title: GSM8497732: 012323-OJO001; Homo sapiens; RNA-Seq
library_construction_protocol: The Allprep DNA/RNA Mini Kit (QIAGEN) was used for RNA isolation and the manufacturer's protocol was followed using the Qiacube. The extraction process was performed on LC (CD19+), non-leukemic cells (CD19-), and the whole cell population (cells without separation). RNA quantification was performed using Qubit™ RNA High Sensitivity and Nanodrop according to the manufacturer's protocols. RNA obtained from the CD19 negative fraction of the samples was used to prepare the libraries. Libraries were generated using Illumina Stranded Total RNA prep with Ribo-Zero Plus library preparation kit according to manufacturer's instructions. Briefly, ribosomal RNA was first depleted from total RNA (100 ng). Following purification, the RNA was fragmented, and first- and second-strand cDNA synthesized.  Libraries were created from the cDNA by first adenylating the 3′ end and ligating anchors to the ends. An amplification step of 13 cycles of PCR was then performed to add unique indexes to the ligated products. Resulting libraries were quantified using the Qubit dsDNA High Sensitivity Assay Kit and the size and purity assessed with the Agilent 2100 bioanalyzer. Sequencing was performed on Illumina's NextSeq 2000 instrument using a NextSeq 2000 P2 200 cycles kit with pair-end 76 bp reads. FASTQ files were uploaded to Partek Flow for analysis, as follows: contaminants (rDNA, tRNA, mtDNA) were removed with Bowtie 2 (v2.2.5) and the reads aligned to hg38 with STAR v2.7.8a and quantified with RefSeq Transcripts 93.