Illumina HiSeq 2000 paired end sequencing; RNA-Seq of mouse beta cell line MIN6 after siRNA knockdown of lncRNA Pax6os1
RNA-Seq of mouse beta cell line MIN6 after siRNA knockdown of lncRNA Pax6os1
MIN6 cells were grown in DMEM supplemented with 15 % FBS. MIN6 cells were transfected with a pool of 3 siRNAs targeting Pax6os1 (kd) or 3 control siRNAs (con) using Lipofectamine siRNAMax. Cells were harvested 72 hours post-transfection. RNA was extracted with TRIzol (Invitrogen) using the manufacturer's protocol, with the addition of glycogen as a co-precipitate. Resuspended RNA was treated with DNAse I (Invitrogen), which was then heat inactivated. Polyadenylated RNA was purified with oligo-dT beads and the library constructed using the TruSeq Stranded kit (Illumina). Illumina Universal Paired-end adapters were used, and custom barcoding sequences incorporated during the amplification rounds (15 cycles) for multiplexing.