GSM3581941: EE rep2; Mus musculus; RNA-Seq
For whole DRG, RNA was extracted using RNeasy kit (Qiagen), according to manufacturer's guidelines. Residual DNA contamination was removed by treating the spin column with 40 units of RNase-free DNase I (Qiagen) for 15 min at room temperature prior to RNA elution. RNA concentrations and purity were verified for each sample following elution with Agilent 2100 Bioanalyzer (Agilent). RNA with RIN factor above 7.5 was used for library preparation. For Large diameter DRG neurons, laser capture microdissection was performed to select for neurons with diameter of at least 30 microns. RNA was extracted using Arcturus PicoPure RNA isolation kit (Life Technologies), following manufacturers protocol. RNA concentrations and quality were measured using Agilent 2100 Bioanalyzer (Agilent). RNA with RIN factor between 6 and 7 were used for library preparation. For whole DRG, libraries were generated using the TruSeq mRNA Sample Preparation Kit  (Illumina, San Diego CA) at Ospedale San Raffaele according to the Sample Preparation Guide protocol. Ligation and library integrity was verified using a High Sensitivity DNA chip on the Agilent 2100 Bioanalyzer (Agilent). Sample libraries ligated with unique adapter sequences were multiplexed and were sequenced using Illumina HiSeq 2500 100-cycle, pair end sequencing.  For laser captured DRGs, libraries were prepared using the RiboZero ScriptSeq V2 kit working from the low input protocol using 100 ng of RNA per sample.  Libraries were sequenced using Illumina HiSeq 2500 generating 125 bp, paired end reads