GSM1574332: P1_Lin28_tg_mut_n2; Mus musculus; RNA-Seq
Sciatic nerves were freed from perineurial fibroblast layer and homogenized with a chilled pestle. Total RNA was subsequently extracted using Qiazol according to manufacturers description. The TruSeq RNA Sample Prep Kit v2 (Illumina, Inc.) was used in the succeeding steps. Briefly, total RNA samples (100-1000 ng) were depleted of ribosomal RNA using Ribo Zero Gold (Epicentre¨) and then reverse-transcribed into double-stranded cDNA. The cDNA samples were fragmented, end-repaired and polyadenylated before ligation of TruSeq adapters containing the index for multiplexing Fragments containing TruSeq adapters on both ends were selectively enriched with PCR. The quality and quantity of the enriched libraries were validated using Qubit¨ (1.0) Fluorometer and the Caliper GX LabChip¨ GX (Caliper Life Sciences, Inc.). The product is a smear with an average fragment size of approximately 260 bp. The libraries were normalized to 10nM in Tris-HCl 10 mM (pH 8.5) with 0.1% Tween 20. The TruSeq PE Cluster Kit v3-cBot-HS or TruSeq SR Cluster Kit v3-cBot-HS (Illumina, Inc.) was used for cluster generation using 10 pM of pooled normalized libraries on the cBOT. Sequencing were performed on the Illumina HiSeq 2000 paired end at 2 X101 bp or single end 100 bp using the TruSeq SBS Kit v3-HS (Illumina, Inc.)
GEO
GSM1574332