characteristics.text: ERC000011
characteristics.characteristic: ENA-CHECKLIST
characteristics.text: 2022-04-06T00:30:45Z
characteristics.characteristic: ENA-FIRST-PUBLIC
characteristics.text: 2022-04-06T00:30:45Z
characteristics.characteristic: ENA-LAST-UPDATE
characteristics.text: SAMEA12921931
characteristics.characteristic: External Id
characteristics.text: Max Planck Institute of Immunobiology and Epigenetics (MPI-IE)
characteristics.characteristic: INSDC center name
characteristics.text: 2022-04-06T00:30:45Z
characteristics.characteristic: INSDC first public
characteristics.text: 2022-04-06T00:30:45Z
characteristics.characteristic: INSDC last update
characteristics.text: public
characteristics.characteristic: INSDC status
characteristics.text: ERS10521077
characteristics.characteristic: SRA accession
characteristics.text: E-MTAB-11427:SH_Knockdown_3
characteristics.characteristic: Submitter Id
characteristics.text: ArrayExpress
characteristics.ontologyTerms: http://www.ebi.ac.uk/efo/EFO_0009737
characteristics.characteristic: broker name
characteristics.text: long term hematopoietic stem cell
characteristics.ontologyTerms: http://purl.obolibrary.org/obo/CL_0002034
characteristics.characteristic: cell type
characteristics.text: human
characteristics.ontologyTerms: http://purl.obolibrary.org/obo/NCBITaxon_9606
characteristics.characteristic: common name
characteristics.text: Protocols: CD34+ bone marrow cells (StemCell Technology) were FACS sorted for LT-HSC (CD34+CD38-CD45RA-CD90+CD49f+) and prestimulated in culture for 24h. Cells were transduced with lentivirus of two different shGPR and shRenilla. The selection of the shRNA depended on GFP and puromycin. After 48h post-transduction, puromycin selection was performed for 48h. Positively transduced cells were marked by GFP and sorted using the FACS Aria III (Becton Dickinson) at the MPI FACS facility, Freiburg, Germany. The following sort parameters were used: 100 um nozzle with approximately 1,000 evt/s. 200 GFP+ cells were collected into ice-cold RNA lysis buffer (ARCTURUS PicoPure RNA Isolation Kit (Life Technologies, Invitrogen)) and stored at -80C until further usage. Total RNA isolation was performed from the indicated populations using ARCTURUS PicoPure RNA Isolation Kit (Life Technologies, Invitrogen) according to the manufacturers instructions. DNAse treatment was performed using RNAse-free DNAse Set (Qiagen). Total RNA was used as starting material for cDNA-libraries preparation using the Low Input RNA: cDNA Synthesis, Amplification and Library Generation (NEB #E6420). 14 PCR cycles were used for cDNA amplification and 10 cycles for PCR enrichment.
characteristics.characteristic: description
characteristics.text: adult
characteristics.characteristic: developmental stage
characteristics.text: normal
characteristics.ontologyTerms: http://purl.obolibrary.org/obo/PATO_0000461
characteristics.characteristic: disease
characteristics.text: GPRC5C knockdown
characteristics.characteristic: genotype
characteristics.text: 1
characteristics.characteristic: individual
characteristics.text: Homo sapiens
characteristics.characteristic: organism
characteristics.text: Homo sapiens
characteristics.ontologyTerms: http://purl.obolibrary.org/obo/NCBITaxon_9606
characteristics.characteristic: organism
characteristics.text: bone marrow
characteristics.characteristic: organism part
characteristics.text: Homo sapiens
characteristics.characteristic: scientific_name
characteristics.text: Frozen -80C
characteristics.characteristic: specimen with known storage state
characteristics.text: SH_Knockdown_3
characteristics.characteristic: title