 |
 |
GEO help: Mouse over screen elements for information. |
|
| Status |
Public on Dec 31, 2015 |
| Title |
P7_Naive |
| Sample type |
SRA |
| |
|
| Source name |
CD4+ T cells
|
| Organism |
Homo sapiens |
| Characteristics |
status: pre-T1D
barcode: AGCCTAAGCT
|
| Treatment protocol |
Fresh heparinised venous blood or buffy coats were processed by Ficoll-Paque® centrifugation (Amersham Pharmacia Biotech AB) to recover peripheral blood mononuclear cells (PBMCs). PBMCs were stored in liquid nitrogen prior to T cell subset sorting. CD4+ T cells were enriched with anti-human CD4 microbeads (clone M-T466) (Miltenyi Biotec) then labeled with the following antibodies: CD4-Pacific Blue (clone RPA-T4), CD45RA-APC (clone HI100), CD45RO-PE (clone UCHL1), CD27-FITC (clone M-T271), CCR7-Percp*Cy5.5 (clone 150503) (BD Bioscience) and CD25-PE-Cy7 (clone BC96) (eBiosciences). The cells were FACS-sorted into 6 subsets: naïve (CD45RA+ CD45RO- CD25-), resting Treg (rTreg) (CD45RA+ CD45RO- CD25+), activated Treg (aTreg) (CD45RA- CD45RO+ CD25 ++), central memory (Tcm) (CD45RA- CD45RO+ CD25lo/- CD27+ CCR7+), transitional memory (Ttm) (CD45RA- CD45RO+ CD25lo/- CD27+ CCR7-) and effector memory (Tem) (CD45RA- CD45RO+ CD25lo/- CD27- CCR7-).
|
| Growth protocol |
No cell culture involved
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted from sorted CD4+ T cell subsets with TRI Reagent (Ambion) following the manufacturer’s protocol with one modification: RNA was precipitated with ethanol at -80°C (instead of isopropanol on ice) in order to obtain all RNAs including miRNAs.
Small RNA libraries were constructed for sequencing on Illumina GAII/HiSeq platforms, essentially as described previously (18; 29), but with addition of barcoding. In order to multiplex the sequencing runs, one of four barcodes (AGCCTAAGCT, CATTGTTAGC, TCAACGGTAG or GTGCATCCTA) was added to the 5’ end of each library.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
size fractionation |
| Instrument model |
Illumina HiSeq 2000 |
| |
|
| Data processing |
Base calling
Barcode deconvolution and stripping
Alignment to mature miRNA database (miRBase)
Alignment of remaining reads to miRNA precursor database (miRBase)
Read abundance counting and normalization to library read size
Genome_build: miRBase 17
Supplementary_files_format_and_content: Excel spread sheet: miRNA identity and count
|
| |
|
| Submission date |
Feb 25, 2013 |
| Last update date |
May 15, 2019 |
| Contact name |
Mark Chong |
| E-mail(s) |
mchong@svi.edu.au
|
| Phone |
61-3-92313444
|
| Organization name |
St Vincent's Institute of Medical Research
|
| Street address |
9 Princes Street
|
| City |
Fitzroy |
| State/province |
Victoria |
| ZIP/Postal code |
3065 |
| Country |
Australia |
| |
|
| Platform ID |
GPL11154 |
| Series (1) |
| GSE44639 |
Altered microRNA expression in individuals at high risk of type 1 diabetes |
|
| Relations |
| SRA |
SRX245619 |
| BioSample |
SAMN01924905 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM1088244_P7_Naive.txt.gz |
9.6 Kb |
(ftp)(http) |
TXT |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
|
|
|
|
 |
