GSM4728723: vargas7_FIB-2; Entamoeba histolytica HM-1:IMSS; RIP-Seq
RNA molecules were released by protein digestion adding SDS (0.5% to 1%) and proteinase K (1.2mg/mL) for 1h at 55°C and purified by homogenization in 3 vols of TRIzol and filtering through RNA purification Direct-zol™ RNA Miniprep Kit (Zymo Research) columns Purified RNA was fragmented for 5 min with Ambion reagent at 70°C (Ambion, #AM8740), recovered with the RNeasy MinElute Cleanup Kit (Qiagen #74204), dephosporylated with Antartic phosphatase at 37°C for 30 min (NEB #M0289S) and phosphorylated with T4 Polynucleotide Kinase (NEB #M0201S). Libraries were constructed according to manufacter's recommendations using TruSeq Small RNA Sample Prep Kit (Illumina, #RS-200-0012) and were purified with AMPure XP beads (Agencourt, #A63880). Then, sequencing was performed using HiSeq 2500 platform (Illumina) in a multiplexed 51+7 bases single read using a TruSeq SR Cluster kit v3-cBot-HS (Illumina, #GD-401-3002) and a TruSeq SBS kit v3-HS 50 cycles (Illumina, #FC-401-3002).