De novo assembly of maritime pine transcriptome: implications for forest breeding and biotechnology

Abstract

SummaryMaritime pine (Pinus pinaster Ait.) is a widely distributed conifer species in Southwestern Europe and one of the most advanced models for conifer research. In the current work, comprehensive characterization of the maritime pine transcriptome was performed using a combination of two different next-generation sequencing platforms, 454 and Illumina. De novo assembly of the transcriptome provided a catalogue of 26 020 unique transcripts in maritime pine trees and a collection of 9641 full-length cDNAs. Quality of the transcriptome assembly was validated by RT-PCR amplification of selected transcripts for structural and regulatory genes. Transcription factors and enzyme-encoding transcripts were annotated. Furthermore, the available sequencing data permitted the identification of polymorphisms and the establishment of robust single nucleotide polymorphism (SNP) and simple-sequence repeat (SSR) databases for genotyping applications and integration of translational genomics in maritime pine breeding programmes. All our data are freely available at SustainpineDB, the P. pinaster expressional database. Results reported here on the maritime pine transcriptome represent a valuable resource for future basic and applied studies on this ecological and economically important pine species.

Ecological and association genetics in two Mediterranean pine species

“…Thus, 285 SNPs were discarded and the final list comprised 4712 SNPs. Then, flanking sequences of each retained SNP were aligned against the most up-to-date maritime pine UniGene (PineV3, Canales et al 2014: http:// www.scbi.uma.es/sustainpinedb/home_page) to identify its corresponding contig and position within that contig. Alignments were performed by carrying out BLASTN searches in the BLASTALL version 2.2.26 suite (Evalue = 10 À5 ).…”

“…An additional set of in silico SNPs was then obtained from 281 candidate genes with functions relating to abiotic and biotic (plant defence) stress responses in forest trees selected from the following sources: (i) 66 genes from published and unpublished maritime pine sequences available from GenBank, (ii) 149 genes from sequencing studies in other conifers (Wachowiak et al 2009;Kujala & Savolainen 2012), (iii) 53 transcripts displaying differential expression in the presence and absence of pine wood nematode infection ), (iv) 10 genes associated with adventitious shoot induction and plant development in pines (Alonso et al 2007; Ord as et al unpublished for knox genes) and (v) 3 genes potentially involved in cavitation resistance in beech (Lalag€ ue et al 2014). These sequences were blasted (BLASTN, E-value = 10 À5 ) against PineV3 UniGene to retrieve the best matching contig and the SNP flanking sequences from the catalogue of 55 607 available SNPs obtained by Canales et al (2014; Fig. 2).…”

“…Such discoveries should also lead to changes in silviculture practices to take into account the evolutionary processes inferred from neutral and selected markers (Lef evre et al 2014). Major efforts have been devoted to the sequencing and assembly of the maritime pine transcriptome (Canales et al 2014), for studies of the molecular basis of the phenotypic response to biotic and abiotic constraints . Moreover, since the pioneering work of Lepoittevin et al (2010), describing the design of the first multiplex single nucleotide polymorphism (SNP) genotyping assay in maritime pine, medium-scale SNP arrays have been developed (Tables S1 and S2, Supporting information).…”

Forest tree genomics: 10 achievements from the past 10 years and future prospects

“…Conifers are the primary source for wood production worldwide (Canales et al 2014). By unraveling the complexity of the regulated gene network involved in conifer embryo development, it should be possible to develop genomic and epigenomic tools for the early selection of improved varieties.…”

Nitrogen Economy and Nitrogen Environmental Interactions in Conifers

“…However, only limited knowledge is currently available on the expression of key genes involved in arginine biosynthesis and the transcriptional regulation of this process in conifers. In last few years, the assembly of pine and spruce transcriptomes [36,37] and genomes have been reported [38,39] and these new resources will facilitate the identification and functional characterization of genes involved in arginine metabolism. development [36].…”