|
| Status |
Public on Jul 14, 2015 |
| Title |
HITSCLIP-HIV- (3 donor mix) |
| Sample type |
SRA |
| |
|
| Source name |
monocyte-derived macrophages, noninfected, Ago2 IP
|
| Organism |
Homo sapiens |
| Characteristics |
donor: mix of 3 donors
cell type: monocyte-derived macrophages
infection: none
antibody: anti-AGO2, clone 11A9 (Sigma)
|
| Treatment protocol |
After another week, the differentiated macrophages were infected with HIV-1JR-FL with a multiplicity of infection (MOI) of 0.1. The cells were washed the next day with PBS supplemented with 2% FCS.
|
| Growth protocol |
Primary human monocytes from HIV-1 negative blood donors were isolated with anti-CD14 coated magnetic beads (Miltenyi Biotech) from buffy coats and cultured one week in RPMI supplemented with 1% penicillin/streptomycin, 10% human serum (Sigma) and 0.02 µg/ml M-CSF (macrophage colony stimulating factor). Seven days after isolation, the macrophages were washed in PBS supplemented with 2% FCS to remove not adherent and dead cells and the medium was replaced and cultured in RPMI supplemented with 1% penicillin/streptomycin and 10% FCS.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Ago2 immunoprecipitation and PAR-CLIP (as described in Hafner et al., 2010; Kishore et al., 2012; Imig et al., 2015). Small RNA sequencing: Trizol-based total RNA isolation.
According to Kishore et al., Nat Meth, 2011.
Single end reads, 5'-3' adapter, multiplexed barcoded.
|
| |
|
| Library strategy |
RIP-Seq |
| Library source |
transcriptomic |
| Library selection |
other |
| Instrument model |
Illumina HiSeq 2000 |
| |
|
| Description |
HITS-CLIP
Argonaut2 immunoprecipitation
Processed data file: HITSCLIP-Summary.xlsx
|
| Data processing |
Adapters sequences removed from all samples using fastX toolkit.
All samples were aligned to human and HIV genomes simultaneously using Bowtie 1.0.
PAR-CLIP samples were analyzed using Paralyzer tool.
Gene expression was quantified using HTseq count tool for sRNA-seq samples.
HITS-CLIP samples were used for checking the reads on HIV genome using manual scripts.
Genome_build: Human: GRCh37.72
Genome_build: HIV-1: HIV-1JR-FL (GenBank: U63632.1) and the LTR sequences derived from HIV-1JRCSF (GenBank: M38429.1).
Supplementary_files_format_and_content: Tab-delimited text files containing Paralyzer output (PAR-CLIP samples), raw gene expressions (sRNA-seq), and coverage on HIV genome (HITS-CLIP).
|
| |
|
| Submission date |
Jul 13, 2015 |
| Last update date |
May 15, 2019 |
| Contact name |
Pejman Mohammadi |
| Organization name |
New York Genome Center
|
| Street address |
101 Av. of the Americas
|
| City |
New York |
| ZIP/Postal code |
10013 |
| Country |
USA |
| |
|
| Platform ID |
GPL11154 |
| Series (1) |
| GSE70851 |
HIV-1 RNAs are not part of the Argonaute 2 associated RNA interference pathway in macrophages |
|
| Relations |
| BioSample |
SAMN03856994 |
| SRA |
SRX1093016 |
