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Transcriptome Analysis of the Emerald Ash Borer (EAB), Agrilus planipennis: De Novo Assembly, Functional Annotation and Comparative Analysis.

Duan J, Ladd T, Doucet D, Cusson M, vanFrankenhuyzen K, Mittapalli O, Krell PJ, Quan G - PLoS ONE (2015)

Bottom Line: The EAB transcriptome assembly was compared with 13 other sequenced insect species, resulting in the prediction of 536 unigenes that are Coleoptera-specific.This study provides one of the most fundamental and comprehensive transcriptome resources available for EAB to date.Identification of the tissue- stage- or species- specific unigenes will benefit the further study of gene functions during growth and metamorphosis processes in EAB and other pest insects.

View Article: PubMed Central - PubMed

Affiliation: Great Lakes Forestry Centre, Canadian Forest Service, Natural Resources Canada, Sault Ste. Marie, Ontario, Canada; Department of Molecular and Cellular Biology, University of Guelph, Guelph, Ontario, Canada.

ABSTRACT

Background: The Emerald ash borer (EAB), Agrilus planipennis, is an invasive phloem-feeding insect pest of ash trees. Since its initial discovery near the Detroit, US- Windsor, Canada area in 2002, the spread of EAB has had strong negative economic, social and environmental impacts in both countries. Several transcriptomes from specific tissues including midgut, fat body and antenna have recently been generated. However, the relatively low sequence depth, gene coverage and completeness limited the usefulness of these EAB databases.

Methodology and principal findings: High-throughput deep RNA-Sequencing (RNA-Seq) was used to obtain 473.9 million pairs of 100 bp length paired-end reads from various life stages and tissues. These reads were assembled into 88,907 contigs using the Trinity strategy and integrated into 38,160 unigenes after redundant sequences were removed. We annotated 11,229 unigenes by searching against the public nr, Swiss-Prot and COG. The EAB transcriptome assembly was compared with 13 other sequenced insect species, resulting in the prediction of 536 unigenes that are Coleoptera-specific. Differential gene expression revealed that 290 unigenes are expressed during larval molting and 3,911 unigenes during metamorphosis from larvae to pupae, respectively (FDR< 0.01 and log2 FC>2). In addition, 1,167 differentially expressed unigenes were identified from larval and adult midguts, 435 unigenes were up-regulated in larval midgut and 732 unigenes were up-regulated in adult midgut. Most of the genes involved in RNA interference (RNAi) pathways were identified, which implies the existence of a system RNAi in EAB.

Conclusions and significance: This study provides one of the most fundamental and comprehensive transcriptome resources available for EAB to date. Identification of the tissue- stage- or species- specific unigenes will benefit the further study of gene functions during growth and metamorphosis processes in EAB and other pest insects.

No MeSH data available.


Related in: MedlinePlus

Abundance and distribution of the EAB transcripts compared to T. castaneum genome.The transcripts with ORFs longer than 300 bp were compared to the T. castaneum genome. A window of 10 kb was adopted to analyze the density distribution of the best hit region. External track shows T. castaneum gene density in both + (outside) and—(inside) strands. The middle track shows the density of the alignments of EAB transcripts to T. castaneum genome, in both + (outside) and—(inside) strands. Inner-most track shows the GC profile of T. castaneum genome.
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pone.0134824.g003: Abundance and distribution of the EAB transcripts compared to T. castaneum genome.The transcripts with ORFs longer than 300 bp were compared to the T. castaneum genome. A window of 10 kb was adopted to analyze the density distribution of the best hit region. External track shows T. castaneum gene density in both + (outside) and—(inside) strands. The middle track shows the density of the alignments of EAB transcripts to T. castaneum genome, in both + (outside) and—(inside) strands. Inner-most track shows the GC profile of T. castaneum genome.

Mentions: The completeness of the transcriptome assembly was evaluated in two ways. The CEGMA (Core Eukaryotic Genes Mapping Approach) program was used to assess the representation of core eukaryotic proteins in the assembly. CEGMA defines a representative set of 248 ultra-conserved Core Eukaryotic Genes (CEGs), which are mostly housekeeping genes [41], and therefore are expected to be expressed in EAB. Analysis of the assembled unigenes identified 229 of the 248 core proteins (92.3%) as complete (defined as covering more than 75% of the length of the core proteins by global alignment). Completeness was also evaluated by the ability of reconstructing of the unigenes with full-length proteins. All unigenes were scanned for potential open reading frames (ORFs). A total of 15,079 transcripts with ORFs longer than 300 bp were predicted. These transcripts were compared to the T. castaneum official gene set by reciprocal best-hits BLAST method. We identified 7,580 EAB transcripts with corresponding homologs in T. castaneum with a cut-off E-value of 10−6 (Fig 2). Of these, 2,143 (28.3%) EAB transcripts can be matched with 100% alignment coverage, while 5,505 (72.6%) can be matched with > 70% coverage. All predicted transcripts were further compared to the T. castaneum genome. The density distribution of the best hit region was analyzed. A high correlation (R = 0.81) of the best hit regions with T. castaneum was observed (Fig 3), which indicates that most of the predicted EAB transcripts are represented in T. castaneum. Taken together, both analyses suggest that our EAB transcriptome assembly had a broad representation.


