De novo transcriptome assembly

Raw sequence reads were pre-processed for base quality (Q20 from left and right) and adapter content using BBDuk package from the software BBMap version 37.68 [41] as well as rRNA filtered using SortMeRNA version 3.0.3 (Kopylova 2012).

De novo transcriptome assemblies of all five species were performed with Trinity version 2.6.5 [28, 42], SOAPdenovo-Trans version 1.04 [29] as well as TransLiG version 1.3 [30] using all three biological replicates each (Table ​(Table1).1). Based on the library protocols that were used to sequence the RNA-seq data, Trinity assembly was performed with default values and –SS_lib_type RF for Acer data, FR for Vaccinium and no lib type for Arabidopsis. The replicates are indicated via the –samples_file parameter. Strandness for the TransLiG assemblies were indicated with the -m parameter. SOAPdenovo-Trans doesn’t offer a strand-specific option and thus analyses were run with default parameters. The maximum read length and the estimated average insert size was indicated in the SOAPdenovo-Trans config file. The insert size was estimated for each sample using raw sequenced reads and BBMerge [41] and averaged for each library type. For the input of SOAPdenovo-Trans and TransLiG the input files of replicates were concatenated. By default, the minimum contig length for Trinity and TransLiG is 201 bp while it is 100 bp for SOAPdenovo-Trans. For a more balanced evaluation of the assembly quality and as it was not possible to change the minimal contig length in SOAPdenovo-Trans, contigs smaller than 201 bp were removed from all SOAPdenovo-Trans assemblies.