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Sample GSM5252574 Query DataSets for GSM5252574
Status Public on Apr 20, 2021
Title 197441
Sample type RNA
 
Source name Subcutaneous tumor
Organism Mus musculus
Characteristics disease: Melanoma
genotype: Hgftg;Cdk4R24C/R24C
mouse strain: B6.cBrd
treatment: Non-responders to 10mg/kg anti-mousePD-L1 mIgG1 D265A in PBS
Treatment protocol See publication Materials and Methods: Immunotherapy treatment
Growth protocol See publication Materials and Methods: Melanoma allograft model
Extracted molecule total RNA
Extraction protocol Tumor samples were lysed and homogenized for RNA isolation using the RNeasy Mini Kit following manufacturer’s protocol for purification of total RNA with on-column DNase digestion (Qiagen). RNA purity and concentration were determined spectrophotometrically, and RNA quality was assessed using the Agilent Technology Tape Station RNA Screen Tape.
Label Biotin
Label protocol A total input of 800ng of isolated RNA was applied to the Message Amp II-Biotin Enhanced aRNA Amplification Kit according to manufacture protocol for first and second Strand cDNA synthesis followed by in-vitro transcription (IVT) reaction to synthesize the complementary RNA (cRNA) for transcriptional amplification and biotin labeling.
 
Hybridization protocol A total of 20 ug of biotinylated cRNA was then fragmented for hybridization on the Affymetrix GeneChip Mouse Genome 430 2.0 Array.
Scan protocol GeneChip Operating Software Tool was used for data capture and quality
Data processing Raw CEL files from the scan were analyzed using a microarray pipeline written in ‘R’. Data were normalized using the Robust Multichip Average (RMA) method. Linear modeling was performed using Limma’s lmFit function (LIMMA, RRID:SCR_010943, Limma powers differential expression analyses for RNA-sequencing and microarray studies), and differential gene expression was determined using the contrasts.fit and eBayes functions. Top pathways for each comparison (Non-responders, Responders, and Relapse; normalized with control IgG treatment samples) were determined using (Gene Set Enrichment Analysis) GSEA (SeqGSEA, RRID:SCR_005724, http://software.broadinstitute.org/cancer/software/gsea/wiki/index.php/Gsea_Citation). The top 10 up and down regulated pathways were generated from Reactome (Reactome diagram viewer: data structures and strategies to boost performance) based on NES scores if Q-value meets threshold of less than 0.1. The top genes from Reactome were then used to run GSVA (Gene Set Variation Analysis for microarray and RNA-seq data) to find enrichment for each sample. Heatmaps were generated for the GSVA analysis, columns (samples) were sorted by group and rows (pathways) were clustered by Euclidean distance. Individual sample differential expression heatmap was analyzed by computing the log fold-change between a sample against the mean expression of the IgG samples, upper and lower limits shown on the heat map are log values from -2 to 2. Gene expression data from this report is available through the Collaborative Bioinformatics Resource of NIH, Center for Cancer Research (CCR). https://zenodo.org/record/3770853#.X9PmhhNKg_N.
 
Submission date Apr 19, 2021
Last update date Apr 20, 2021
Contact name abdalla abdelmaksoud
E-mail(s) abdalla.abdelmaksoud@nih.gov
Organization name NCI
Street address 37 Convent Dr
City bethesda
State/province maryland
ZIP/Postal code 20814
Country USA
 
Platform ID GPL1261
Series (1)
GSE172320 Gene Expression profile for the characterization of PD-L1 blockade in melanoma model

Data table header descriptions
ID_REF
VALUE RMA normalized

Data table
ID_REF VALUE
1415670_at 9.440748416
1415671_at 11.5695729
1415672_at 11.94745573
1415673_at 10.25297647
1415674_a_at 10.28698815
1415675_at 9.333084501
1415676_a_at 11.59448681
1415677_at 8.706261715
1415678_at 10.17666484
1415679_at 11.34550867
1415680_at 8.583080664
1415681_at 10.06008011
1415682_at 8.127361483
1415683_at 10.83942612
1415684_at 8.369822674
1415685_at 9.30851873
1415686_at 10.36229864
1415687_a_at 11.64852909
1415688_at 11.23189614
1415689_s_at 8.809017275

Total number of rows: 45101

Table truncated, full table size 1025 Kbytes.




Supplementary file Size Download File type/resource
GSM5252574_RE714_24_197441.CEL.gz 3.3 Mb (ftp)(http) CEL
Raw data provided as supplementary file
Processed data included within Sample table

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