An annotated transcriptome of highly inbred Thuja plicata (Cupressaceae) and its utility for gene discovery of terpenoid biosynthesis and conifer defense

Abstract

Western redcedar (Thuja plicata; Cupressaceae; WRC) is an ecologically and economically important conifer species of the Pacific Northwest. Regeneration of WRC forests is affected by ungulate browsing, which removes current growth and hampers development of young trees. Monoterpenes make WRC foliage less palatable and can deter browsing. Genomic resources are required to advance knowledge of terpene accumulation and breeding of WRC for herbivore resistance. Unlike most conifers, WRC readily selfs to produce genotypes of reduced heterozygosity. We used seedlings of eight different fifth-generation selfed lines for monoterpene analysis and transcriptome sequencing. Trinity, Velvet/Oases, TransABySS, and SOAPdenovoTrans were used to generate independent transcriptome assemblies for each line. Sequence redundancy was reduced using the EvidentialGene pipeline. The best assembly, as determined by metrics of completeness, contiguity, and accuracy, was used to produce a WRC reference gene set of 28,279 sequences, of which 77% were annotated with significant BLASTp hits and 89% with significant InterProScan hits. An orthology-based approach was used to annotate gene families. Manually curated annotation identified 33 putative full-length terpene synthases (TPS). A maximum likelihood phylogeny revealed that WRC TPS cluster apart from those of Pinaceae within the gymnosperm TPS-d clade. Use of selfed lines enabled the development and annotation of a reduced-redundancy gene set for a gymnosperm of the Cupressaceae family. This gene set serves as a foundation for future functional characterization of WRC TPS and other defense genes and as a resource for the annotation of protein coding sequences in the WRC genome.