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Sample GSM837608 Query DataSets for GSM837608
Status Public on Nov 23, 2011
Title HIV-1 3 [miRNA]
Sample type RNA
 
Source name PBMC
Organism Homo sapiens
Characteristics tissue: Blood
infection: HIV-1
Treatment protocol One half of the cells were subsequently infected with 0.1 MOI of virus particles using standard protocols as described and the remaining half of the culture is maintained under similar conditions and used as control. Seven days post infection (pi), cells were collected and frozen for total RNA isolation.
Growth protocol We purchased normal donor blood from the American Red Cross Blood Bank in Pittsburgh using appropriate IRB approval forms from the University of Pittsburgh. PBMCs were isolated by Ficoll-Hypaque gradient centrifugation. Freshly isolated normal donor PBMCs (5×106/mL) were stimulated with 5 µg/ml PHA-P (Sigma, St. Louis, MO) for three days. Cells were washed, divided into two parts and cultured in RPMI medium (GIBCO, CA) containing 10% FBS (Hyclone, Logan, UT), 1% L-glutamine (Cambrex, MD), 1% penicillin-streptomycin (GIBCO, CA), and IL-2 (200 U/mL, Chiron, Emoryville, CA).
Extracted molecule total RNA
Extraction protocol Total RNA was isolated using the TRIzol method (InVitrogen) as suggested by the manufacturer. Next, to enhance the sensitivity and detection, we enriched the small RNA using a microRNA isolation kit (SABiosciences) with two separating columns as per the manufacturer's instructions.
Label SYBR Green/Rox
Label protocol One ug of total RNA enriched for miRNA was reverse transcribed to cDNA using SA Biosciences RT2 miRNA first strand Kit (MA-03). Then the cDNA product along with SA Biosciences RT2 Real-time SYBR Green/Rox PCR master mix (PA-012) was used for detection.
Cycles Duration & Temperature 1 10 minutes at 95°C 40 15 seconds at 95°C -> 40 seconds at 60°C -> 30 seconds at 72°C
 
Hybridization protocol n/a
Scan protocol n/a
Description Infected PBMC from donor 3
Data processing Raw Ct values obtained after the 7900 HT RT-PCR run were uploaded onto SA Biosciences online analysis tool to generate fold changes for differential expression of miRNA in Infected group i.e. HIV-1 (N=6) over Uninfected group i.e Control group (N=6).
 
Submission date Nov 21, 2011
Last update date Nov 30, 2011
Contact name Pruthvi S. R. Nagilla
E-mail(s) [email protected]
Phone 412 624-3062
URL http://www.idm.pitt.edu/directory/bios/ayyavoo.asp
Organization name University of Pittsburgh
Department Infectious Diseases and Microbiology
Street address 416 Parran Hall, 130 DeSoto Street
City Pittsburgh
State/province PA
ZIP/Postal code 15261
Country USA
 
Platform ID GPL14907
Series (2)
GSE33837 Comparative Expression Profile of miRNA and mRNA in Primary Peripheral Blood Mononuclear Cells Infected with Human Immunodeficiency Virus (HIV-1) [miRNA]
GSE33879 Comparative Expression Profile of miRNA and mRNA in Primary Peripheral Blood Mononuclear Cells Infected with Human Immunodeficiency Virus (HIV-1)

Data table header descriptions
ID_REF
VALUE Ct value

Data table
ID_REF VALUE
1 29.63348
2 30.413052
3 31.979336
4 21.406752
5 21.184765
6 31.810959
7 30.834778
8 31.909506
9 33.593647
10 32.475506
11 31.51313
12 24.139013
13 28.442263
14 32.656475
15 32.458496
16 34.554028
17 32.784416
18 31.344479
19 30.13908
20 29.425592

Total number of rows: 768

Table truncated, full table size 10 Kbytes.




Supplementary data files not provided
Processed data are available on Series record

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