Protocols: CD34+ BM cells from Lonza were stained with CD34(APCCy7), CD38(PECy7), CD90(BV605), CD45RA(BV421) and CD49f (PECy5) and Live/Dead(Zombie). Two population were sorted the LT-HSC(CD34+CD38-CD90+CD45RA-Cd49f+) and the ST-HSCs(CD34+CD38-CD90-CD45RA-Cd49f-). Sort was performed on FACS Aria III (Becton Dickinson) at the MPI FACS facility, Freiburg, Germany. The following sort parameters were used: 100 um nozzle with approximately10,000 evt/s. Sorted 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 SMARTseq v4 Ultra Low RNA Kit (Takara) according to the manufacturers indications. Sequencing libraries were prepared from 10ng of cDNA libraries as input using NEBNext Ultra II FS DNA library prep kit (#E7805S, NEB) according to manufacturer's protocol together with unique dual indexing barcodes (NEB, #E6440). Libraries were amplified with 7 PCR-cycles before Ampure bead (Beckman-Coultier) clean-up and subsequent quality control.
ENA first public
2022-01-01
ENA last update
2022-01-01
External Id
SAMEA7702570
INSDC center alias
Max Planck Institute of Immunobiology and Epigenetics (MPI-IE)
INSDC center name
Max Planck Institute of Immunobiology and Epigenetics (MPI-IE)
INSDC first public
2022-01-01T00:12:37Z
INSDC last update
2022-01-01T00:12:37Z
INSDC status
public
Submitter Id
E-MTAB-9892:Lonza_19TL065744_ST_HSCs
age
20 to 30
broker name
ArrayExpress
cell type
ST-HSC (CD34+CD38-CD45RA+CD90-CD49f-)
common name
human
developmental stage
adult
disease
normal
individual
4
organism part
bone marrow
sample name
E-MTAB-9892:Lonza_19TL065744_ST_HSCs
sex
female
specimen with known storage state
frozen specimen (-80 degree celsius)