GSM1531974: 15-1; Mus musculus; RNA-Seq
Whole livers from 6 control and 6 HFD male fetuses were frozen in RNALater and total RNA was extracted using RNeasy (Qiagen). An mRNA library was prepared by the OHSU Massively Parallel Sequencing Shared Resource, using the Illumina TruSeq RNA Sample Prep Kit v2. Starting with total RNA, mRNA was purified using polyA selection, then chemically fragmented and converted into single-stranded cDNA using random hexamer priming. Next, the second strand was generated to create double-stranded cDNA. Blunt-end DNA fragments were then generated using fill-in reactions and exonuclease activity. An ‘A’- base was added to the blunt ends of each strand, preparing them for ligation to the coded sequencing adapters. Once the adapters were ligated, the constructs were then subject to 10 rounds of PCR. The amplified libraries were purified and underwent quantitative PCR with an ABI StepOne real-time PCR system. Libraries are diluted to an empirically determined concentration appropriate for the flow cell in use and applied using an Illumina cBot. Flow cells were sequenced on an Illumina HiSeq 2000.
GEO Accession
GSM1531974