GSM7330671: ILEUM_ARE_biol rep 2; Mus musculus; RNA-Seq
RNA was isolated by means of the RNeasy Micro kit following manufacturer's instructions (Qiagen). Per sample, an amount of 1µg of total RNA was used as input. Using the Illumina TruSeq® Stranded mRNA Sample Prep Kit (protocol version:# 1000000040498 v00 October 2017) poly-A containing mRNA molecules were purified from the total RNA input using poly-T oligo-attached magnetic beads. In a reverse transcription reaction using random primers, RNA was converted into first strand cDNA and subsequently converted into double-stranded cDNA in a second strand cDNA synthesis reaction using DNA PolymeraseI and RNAse H. The cDNA fragments were extended with a single 'A' base to the 3' ends of the blunt-ended cDNA fragments after which multiple indexing adapters were ligated introducing different barcodes for each sample. Finally, enrichment PCR was carried out to enrich those DNA fragments that have adapter molecules on both ends and to amplify the amount of DNA in the library. Sequence-libraries of each sample were equimolarly pooled and sequenced on Illumina NovaSeq 6000 (S1 flow cell, 100 cycles, single read, lane 1+2) at the VIB Nucleomics Core (www.nucleomics.be).