GSM7025974: THP1-Mɸ 3; Homo sapiens; RNA-Seq
RNA was harvested using RNeasy RNA extraction kit (Qiagen). 2 ug of total RNA was used for the construction of sequencing libraries. The input for library construction was 500 ng of total RNA which was delivered in a 10 µL volume. Then total RNA samples (500 ng) were hybridized with Ribo-Zero Gold to substantially deplete cytoplasmic and mitochondrial rRNA from the samples. Stranded RNA sequencing libraries were prepared as described using the Illumina TruSeq Stranded Total RNA Library Prep Gold kit. The average insert size of libraries constructed with the TruSeq Stranded Total RNA Library Prep Kit was 200 bp. Purified libraries were qualified on an Agilent Technologies 4200 TapeStation using a D1000 ScreenTape assay (cat# 5067-5582). The molarity of adapter-modified molecules was defined by quantitative PCR using the Kapa Biosystems Kapa Library Quant Kit (Cat#KK4824, KAPA). Individual libraries were normalized to 1.30 nM in preparation for Illumina sequence analysis. Sequencing libraries (1.3 nM) were chemically denatured and applied to an Illumina NovaSeq flow cell using the NovaSeq XP chemistry workflow (Cat# 20021664, Illumina, USA). Following transfer of the flow cell to an Illumina NovaSeq instrument, a 2 x 151 cycle paired end sequence run was performed using a NovaSeq S4 reagent Kit (Cat# 200122866, Illumina, USA).