2.5. Likely Protein Identification and Annotation

We next extracted the likely proteins from the assembled transcriptome using TransDecoder. Then, the likely proteins were searched against the UniProtKB/Swiss-Prot database to identify known proteins, functional PFAM domains were identified using HMMER [26], signal peptides were predicted using SignalP [27], and transmembrane domains were predicted using TMHMM Sever v2.0 [28]. The EggNOG database v4.1 [29] was searched against to identify proteins in EuKaryotic Orthologous Groups (KOG), Clusters of Orthologous Groups (COGs), and non-supervised orthologous groups (NOGs). All the annotation for the assembled genes and likely proteins were subjected to the Trinotate v3.1.1 (http://trinotate.github.io) for combination.

Based on the gene annotation and likely protein annotation, we obtained the Unigenes, which were annotated into olfactory gene families, such as OBP, OR, IR, GR, and SNMP, using their names as key words. For the CSP transcripts, we aligned the Unigenes to all the CSP transcripts from NCBI GenBank. Hits with an e-value > 1 × 10⁻⁵ were filtered.