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Characterization of the Conus bullatus genome and its venom-duct transcriptome.

Hu H, Bandyopadhyay PK, Olivera BM, Yandell M - BMC Genomics (2011)

Bottom Line: Our results provide the first global view of venom-duct transcription in any cone snail.A notable feature of Conus bullatus venoms is the breadth of A-superfamily peptides expressed in the venom duct, which are unprecedented in their structural diversity.We also find SNP rates within conopeptides are higher compared to the remainder of C. bullatus transcriptome, consistent with the hypothesis that conopeptides are under diversifying selection.

View Article: PubMed Central - HTML - PubMed

Affiliation: Eccles institute of Human Genetics, University of Utah, and School of Medicine, Salt Lake City, UT 84112, USA.

ABSTRACT

Background: The venomous marine gastropods, cone snails (genus Conus), inject prey with a lethal cocktail of conopeptides, small cysteine-rich peptides, each with a high affinity for its molecular target, generally an ion channel, receptor or transporter. Over the last decade, conopeptides have proven indispensable reagents for the study of vertebrate neurotransmission. Conus bullatus belongs to a clade of Conus species called Textilia, whose pharmacology is still poorly characterized. Thus the genomics analyses presented here provide the first step toward a better understanding the enigmatic Textilia clade.

Results: We have carried out a sequencing survey of the Conus bullatus genome and venom-duct transcriptome. We find that conopeptides are highly expressed within the venom-duct, and describe an in silico pipeline for their discovery and characterization using RNA-seq data. We have also carried out low-coverage shotgun sequencing of the genome, and have used these data to determine its size, genome-wide base composition, simple repeat, and mobile element densities.

Conclusions: Our results provide the first global view of venom-duct transcription in any cone snail. A notable feature of Conus bullatus venoms is the breadth of A-superfamily peptides expressed in the venom duct, which are unprecedented in their structural diversity. We also find SNP rates within conopeptides are higher compared to the remainder of C. bullatus transcriptome, consistent with the hypothesis that conopeptides are under diversifying selection.

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C. bullatus genome size estimated using Illumina reads. Blue-line: C. bullatus; Red-line: D. melanogaster. Green line: C. elegans. x-axis: depth of coverage of transciptome contigs by aligned genomic reads. y-axis: frequency. In all cases the best estimate for genome size is the product of the total length of genomic reads and the mode of the frequency distribution.
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Figure 4: C. bullatus genome size estimated using Illumina reads. Blue-line: C. bullatus; Red-line: D. melanogaster. Green line: C. elegans. x-axis: depth of coverage of transciptome contigs by aligned genomic reads. y-axis: frequency. In all cases the best estimate for genome size is the product of the total length of genomic reads and the mode of the frequency distribution.

Mentions: We have developed a novel method for determining genome size, using 2nd generation genomic and RNA-Seq reads (see Methods). For proof of principle, we first estimated the genome size of D. melanogaster. To do so, we simulated 4,342,253 59bp genomic reads for the fly-genome, and blasted the annotated fly transcriptome against the simulated reads (red line in Figure 4). The depth of coverage peak is at 1.50 (Figure 4). Thus, the estimated genome size for D. melanogaster is 4,342,253*59/1.50 = 170.8 MB. Compared to the current size of fly genome (166.6 MB), the error is 2.5%. We also estimated the genome size of C. elegans. This time we randomly sheared the annotated transcriptome of C. elegans into short contigs with the same N50 as our C. bullatus transcriptome assembly, and randomly selected a 57mb subset of these contigs. We did this to simulate the fragmented nature of our de novo transcriptome assembly. We also simulated 2,630,408 genomic C. elegans reads, and blasted them to the subset of simulated C. elegans transcriptome. As shown in Figure 4 (green line), the peak depth of coverage for the transcriptome is 1.45×. We repeated this experiment three times; there was no variance in this value. This gives us an estimate of genome size of 107.0MB, which is 6.7% higher than estimated genome size (100.3MB), again a good fit to the published genome size. For Conus bullatus, the estimated coverage depth is 1.70× from 4.36GB of sequence reads, thus the best estimate for the size of the Conus bullatus genome is 2.56 GB.


