GSM9724214: HT29,untreated,sample2; Homo sapiens; RNA-Seq
Total RNA was isolated using the RNeasy Mini Kit (74106, Qiagen), including an on column DNase digestion (79254, Qiagen),  according to the manufactures instructions.  Quality and quantity of the total RNA was assessed by the 2100 Bioanalyzer using “Agilent RNA 6000 Nano” (G2938-90034, Agilent Technologies). Total RNA samples having RIN>8 were subjected to library generation using the TruSeq stranded mRNA library preparation kit, according to the manufacturer's instructions (Document # 1000000040498 v00 , Illumina) with incorporation of xGen UDI-UMI adapters (Integrated DNA Technologies, Inc., Coralville). The stranded mRNA libraries were analyzed on a 2100 Bioanalyzer instrument following the manufacturer's protocol “Agilent DNA 7500 kit” (G2938-90025, Agilent Technologies), diluted to 10nM and pooled equimolar into multiplex sequencing pools for paired end sequencing on the NovaSeq 6000 Illumina sequencing instrument. Paired-end sequencing was performed using 150 cycles for Read 1, 19 cycles for Read i7, 10 cycles for Read i5 and 150 cycles for Read 2, using the NovaSeq6000 Reagent Kit v1.5 (300 cycles) (20028400, Illumina). On average 40 M paired reads per sample were generated.