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Raw reads were first processed to remove adapter and low-quality sequences using Trimmomatic31.
An additional 0.7X Ampure bead clean-up was performed to remove adapter dimer before 11 PCR cycles of library amplification.
Reads were trimmed to remove adapter sequences.
RADseq data were trimmed to remove adapter sequence and error-prone regions.
Fastx clipper was used to remove adapter sequences matching the following ten-bases sequences: GAAGAGCGGT.
Small RNA reads were trimmed to remove adapter sequences and low quality reads.
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Prior to assembly, raw reads were filtered with Prinseq v0.20.4 (ref. 17), in order to remove duplicated sequences and the reads showing a quality score mean below 30 (Q<30) and with Cutadapt v1.9, to remove adapters (Table 1).
Deconvolution and filtering of sequence reads to remove adapter-dimer sequences yielded a mean of 11.3 million reads (range: 2.7 million to 18.0 million reads) per individual RNA-seq library.
Size selection of the cDNA constructs was performed using a 3%% gel cassette on the Pippin Prep (Sage Sciences, Beverly, MA) to reduce the library to mature miRNAs and other regulatory RNAs in the 20 30 bp size range and to remove adapter-adapter products.
BS-Seeker2 and BSMAP have built-in functions to remove adapters, while Bismark does not.
Illumina reads were trimmed to remove adapters and low quality bases using the software Sickle v1.2 (github.com/najoshi/sickle).
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