 12, 2012 |
| Contact name |
Chin Chen |
| E-mail(s) |
[email protected]
|
| Phone |
650-498-7599
|
| Fax |
650-724-3106
|
| Organization name |
Stanford University School of Medicine
|
| Department |
Pediatric Gastroenterology
|
| Lab |
Eric Sibley
|
| Street address |
300 Pasteur Drive
|
| City |
Stanford |
| State/province |
CA |
| ZIP/Postal code |
94305-5208 |
| Country |
USA |
| |
|
| Platform ID |
GPL1261 |
| Series (1) |
| GSE29048 |
Expression Profiling Identifies Novel Gene Targets and Functions for Pdx1 in the Duodenum of Mature Mice |
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