De novo characterization of the gametophyte transcriptome in bracken fern, Pteridium aquilinum

<p>Abstract</p> <p>Background</p> <p>Because of their phylogenetic position and unique characteristics of their biology and life cycle, ferns represent an important lineage for studying the evolution of land plants. Large and complex genomes in ferns combined with the absence of economically important species have been a barrier to the development of genomic resources. However, high throughput sequencing technologies are now being widely applied to non-model species. We leveraged the Roche 454 GS-FLX Titanium pyrosequencing platform in sequencing the gametophyte transcriptome of bracken fern (<it>Pteridium aquilinum</it>) to develop genomic resources for evolutionary studies.</p> <p>Results</p> <p>681,722 quality and adapter trimmed reads totaling 254 Mbp were assembled <it>de novo </it>into 56,256 unique sequences (i.e. unigenes) with a mean length of 547.2 bp and a total assembly size of 30.8 Mbp with an average read-depth coverage of 7.0×. We estimate that 87% of the complete transcriptome has been sequenced and that all transcripts have been tagged. 61.8% of the unigenes had blastx hits in the NCBI nr protein database, representing 22,596 unique best hits. The longest open reading frame in 52.2% of the unigenes had positive domain matches in InterProScan searches. We assigned 46.2% of the unigenes with a GO functional annotation and 16.0% with an enzyme code annotation. Enzyme codes were used to retrieve and color KEGG pathway maps. A comparative genomics approach revealed a substantial proportion of genes expressed in bracken gametophytes to be shared across the genomes of <it>Arabidopsis</it>, <it>Selaginella </it>and <it>Physcomitrella</it>, and identified a substantial number of potentially novel fern genes. By comparing the list of <it>Arabidopsis </it>genes identified by blast with a list of gametophyte-specific <it>Arabidopsis </it>genes taken from the literature, we identified a set of potentially conserved gametophyte specific genes. We screened unigenes for repetitive sequences to identify 548 potentially-amplifiable simple sequence repeat loci and 689 expressed transposable elements.</p> <p>Conclusions</p> <p>This study is the first comprehensive transcriptome analysis for a fern and represents an important scientific resource for comparative evolutionary and functional genomics studies in land plants. We demonstrate the utility of high-throughput sequencing of a normalized cDNA library for <it>de novo </it>transcriptome characterization and gene discovery in a non-model plant.</p

Volatile Emission in Bracken Fern Is Induced by Jasmonates but Not by Spodoptera littoralis or Strongylogaster multifasciata Herbivory

Jasmonate-mediated regulation of VOC emission has been extensively investigated in higher plants, however, only little is known about VOC production and its regulation in ferns. Here, we investigate whether the emission of VOCs from bracken fern Pteridium aquilinum is triggered by herbivory and if so - whether it is regulated by the octadecanoid signaling pathway. Interestingly, feeding of both generalist (Spodoptera littoralis) and specialist (Strongylogaster multifasciata) herbivores as well as application of singular and continuous mechanical wounding of fronds induced only very low levels of VOC emission. In contrast, treatment with jasmonic acid (JA) led to the emission of a blend of VOCs that was mainly comprised of terpenoids. Likewise, treatment with the JA precursor 12-oxo-phytodienoic acid (OPDA) and α-linolenic acid also induced VOC emission, albeit to a lower intesity than the JA treatment. Accumulation of endogenous JA was low in mechanically wounded fronds and these levels were unaffected by the application of oral secretions from both generalist or specialist herbivores. The emission of terpenoids upon JA treatment could be blocked with fosmidomycin and mevinolin, which are inhibitors of the MEP- and MVA pathways, respectively. These results indicate that similar to higher plants, terpenoid VOCs are produced via these pathways in bracken fern and that these pathways are JA-responsive. However, the very low amounts of terpenoids released after herbivory or mechanical damage are in stark contrast to what is known from higher plants. We speculate that S. multifasciata and S. littoralis feeding apparently did not induce the threshold levels of JA required for activating the MEP and MVA pathways and the subsequent volatile emission in bracken fern