Transcriptome assembly
We combined all libraries and assembled them de novo using
Trinity software (Grabherr et al., 2011), SOAPdenovo-Trans (k-mer sizes
39, 41) (Xie et al., 2014) and Oases (k-mer sizes 39, 43 and 47)
(Schulz, Zerbino, Vingron, & Birney, 2012). Next, we combined all the
resulting contigs and ran the EvidentialGene pipeline (Gilbert, 2013).
We then combined okayset and okayalt outputs of the pipeline and
clustered them using RapClust (Srivastava, Sarkar, Malik, & Patro,
2016), and merged the genes within the clusters using Lace (Davidson,
Hawkins, & Oshlack, 2017). To remove contamination, the resulting
contigs were aligned against NCBI non-redundant protein database
(Pruitt, Tatusova, & Maglott, 2007) using BLAST, and only the
transcripts that had a best hit in plant kingdom were retained. We also
removed the transcripts mapping to ribosomal genes and having ambiguous
sites (Ns). Minimum read coverage of three was used for all the
assemblies.