alias: E-MTAB-7391:MPP2c_p
title: Illumina HiSeq 2000 paired end sequencing; 3'-Seq HSC/MPP1-4
design: 3'-Seq HSC/MPP1-4
library_construction_protocol: Bone marrow (BM) was isolated from pooled femora, tibiae, ilia and vertebrae of C57BL6 mice (3 female mice per biological replicate) by gentle crushing in PBS using a mortar and pistil. HSC and MPP1-4 progenitors were identified by immunophenotype (Lineage- Sca1+ Kit+ CD150-/+ CD48-/+ CD34-/+), purified by FACS and subsequently subjected to 3'-seq. Total RNA isolation was performed using ARCTURUS PicoPure RNA isolation kit (Life Technologies, Invitrogen) according to the manufacturer's instructions. Total RNA was used for quality controls and for normalization of the starting material. Sequencing libraries were generated with 3 ng of total RNA using the QuantSeq 3' mRNA-Seq Library Prep Kit REV (Lexogen) according to the manufacturer's indications except for adaptations (version 015UG009V0221: Do not skip step 2; 15 minute incubation time step 4; Cycle Number was adjusted to 19 using the PCR Add-on Kit for Illumina (Lexogen))
identifiers.namespace: University of Heidelberg
identifiers.id: E-MTAB-7391:MPP2c_p