|
| Status |
Public on Mar 23, 2016 |
| Title |
MGH_NCI-H1703_mutant KRAS_rep2 |
| Sample type |
RNA |
| |
|
| Source name |
NSCLC NCI-H1703 mutant KRAS
|
| Organism |
Homo sapiens |
| Characteristics |
genotype: mut p53, mut p16, wild type KRAS
derivation: stage 1 lung squamous cell carcinoma of a 54-year-old Caucasian male smoker.
cell line: NCI-H1703
|
| Treatment protocol |
NCI-H1703 clones maintained at 1 mg/mL puromycin.
|
| Growth protocol |
NCI-H1703 cells were grown in RPMI media with 10% BGS at 37°C.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted and purified using the miRNeasy kit (Qiagen Inc., Valencia, CA, USA) following the manufacturer's instructions including treatment with DNAase.
|
| Label |
Cy3
|
| Label protocol |
Labeling was performed by NimbleGen Systems Inc., Madison, WI USA, following their standard operating protocol. See www.nimblegen.com.
|
| |
|
| Hybridization protocol |
Hybridization was performed by NimbleGen Systems Inc., Madison, WI, USA following their standard operating protocol. See www.nimblegen.com.
|
| Scan protocol |
Scanning was performed by NimbleGen Systems Inc., Madison, WI USA, following their standard operating protocol. See www.nimblegen.com.
|
| Description |
This sample is of KRAS mutant NSCLC NCI-H1703 cells. It is the second of three wild-type biological replicates used in this experiment, each from separate cultures.
|
| Data processing |
We performed raw data quality control by principle component analysis and subsequent clustering by visualizing the distribution of expression levels to identify experimental outliers. After outlier removal, raw data files for each sample were normalized, background-corrected and saved to logarithmic scale using a Robust Multi-Array Analysis. Normalized data were analyzed and presented using R project. The normalized data from the experiments were fit into linear models using the Bioconductor package limma. Pair-wise comparisons of interest were performed using moderated t-statistics to test for significant differential expression. False discovery rate (FDR) at the .05 level for each probe was computed. Probes with p<0.05 and showing a fold change > 1 were considered differentially expressed. All probes were annotated with gene names. To better visualize the effect of differentially expressed genes, pathway analysis was performed on those transcripts that had a significance of P < 0.05. Enriched networks, pathways, cellular processes etc were generated through the use of IPA (Ingenuity® Systems) in all three contrasts. The Database for Annotation, Visualization and Integrated Discovery (DAVID) was used to annotate genes with chromosomal location in order to cluster highly repressed genes in mutant samples.
|
| |
|
| Submission date |
Mar 22, 2016 |
| Last update date |
Mar 23, 2016 |
| Contact name |
Meng Wang |
| E-mail(s) |
wang.meng@mgh.harvard.edu
|
| Phone |
6177268162
|
| Organization name |
Mass. General Hospital
|
| Department |
Radiation Oncology
|
| Lab |
Radiation Oncology
|
| Street address |
CNY149-4406 13th st Charlestown
|
| City |
Boston |
| State/province |
MA |
| ZIP/Postal code |
02129 |
| Country |
USA |
| |
|
| Platform ID |
GPL18943 |
| Series (1) |
| GSE79486 |
KRAS mutation (G12V) expression by pBABE-Puro vector in NCI-H1703 cells |
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