GSM9290668: P22 Peanut Knockout 3; Mus musculus; RNA-Seq
Retinas were dissected and flash frozen in Trizol reagent.RNA was harvested using chloroform and extraction performed using Zymo RNA Clean and Concentrator . P0 RNAseq library preparation was performed with 500ng to 1ug of total RNA. Ribosomal RNA was removed by an RNase-H method using RiboErase kits (Kapa Biosystems). mRNA was then fragmented in reverse transcriptase buffer and heating to 94 degrees for 8 minutes. mRNA was reverse transcribed to yield cDNA using SuperScript III RT enzyme (Life Technologies, per manufacturer's instructions) and random hexamers. A second strand reaction was performed to yield ds-cDNA. cDNA was blunt ended, had an A base added to the 3' ends, and then had Illumina sequencing adapters ligated to the ends.  P22 RNAseq library contruction was performed following manufacturer protocol of Takara SMARTer Stranded Total RNA High Input using Illumina TruSeq stranded primers.