title: GSM4586401: RNA_p1_Baseline; Homo sapiens; RNA-Seq
library_construction_protocol: Cells were isolated by Miltenyi CD34 microbead kit according to the manufacturer's instructions. In brief, selection was performed immediately after defrosting the cryopreserved bone marrow patient samples. Samples were thawed in 37°C and DNase-I (Sigma D4513) was added to the samples to prevent cells clumping. Cells were resuspended in 2% FBS + IMDM medium. Cell pellets were resuspended in FcR blocking reagent and CD34 microbeads; incubated for 30 minutes. Positive selection of CD34+ cells were performed on MACS MS columns (Miltenyi Biotec 130-042-201). Cells were lysed with Trizol reagent (Invitrogen) and total RNA was extracted using miRNeasy Micro Kit (Qiagen) following manufacturer's protocol. Preparation of total RNA for sequencing on the Illumina NovaSeq 6000 NGS platform was carried out at the University of Miami's Hussman Institute for Human Genomics (http://hihg.med.miami.edu/cgt). Briefly, 1ng of total RNA was ribo-depleted and library construction was performed using the 'Nugen Ovation SoLo RNA-Seq (Human) (Rev. M01406 v3) with AnyDeplete Probe Standard Human/Mouse (rRNA)' according to manufacturer's protocol. Samples were barcoded to allow for multiplexing. Cluster generation and sequencing took place on the Illumina NovaSeq 6000 using the reagents provided in the Illumina NovaSeq 6000 S1 Reagent Kit (200 cycle) with >60M paired-end 100 reads per sample