Novel bioinformatic pipeline for de novo transcriptomics analysis: application in a Patagonian native tree species (Nothofagus pumilio)

MAXIMILIANO ESTRAVIS-BARCALA; VERÓNICA ARANA; PAULA MARCHELLI; KATRIN HEER; BIRGIT ZIEGENHAGEN; NICOLAS BELLORA

Rosario

Simposio; Genomica Funcional de Plantas 2017- (GFP2017); 2017

P { margin-bottom: 0.1in; direction: ltr; color: rgb(0, 0, 10); line-height: 120%; text-align: left; }P.western { font-family: "Liberation Serif",serif; font-size: 12pt; }P.cjk { font-family: "Droid Sans Fallback"; font-size: 12pt; }P.ctl { font-family: "FreeSans"; font-size: 12pt; }A:link { }Nothofaguspumilio (commonname: Lenga) is the most abundant tree species of the templatePatagonian forest, the southernmost woody ecosystem on Earth. Despiteits ecological and economical importance, physiological and molecularstudies in N.pumilioare scarce, and the genomic resources for the whole genus are verylimited (only a partial draft transcriptome for N.nervosa,a related species). Given our interest in the overall effect of hightemperatures in N.pumilio?sphysiology in a context of climate change, we performed NGS on leafpoly-A-selected RNA from plants exposed to contrasting temperatures(20° and 34°C) in a growth chamber controlled environment. Twobiological replicates from independent experiments were sequenced foreach temperature. The non-model nature of this species implied a denovo assemblyand all downstream analysis with no reference genome. In this talk wepresent some problems of great interest to the non-model plantspecies community and how we managed to overcome them with a hands-onprogramming approach. We will show an original bioinformatic pipelinethat integrates denovotranscriptome assembly and transcriptome mapping, with both standardand new tools developed in our lab aimed to solve basic problemsfaced by working without a reference genome, such as mis-assembledcontigs, sequence redundancy and strand specificity. Besides, thispipeline allows the annotation against a custom database (in ourcase, 12 broad-leaf tree transcriptomes from an open-access website,http://www.hardwoodgenomics.org/).Finally, we will show differential expression and transcriptome-basedphylogeny results using our corrected transcriptome.