GSM9066972: Stem-Line II; Day 17; CD16 positive; Rep 1; Homo sapiens; RNA-Seq
RNA was harvested from purified CD34+ derived neutrophils (CD16+ MACS or bulk). Therefore, cells were washed thrice with PBS and resuspended in RLT+ buffer before storage at -80C. Cell pellets were lysed in 600ul RNA Plus buffer supplemented with 1% mercapto-ethanol. Resuspended cells were homogenized with QiaShredder Columns and isolated with the Qiagen RNAeasy plus kit. RNA was eluted with 40ul of nuclease free water to obtain RNA concentrations between 10 and 100 ng/uL. NEBNext Ultra II Directional RNA Library Prep Kit for Illumina was used to process samples according to protocol. rRNA was depleted from total RNA using the Qiagen fast select kit. After fragmentation of the rRNA reduced RNA, a cDNA synthesis used for ligation with sequencing adapters. A final concentration of 0.8 nM of DNA was used for sequencing.
GEO
GSM9066972_r1