|
| Status |
Public on Sep 01, 2014 |
| Title |
Ly49C/I+ NK cells_rep2 |
| Sample type |
RNA |
| |
|
| Source name |
IL2 cultured splenocytes_Ly49C/I+
|
| Organism |
Mus musculus |
| Characteristics |
strain: C57BL/5
tissue: Spleen
cell type: natural killer
|
| Growth protocol |
Splenocytes were removed from C57BL/6 mice and were then incubated with antibodies against Thy1.2 and rabbit complement to eliminate T cells and were then cultured in media and 1000 IU IL-2 for 7 days.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Flow cytometric cell sorting was performed by gating on CD3-CD122+Ly49G2+Ly49C/I- versus CD3-CD122+Ly49G2-Ly49C/I+. Total mRNA was extracted using the Qiagen RNeasy kit. RNA quality was checked on an Agilent Bioanalyzer. All samples used for microarray analysis had a high quality score (RIN >9). 100 ng of RNA was reverse transcribed and amplified using the Ambion WT expression kit following manufacturer’s suggested protocol.
|
| Label |
Biotin
|
| Label protocol |
Sense strand cDNA was fragmented and labeled using Affymetrix WT terminal labeling kit.
|
| |
|
| Hybridization protocol |
Three replicates of each group were hybridized to Affymetrix mouse Gene ST 1.0 GeneChip in Affymetrix hybridization oven at 45C, 60rpm for 16 hrs.
|
| Scan protocol |
Wash and stain were performed on Affymetrix Fluidics Station 450 and scanned on Affymetrix GeneChip scanner 3000.
|
| Description |
IL2 cultured splenocytes sorted for Ly49C/I expression
|
| Data processing |
Data were collected using Affymetrix AGCC software. Statistical and clustering analysis was performed with Partek Genomics Suite software using the RMA normalization algorithm. Differentially expressed genes were identified with ANOVA analysis. Genes that were up- or down-regulated more than 1.5 fold and with a p<0.001 were considered significant. Significant genes were analyzed for enrichment for pathways using DAVID bioinformatics database (http://david.abcc.ncifcrf.gov/) and Ingenuity Pathway Analysis software.
Quantile normalized gene level expression values from GeneBASE
|
| |
|
| Submission date |
Apr 09, 2014 |
| Last update date |
Sep 01, 2014 |
| Contact name |
Stephen Kent Anderson |
| E-mail(s) |
andersonst@mail.nih.gov
|
| Phone |
301-846-1330
|
| Organization name |
NCI-Frederick
|
| Department |
CCR
|
| Lab |
Experimental Immunology
|
| Street address |
Building 560, room 31-33
|
| City |
Frederick |
| State/province |
MD |
| ZIP/Postal code |
21702-1201 |
| Country |
USA |
| |
|
| Platform ID |
GPL10740 |
| Series (1) |
| GSE56662 |
Expression data from murine natural killer cell subsets based on Ly49 receptor expression in C57BL/6 mice (C/I+ versus Ly49G2+ NK cells) |
|
