Annotation and gene ontology (GO)
analysis
To obtain a functional annotation for the transcripts, we used blastp
(Camacho et al., 2009) to search for the best match among Arabidopsis
representative set of proteins (Berardini et al., 2015) downloaded from
TAIR FTP server
(ftp://ftp.arabidopsis.org/home/tair/Proteins/TAIR10_protein_lists/).
The retrieved best hit was assumed to be an Arabidopsis ortholog and the
functional annotation and GO category assignment of the ortholog was
transferred to the P. lanceolata query transcript. The
Arabidopsis GO annotations were downloaded from the TAIR FTP server
(ftp://ftp.arabidopsis.org/home/tair/Ontologies/Gene_Ontology/). Gene
ontology enrichment analysis was carried out using piano software
(Varemo, Nielsen, & Nookaew, 2013) and the log2 fold
changes and false discovery rate (FDR) adjusted P-values of
differentially expressed genes were imported from DESeq2 results.
The GO hierarchy enrichment tests were carried out using the GOATools
toolkit (Klopfenstein et al., 2018). To focus on signaling responses, we
selected the responses and signaling sub-branches of the Biological
process category. The GO enrichment results were plotted using R with
fold change values obtained from the piano package. The GO enrichment
analyses of RDA loadings was carried out with piano software (Varemo et
al., 2013) using RDA loadings as gene level statistics and GO categories
as gene set collection and mean statistics. Again, the results were
plotted in R.