The head regeneration small RNA transcriptome of the planarian Dugesia japonica

Animals were kept at 21°C in mountain spring water and fed with veal liver once per week. Asexual planarians of the species Dugesia japonica were starved for one week prior to decapitation with a blade. Regeneration was allowed to occur for 4 different periods of time (0h, 24h, 72h, 120h). 5 regenerating animals from the same time points were pooled and immediately homogenized in Trizol(Invitrogen). Total RNA was extracted with Trizol (Invitrogen) according to the manufacturer's instructions. Washed RNA was dissolved in 20µl nuclease-free water and the concentration was determined using the NANODROP 2000C system (Thermo scientific). RNA integrity was assessed by applying the total RNA on a 1% ethidium bromide-stained agarose gel with subsequence electrophoresis at 100V and 15 minutes. Illumina's TruSeq RNA Preparation Kit was used to purify 15-40nt small RNAs from total RNA. Size selection and quality control were performed by BGI, Hong Kong. Libraries for each time point were sequenced on an Illumina HiSeq 4000 sequencer (single-end, 50bp).