Similar protocols

Pipeline publication

[…] Nr: GSE25305). Differential expression of probe sets was assessed by fitting a linear model, including the effects of feeding (2 levels: Control and TTA) and the effects of sampling point (2 levels: 8.weeks and 17.weeks) and their interaction. The specific comparisons: TTA vs. Control at sampling point 8.weeks and TTA vs.Control at sampling point 17.weeks were made by extracting the appropriate contrasts from the linear model. For each contrast moderated t-statistics were calculated using an empirical Bayes method []. Probes without annotation were removed from the dataset before controlling the false discovery rate [] simultaneously across probe sets and contrasts (method: “ global” in the limma function “decideTests”). Probe sets with a q-value ≤ 0.05 and a log2FC ≥ 0.5 were declared DE for the corresponding contrast. A comprehensive list of all DE probes for each contrast can be found in Additional file : Table S3., Probe annotation and GOs were retrieved using the top Blast function implemented in Blast2GO []. Full length probe sequences were blasted against protein sequences from the Swissprot database in a BlastX search. The E-value cut off was set to 10−6. Hypergeometric testing for over-representation of GO terms from the category biological process [] among the genes DE for the contrast TTA vs.Control at the 17.weeks sampling point was conducted using the GOstats package []. Before testing, probes matching to the same gene were collapsed to the probe showing the largest variance., Correspondence analysis was conducted using the R package “made4” []. Probe sets with missing values were removed from the dataset prior to correspondence analysis., Single strand cDNA was synthesized from 500ng of total RNA using oligo dT primers and the Taq Man reverse transcription Kit (Applied Biosystems, CA, USA). qRT-PCR was performed on a Light-Cycler 480 (Roche, Switzerland). For the PCR reaction, 2x SYBR green I master Mix (Roche), 0.41nM of each primer and the cDNA template were mixed in a total reaction volume of 10μl. Primer sequences are listed in Table […]

Pipeline specifications

Software tools limma, Blast2GO, BLASTX, GOstats
Organisms Salmo salar
Diseases Hypertrophy