Transcriptome Analysis of the Emerald Ash Borer (EAB), Agrilus planipennis: De Novo Assembly, Functional Annotation and Comparative Analysis.

Duan J, Ladd T, Doucet D, Cusson M, vanFrankenhuyzen K, Mittapalli O, Krell PJ, Quan G - PLoS ONE (2015)

Abundance and distribution of the EAB transcripts compared to T. castaneum genome.The transcripts with ORFs longer than 300 bp were compared to the T. castaneum genome. A window of 10 kb was adopted to analyze the density distribution of the best hit region. External track shows T. castaneum gene density in both + (outside) and—(inside) strands. The middle track shows the density of the alignments of EAB transcripts to T. castaneum genome, in both + (outside) and—(inside) strands. Inner-most track shows the GC profile of T. castaneum genome.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4526369&req=5

pone.0134824.g003: Abundance and distribution of the EAB transcripts compared to T. castaneum genome.The transcripts with ORFs longer than 300 bp were compared to the T. castaneum genome. A window of 10 kb was adopted to analyze the density distribution of the best hit region. External track shows T. castaneum gene density in both + (outside) and—(inside) strands. The middle track shows the density of the alignments of EAB transcripts to T. castaneum genome, in both + (outside) and—(inside) strands. Inner-most track shows the GC profile of T. castaneum genome.
Mentions: The completeness of the transcriptome assembly was evaluated in two ways. The CEGMA (Core Eukaryotic Genes Mapping Approach) program was used to assess the representation of core eukaryotic proteins in the assembly. CEGMA defines a representative set of 248 ultra-conserved Core Eukaryotic Genes (CEGs), which are mostly housekeeping genes [41], and therefore are expected to be expressed in EAB. Analysis of the assembled unigenes identified 229 of the 248 core proteins (92.3%) as complete (defined as covering more than 75% of the length of the core proteins by global alignment). Completeness was also evaluated by the ability of reconstructing of the unigenes with full-length proteins. All unigenes were scanned for potential open reading frames (ORFs). A total of 15,079 transcripts with ORFs longer than 300 bp were predicted. These transcripts were compared to the T. castaneum official gene set by reciprocal best-hits BLAST method. We identified 7,580 EAB transcripts with corresponding homologs in T. castaneum with a cut-off E-value of 10−6 (Fig 2). Of these, 2,143 (28.3%) EAB transcripts can be matched with 100% alignment coverage, while 5,505 (72.6%) can be matched with > 70% coverage. All predicted transcripts were further compared to the T. castaneum genome. The density distribution of the best hit region was analyzed. A high correlation (R = 0.81) of the best hit regions with T. castaneum was observed (Fig 3), which indicates that most of the predicted EAB transcripts are represented in T. castaneum. Taken together, both analyses suggest that our EAB transcriptome assembly had a broad representation.

Bottom Line: The EAB transcriptome assembly was compared with 13 other sequenced insect species, resulting in the prediction of 536 unigenes that are Coleoptera-specific.This study provides one of the most fundamental and comprehensive transcriptome resources available for EAB to date.Identification of the tissue- stage- or species- specific unigenes will benefit the further study of gene functions during growth and metamorphosis processes in EAB and other pest insects.

View Article: PubMed Central - PubMed

Affiliation: Great Lakes Forestry Centre, Canadian Forest Service, Natural Resources Canada, Sault Ste. Marie, Ontario, Canada; Department of Molecular and Cellular Biology, University of Guelph, Guelph, Ontario, Canada.

ABSTRACT

Background: The Emerald ash borer (EAB), Agrilus planipennis, is an invasive phloem-feeding insect pest of ash trees. Since its initial discovery near the Detroit, US- Windsor, Canada area in 2002, the spread of EAB has had strong negative economic, social and environmental impacts in both countries. Several transcriptomes from specific tissues including midgut, fat body and antenna have recently been generated. However, the relatively low sequence depth, gene coverage and completeness limited the usefulness of these EAB databases.

Methodology and principal findings: High-throughput deep RNA-Sequencing (RNA-Seq) was used to obtain 473.9 million pairs of 100 bp length paired-end reads from various life stages and tissues. These reads were assembled into 88,907 contigs using the Trinity strategy and integrated into 38,160 unigenes after redundant sequences were removed. We annotated 11,229 unigenes by searching against the public nr, Swiss-Prot and COG. The EAB transcriptome assembly was compared with 13 other sequenced insect species, resulting in the prediction of 536 unigenes that are Coleoptera-specific. Differential gene expression revealed that 290 unigenes are expressed during larval molting and 3,911 unigenes during metamorphosis from larvae to pupae, respectively (FDR< 0.01 and log2 FC>2). In addition, 1,167 differentially expressed unigenes were identified from larval and adult midguts, 435 unigenes were up-regulated in larval midgut and 732 unigenes were up-regulated in adult midgut. Most of the genes involved in RNA interference (RNAi) pathways were identified, which implies the existence of a system RNAi in EAB.

Conclusions and significance: This study provides one of the most fundamental and comprehensive transcriptome resources available for EAB to date. Identification of the tissue- stage- or species- specific unigenes will benefit the further study of gene functions during growth and metamorphosis processes in EAB and other pest insects.

No MeSH data available.


Related in: MedlinePlus