|
| Status |
Public on Oct 31, 2014 |
| Title |
Hypo-A2/12_control-3 |
| Sample type |
RNA |
| |
|
| Source name |
cell line HypoA-2/12, control, 24 h exposure, repliclate 3
|
| Organism |
Mus musculus |
| Characteristics |
cell line: HypoA-2/12
cell type: hypothalamic neuronal cells
phenotype: serotonin unresponsive
|
| Treatment protocol |
Cells were grown in monolayers to 90% confluency. Then medium was replaced by serum-free DMEM containing penicillin and streptomycin. After 4 hours, cells were exposed to serotonin.
|
| Growth protocol |
Murine derived hypothalamic neuronal cell lines hypoE-46 and hypoA2/12 (CELLutions Biosystems Inc. Canada) were grown and maintained in DMEM supplemented with 10% heat-inactivated fetal calf serum, 100 units/ml penicillin and 100μg/ml streptomycin at 37 C under 5.0% CO2.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was isolated using TrIzol (Life Technologies) according to manyfacturer's protocol. RNA integrity was checked on chip analysis (Agilent 2100 bioanalyzer, Agilent Technologies, Amsterdam, the Netherlands) according to the manufacturer's instructions. RNA was judged as suitable for array hybridization only if samples exhibited intact bands corresponding to the 18S and 28S ribosomal RNA subunits, and displayed no chromosomal peaks or RNA degradation products (RNA Integrity Number > 8.0).
|
| Label |
biotin
|
| Label protocol |
One hundred nanogram of RNA was used for whole transcript cDNA synthesis with the Ambion WT expression kit [catalog number 4411974] (Applied Biosystems/Life Technologies, Nieuwekerk a/d IJssel, The Netherlands).
|
| |
|
| Hybridization protocol |
Hybridization of 5.5μg labelled cDNA was done overnight for 17 hours, at 60 rpm, at 45ºC in a Hybridization Oven 640 (Affymetrix). The protocol was conducted as described in the Affymetrix Whole Transcript (WT) Sense Target Labeling Assay Manual, chapter 5 (P/N 701880, revision 5). Washing and staining of the arrays were done on an Affymetrix 450 fluidics station using the protocol FS450_0001, as described in the Affymetrix Whole Transcript (WT) Sense Target Labeling Assay Manual, chapter 5 (P/N 701880, revision 5).
|
| Scan protocol |
Arrays were scanned on an Affymetrix GeneTitan instrument (Affymetrix, Santa Clara, CA).
|
| Description |
G145_E09_15_177_m3_2.CEL (h177_control)
|
| Data processing |
Expression estimates were calculated applying the RMA algorithm in the Bioconductor library 'Oligo' (v1.26.6).
|
| |
|
| Submission date |
Apr 30, 2014 |
| Last update date |
Oct 31, 2014 |
| Contact name |
Guido Hooiveld |
| E-mail(s) |
[email protected]
|
| Organization name |
Wageningen University
|
| Department |
Div. Human Nutrition & Health
|
| Lab |
Nutrition, Metabolism & Genomics Group
|
| Street address |
HELIX, Stippeneng 4
|
| City |
Wageningen |
| ZIP/Postal code |
NL-6708WE |
| Country |
Netherlands |
| |
|
| Platform ID |
GPL11533 |
| Series (2) |
| GSE57189 |
Differences in food intake of tumour-bearing cachectic mice are associated with hypothalamic serotonin signalling [A358_GEO_hypocells] |
| GSE57190 |
Differences in food intake of tumour-bearing cachectic mice are associated with hypothalamic serotonin signalling |
|
