title: NextSeq 500 paired end sequencing; RNA-seq analysis of Chronic Lymphocytic Leukemia patients belonging to stereotyped subsets #6 and #8
design: RNA-seq analysis of Chronic Lymphocytic Leukemia patients belonging to stereotyped subsets #6 and #8
library_name: S9_p
library_construction_protocol: PBMCs from CLL patients peripheral blood were isolated using density gradient separation (tumor load of all preparations was checked by flow cytometry and always exceeded 70% for CD19+ cells). Negative selection of CD19+ B cells from CLL patients peripheral blood was performed using the RosetteSep B-cell enrichment kit (StemCell Technologies, Vancouver, BC, Canada) following the manufacturer's instructions. The purity of all preparations was checked by flow cytometry and always exceeded 95% for CD19+ cells. Total RNA was extracted using the QIAamp RNA Blood Mini Kit (QIAGEN, Hilden, Germany) according to the manufacturer's protocol. RNA concentration was estimated usingthe Qubit Fluorometer (Invitrogen, La Jolla, CA). 1μg of total RNA was used for the library construction using the NEB Next Ultra RNA library Prep Kit (New England Biolabs, Ipswich, MA) or the TruSeq RNA Library Prep Kit v2 (Illumina, San Diego, CA). Library quantification was performed with the KAPA SYBR FAST Universal qPCR kit (KAPA Biosystems, Boston, USA).
attributes.tag: Experimental Factor: stereotyped subset
attributes.value: #8