title: GSM3500603: NEMOIEC-KO Il12a-/-_5; Mus musculus; RNA-Seq
library_construction_protocol: Total RNA was extracted with Trizol Reagent (Life Technologies) and RNeasy columns (Qiagen) RNA-seq libraries were prepared using the Illumina® TruSeq stranded® RNA sample preparation Kit. Library preparation started with 1µg total RNA. After poly-A selection (using poly-T oligo-attached magnetic beads), mRNA was purified and fragmented using divalent cations under elevated temperature. The RNA fragments underwent reverse transcription using random primers followed by second strand cDNA synthesis with DNA Polymerase I and RNase H. After end repair and A-tailing, indexing adapters were ligated. The products were then purified and amplified (15 PCR cycles, 20ul template) to create the final cDNA libraries. After library validation and quantification (Agilent tapestation), equimolar amounts of library were pooled. The pool was quantified by using the Peqlab KAPA Library Quantification Kit and the Applied Biosystems 7900HT Sequence Detection System. The pool was sequenced by using an Illumina HiSeq 4000 sequencer with a paired-end (76x7x76 cycles) protocol.
attributes.tag: GEO Accession
attributes.value: GSM3500603