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Sample GSM5252568 Query DataSets for GSM5252568
Status Public on Apr 20, 2021
Title 197438
Sample type RNA
 
Source name Subcutaneous tumor
Organism Mus musculus
Characteristics disease: Melanoma
genotype: Hgftg;Cdk4R24C/R24C
mouse strain: B6.cBrd
treatment: Responders to 10mg/kg anti-mouse PD-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 9.783659155
1415671_at 11.57964227
1415672_at 11.70904286
1415673_at 9.70336264
1415674_a_at 9.991987859
1415675_at 9.326515413
1415676_a_at 11.54926635
1415677_at 8.758652994
1415678_at 10.14778087
1415679_at 11.3229061
1415680_at 8.463832825
1415681_at 9.906438851
1415682_at 8.145214061
1415683_at 11.10674794
1415684_at 8.176330235
1415685_at 8.889456427
1415686_at 10.22910951
1415687_a_at 12.8116761
1415688_at 11.01963614
1415689_s_at 8.695740379

Total number of rows: 45101

Table truncated, full table size 1026 Kbytes.




Supplementary file Size Download File type/resource
GSM5252568_RE714_20_197438.CEL.gz 3.5 Mb (ftp)(http) CEL
Raw data provided as supplementary file
Processed data included within Sample table

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