Assembly and Analysis of Differential Transcriptome Responses of <i>Hevea brasiliensis</i> on Interaction with <i>Microcyclus ulei</i>

<div><p>Natural rubber (<i>Hevea brasiliensis</i>) is a tropical tree used commercially for the production of latex, from which 40,000 products are generated. The fungus <i>Microcyclus ulei</i> infects this tree, causing South American leaf blight (SALB) disease. This disease causes developmental delays and significant crop losses, thereby decreasing the production of latex. Currently several groups are working on obtaining clones of rubber tree with durable resistance to SALB through the use of extensive molecular biology techniques. In this study, we used a secondary clone that was resistant to <i>M</i>. <i>ulei</i> isolate GCL012. This clone, FX 3864 was obtained by crossing between clones PB 86 and B 38 (<i>H</i>. <i>brasiliensis</i> x <i>H</i>. <i>brasiliensis</i>). RNA-Seq high-throughput sequencing technology was used to analyze the differential expression of the FX 3864 clone transcriptome at 0 and 48 h post infection (hpi) with the <i>M</i>. <i>ulei</i> isolate GCL012. A total of 158,134,220 reads were assembled using the <i>de novo</i> assembly strategy to generate 90,775 contigs with an N50 of 1672. Using a reference-based assembly, 76,278 contigs were generated with an N50 of 1324. We identified 86 differentially expressed genes associated with the defense response of FX 3864 to GCL012. Seven putative genes members of the AP2/ERF ethylene (ET)-dependent superfamily were found to be down-regulated. An increase in salicylic acid (SA) was associated with the up-regulation of three genes involved in cell wall synthesis and remodeling, as well as in the down-regulation of the putative gene <i>CPR5</i>. The defense response of FX 3864 against the GCL012 isolate was associated with the antagonistic SA, ET and jasmonic acid (JA) pathways. These responses are characteristic of plant resistance to biotrophic pathogens.</p></div

Distribution of GO terms from BLAST hits of the transcriptome of <i>Hevea brasiliensis</i> (reference-based assembly).

<p>Selected categories of GO terms are displayed at the top of the hierarchy for the biological process, molecular function and cellular component categories.</p

Distribution of contig lengths using Trinity <i>de novo</i> assembly for the <i>Hevea brasiliensis</i> transcriptome.

<p>All the contigs assembled are included.</p

Length distribution of contigs obtained by using the TopHat-Cufflinks assembly (for reference) for the <i>Hevea brasiliensis</i> transcriptome.

<p>All contigs assembled were included.</p

Species distribution (reference-based strategy).

<p>The results generated by sequence homology of transcripts against the NCBI nr database data.</p

Distribution of GO terms from BLAST hits with the transcriptome of <i>Hevea brasiliensis</i> (reference-based assembly).

<p>The biological process, molecular function and cellular component (ontology level 3) categories are shown.</p

Groups with common expression profiles.

<p>Each graph corresponds to a hierarchical clustering of transcripts. X-axis: Clone 1 samples at 0 and 48 hpi, clone 2 at 0 and 48 hpi. Y-axis: centered median of log2 (FPKM). The blue line corresponds to the average expression values per group. Gray lines represent individual transcripts. The number of transcripts per group is shown in the upper left part of each graph.</p

Signs and symptoms caused by <i>M</i>. <i>ulei</i> in <i>H</i>. <i>brasiliensis</i> leaflets.

<p>(A) Leaf deformation and sporulation of the GCL012 isolate 7 d post inoculation in RRIM 600. (B) Leaf showing the formation of stromatic structures of the isolate GCL012 at 24 dpi on RRIM 600. (C) A leaf of the clone FX 3864 showing the formation of chlorotic areas at 10 d post inoculation. (D) Germinating conidia of the GCL012 isolate. The germ tube can be observed (450X).</p

Summary data for sequencing and transcriptome assembly of <i>Hevea brasiliensis</i>. <i>De novo</i> and reference-based strategies.

<p>Summary data for sequencing and transcriptome assembly of <i>Hevea brasiliensis</i>. <i>De novo</i> and reference-based strategies.</p