Characterization of the Conus bullatus genome and its venom-duct transcriptome.

Hu H, Bandyopadhyay PK, Olivera BM, Yandell M - BMC Genomics (2011)

C. bullatus genome size estimated using Illumina reads. Blue-line: C. bullatus; Red-line: D. melanogaster. Green line: C. elegans. x-axis: depth of coverage of transciptome contigs by aligned genomic reads. y-axis: frequency. In all cases the best estimate for genome size is the product of the total length of genomic reads and the mode of the frequency distribution.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3040727&req=5

Figure 4: C. bullatus genome size estimated using Illumina reads. Blue-line: C. bullatus; Red-line: D. melanogaster. Green line: C. elegans. x-axis: depth of coverage of transciptome contigs by aligned genomic reads. y-axis: frequency. In all cases the best estimate for genome size is the product of the total length of genomic reads and the mode of the frequency distribution.
Mentions: We have developed a novel method for determining genome size, using 2nd generation genomic and RNA-Seq reads (see Methods). For proof of principle, we first estimated the genome size of D. melanogaster. To do so, we simulated 4,342,253 59bp genomic reads for the fly-genome, and blasted the annotated fly transcriptome against the simulated reads (red line in Figure 4). The depth of coverage peak is at 1.50 (Figure 4). Thus, the estimated genome size for D. melanogaster is 4,342,253*59/1.50 = 170.8 MB. Compared to the current size of fly genome (166.6 MB), the error is 2.5%. We also estimated the genome size of C. elegans. This time we randomly sheared the annotated transcriptome of C. elegans into short contigs with the same N50 as our C. bullatus transcriptome assembly, and randomly selected a 57mb subset of these contigs. We did this to simulate the fragmented nature of our de novo transcriptome assembly. We also simulated 2,630,408 genomic C. elegans reads, and blasted them to the subset of simulated C. elegans transcriptome. As shown in Figure 4 (green line), the peak depth of coverage for the transcriptome is 1.45×. We repeated this experiment three times; there was no variance in this value. This gives us an estimate of genome size of 107.0MB, which is 6.7% higher than estimated genome size (100.3MB), again a good fit to the published genome size. For Conus bullatus, the estimated coverage depth is 1.70× from 4.36GB of sequence reads, thus the best estimate for the size of the Conus bullatus genome is 2.56 GB.

Bottom Line: Our results provide the first global view of venom-duct transcription in any cone snail.A notable feature of Conus bullatus venoms is the breadth of A-superfamily peptides expressed in the venom duct, which are unprecedented in their structural diversity.We also find SNP rates within conopeptides are higher compared to the remainder of C. bullatus transcriptome, consistent with the hypothesis that conopeptides are under diversifying selection.

View Article: PubMed Central - HTML - PubMed

Affiliation: Eccles institute of Human Genetics, University of Utah, and School of Medicine, Salt Lake City, UT 84112, USA.

ABSTRACT

Background: The venomous marine gastropods, cone snails (genus Conus), inject prey with a lethal cocktail of conopeptides, small cysteine-rich peptides, each with a high affinity for its molecular target, generally an ion channel, receptor or transporter. Over the last decade, conopeptides have proven indispensable reagents for the study of vertebrate neurotransmission. Conus bullatus belongs to a clade of Conus species called Textilia, whose pharmacology is still poorly characterized. Thus the genomics analyses presented here provide the first step toward a better understanding the enigmatic Textilia clade.

Results: We have carried out a sequencing survey of the Conus bullatus genome and venom-duct transcriptome. We find that conopeptides are highly expressed within the venom-duct, and describe an in silico pipeline for their discovery and characterization using RNA-seq data. We have also carried out low-coverage shotgun sequencing of the genome, and have used these data to determine its size, genome-wide base composition, simple repeat, and mobile element densities.

Conclusions: Our results provide the first global view of venom-duct transcription in any cone snail. A notable feature of Conus bullatus venoms is the breadth of A-superfamily peptides expressed in the venom duct, which are unprecedented in their structural diversity. We also find SNP rates within conopeptides are higher compared to the remainder of C. bullatus transcriptome, consistent with the hypothesis that conopeptides are under diversifying selection.

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