Similar protocols

Pipeline publication

[…] with SeqPrep software (https://github.com/jstjohn/SeqPrep). Thereafter, low-quality bases (Phred score < 20) were trimmed, and reads shorter than 50 bp were discarded with Sickle software (https://github.com/najoshi/sickle). rRNA reads were discarded after alignment to SILVA SSU (16S/18S) and SILVA LSU (23S/28S) databases with SortMeRNA software (http://bioinfo.lifl.fr/RNA/sortmerna/). The resulting high-quality reads were then used in the subsequent assembly. The mega-transcriptome was de novo assembled with Trinity software (http://trinityrnaseq.github.io/; version trinityrnaseq-r2013-02-25) using default parameters as previously described (Grabherr et al., ). ORFs were predicted using TransGeneScan software (http://sourceforge.net/projects/transgenescan/). Non-redundant gene catalogs were constructed with an identity of 95% and a coverage of 90% using CD-HIT software (http://www.bioinformatics.org/cd-hit/). To evaluate the expression level of each transcript, the FPKM (fragments per kilobase of exon per million fragments mapped) value of each transcript was obtained with RSEM (RNASeq by expectation maximization) software (http://deweylab.biostat.wisc.edu/rsem/). Differentially expressed genes were identified using edgeR (empirical analysis of digital gene expression data in R) software (http://www.bioconductor.org/packages/release/bioc/html/edgeR.html; Reiner et al., ; Robinson and Smyth, ). For this analysis, the filtering threshold was set to an FDR (false discovery rate) < 0.5 and a |log2FC| > 1., All genes were characterized with BALSTX comparisons against the integrated NCBI NR database with an expectation value of 1e-5 (BLAST Version 2.2.28+, http://blast.ncbi.nlm.nih.gov/Blast.cgi). Species information was obtained from the respective taxonomy annotation NR database. The species abundance in each specimen was determined by calculating the FPKM value of each transcript in the respective sp […]

Pipeline specifications

Software tools TransGeneScan, CD-HIT, RSEM, edgeR