|
| Status |
Public on Aug 24, 2022 |
| Title |
melanoma tumor (mouse under HFD diet)_HFD diet3 |
| Sample type |
RNA |
| |
|
| Source name |
melanoma tumor (mouse under HFD diet)_HFD diet3
|
| Organism |
Mus musculus |
| Characteristics |
tissue: melanoma tumor
|
| Treatment protocol |
Tumor cells ( 0.25x10^6 ) were injected subcutaneously (s.c) in the abdominal flank of mice fed with CD or HFHCD and harvested at day 9 post tumor graft.
|
| Growth protocol |
The melanoma B16F10 cells (ATCC CRL6475) were cultured in RPMI supplemented with 10% fetal bovine serum (FBS), 2 mM glutamine, 50 U/ml penicillin and 100μg/ml streptomycin.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Tumors were harvested, minced, lysed with 350ul RLT buffer containing 1% β-mercaptoethanol and homogenize by using a seringue and a needle ( 10 times). Then RNA were extracted with RNAeasy microkit (Qiagen) according to manufactor's instructions. Total RNA were measured on Nanodrop and store at -80 until analysis.
|
| Label |
Biotin
|
| Label protocol |
100 ng of total RNA is reverse transcribed following the GeneChip® WT Plus Reagent Kit (Thermofisher). Briefly, the resulting double strand cDNA is used for in vitro transcription with T7 RNA polymerase (all these steps are included in the WT cDNA synthesis and amplification kit of Thermofisher).
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| |
|
| Hybridization protocol |
After purification according to Thermofisher protocol, 5.5 ug of Sens Target DNA are fragmented and biotin labelled. After control of fragmentation using Bioanalyzer 2100, cDNA is then hybridized to GeneChip® MouseGene 2.0ST (Affymetrix) at 45°C for 17 hours.
|
| Scan protocol |
After overnight hybridization, chips are washed on the fluidic station FS450 following specific protocols (Affymetrix) and scanned using the GCS3000 7G
|
| Data processing |
The scanned images are then analyzed with Expression Console software (Thermofischer) to obtain raw data (cel files) and metrics for Quality Controls. The observations of some of these metrics and the study of the distribution of raw data show no outlier experiment. RMA normalization is performed using R with CDF v21 of EntrezGene (Brain Array).
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| |
|
| Submission date |
Aug 16, 2022 |
| Last update date |
Aug 24, 2022 |
| Contact name |
Franck Letourneur |
| E-mail(s) |
franck.letourneur@inserm.fr
|
| Organization name |
INSERM
|
| Department |
U1016
|
| Lab |
genomic
|
| Street address |
22 rue mechain
|
| City |
Paris |
| ZIP/Postal code |
75014 |
| Country |
France |
| |
|
| Platform ID |
GPL23092 |
| Series (1) |
| GSE211392 |
Transcriptomic analysis of total RNA from melanoma tumor, originating from control and hyperlipic diet |
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