title: RNA sequencing from planarian tissues
design: RNA sequencing from planarian tissues
library_construction_protocol: 20 animals per sample were collected 7 days after the last RNAi feeding. Animals were homogenized with a motorized pestle in Trizol reagent (Thermo Fisher Scientific) and total RNA extraction was performed following manufacturer's protocol. Purified RNA was DNase treated (Promega) and column purified (Qiagene).  For RNA sequencing, RNA integrity was assessed using Agilent Bioanalyzer 2100 with Nano Rna kit (RIN > 8). Libraries were prepared following the manufacturer protocol for transcriptome sequencing with the Ion Proton System (Thermo Fisher Scientific). mRNA-seq indexed library preparation was performed starting from 1 microg of total RNA with the Ion Total RNA-Seq Kit v2 (Thermo Fisher Scientific) according to the manufacturer's instructions. Indexed libraries were quality controlled on Agilent Bioanalyzer 2100 with High Sensitivity DNA kit, quantified with Qubit HS DNA, normalized and pooled to perform a multiplexed sequencing run.