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A de novo expression profiling of Anopheles funestus, malaria vector in Africa, using 454 pyrosequencing.

Gregory R, Darby AC, Irving H, Coulibaly MB, Hughes M, Koekemoer LL, Coetzee M, Ranson H, Hemingway J, Hall N, Wondji CS - PLoS ONE (2011)

Bottom Line: In total 20.8% of all reads were novel when compared to reference databases.Gene conservation analysis confirmed the close phylogenetic relationship between An. funestus and An. gambiae.This study represents a significant advance for the genetics and genomics of An. funestus since it provides an extensive set of both Expressed Sequence Tags (ESTs) and SNPs which can be readily adopted for the design of new genomic tools such as microarray or SNP platforms.

View Article: PubMed Central - PubMed

Affiliation: University of Liverpool, School of Biological Sciences, Cornwall House, United Kingdom.

ABSTRACT

Background: Anopheles funestus is one of the major malaria vectors in Africa and yet there are few genomic tools available for this species compared to An. gambiae. To start to close this knowledge gap, we sequenced the An. funestus transcriptome using cDNA libraries developed from a pyrethroid resistant laboratory strain and a pyrethroid susceptible field strain from Mali.

Results: Using a pool of life stages (pupae, larvae, adults: females and males) for each strain, 454 sequencing generated 375,619 reads (average length of 182 bp). De novo assembly generated 18,103 contigs with average length of 253 bp. The average depth of coverage of these contigs was 8.3. In total 20.8% of all reads were novel when compared to reference databases. The sequencing of the field strain generated 204,758 reads compared to 170,861 from the insecticide resistant laboratory strain. The contigs most differentially represented in the resistant strain belong to the P450 gene family and cuticular genes which correlates with previous studies implicating both of these gene families in pyrethroid resistance. qPCR carried out on six contigs indicates that these ESTs could be suitable for gene expression studies such as microarray. 31,000 sites were estimated to contain Single Nucleotide Polymorphisms (SNPs) and analysis of SNPs from 20 contigs suggested that most of these SNPs are likely to be true SNPs. Gene conservation analysis confirmed the close phylogenetic relationship between An. funestus and An. gambiae.

Conclusion: This study represents a significant advance for the genetics and genomics of An. funestus since it provides an extensive set of both Expressed Sequence Tags (ESTs) and SNPs which can be readily adopted for the design of new genomic tools such as microarray or SNP platforms.

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Related in: MedlinePlus

Number of hits in reference species in the nr database using BLASTX (E = 1) of the unassembled reads.Corresponding species have been labelled.
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pone-0017418-g002: Number of hits in reference species in the nr database using BLASTX (E = 1) of the unassembled reads.Corresponding species have been labelled.

Mentions: For most of the reads, the best hits matched a mosquito species in the nr database, most frequently An. gambiae. Figure 2 shows the number of best hits for each read matching any of the species represented in the nr database. Key species have been highlighted in Figure 2 with the wide variation in the number of genes in reference species to be noted. Although at a lower frequency, An. funestus reads also matched genes from distant species such as human or mouse probably for conserved genes. Analysis of the contig length distribution resulting from the assembly of reads shows a large peak at 240 bp (Figure 3a). The longest contig is 2,226 bp long and made use of 224 reads, coverage varied from 1 to 53 supporting reads.


A de novo expression profiling of Anopheles funestus, malaria vector in Africa, using 454 pyrosequencing.

Gregory R, Darby AC, Irving H, Coulibaly MB, Hughes M, Koekemoer LL, Coetzee M, Ranson H, Hemingway J, Hall N, Wondji CS - PLoS ONE (2011)

Number of hits in reference species in the nr database using BLASTX (E = 1) of the unassembled reads.Corresponding species have been labelled.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3045460&req=5

pone-0017418-g002: Number of hits in reference species in the nr database using BLASTX (E = 1) of the unassembled reads.Corresponding species have been labelled.
Mentions: For most of the reads, the best hits matched a mosquito species in the nr database, most frequently An. gambiae. Figure 2 shows the number of best hits for each read matching any of the species represented in the nr database. Key species have been highlighted in Figure 2 with the wide variation in the number of genes in reference species to be noted. Although at a lower frequency, An. funestus reads also matched genes from distant species such as human or mouse probably for conserved genes. Analysis of the contig length distribution resulting from the assembly of reads shows a large peak at 240 bp (Figure 3a). The longest contig is 2,226 bp long and made use of 224 reads, coverage varied from 1 to 53 supporting reads.

Bottom Line: In total 20.8% of all reads were novel when compared to reference databases.Gene conservation analysis confirmed the close phylogenetic relationship between An. funestus and An. gambiae.This study represents a significant advance for the genetics and genomics of An. funestus since it provides an extensive set of both Expressed Sequence Tags (ESTs) and SNPs which can be readily adopted for the design of new genomic tools such as microarray or SNP platforms.

View Article: PubMed Central - PubMed

Affiliation: University of Liverpool, School of Biological Sciences, Cornwall House, United Kingdom.

ABSTRACT

Background: Anopheles funestus is one of the major malaria vectors in Africa and yet there are few genomic tools available for this species compared to An. gambiae. To start to close this knowledge gap, we sequenced the An. funestus transcriptome using cDNA libraries developed from a pyrethroid resistant laboratory strain and a pyrethroid susceptible field strain from Mali.

Results: Using a pool of life stages (pupae, larvae, adults: females and males) for each strain, 454 sequencing generated 375,619 reads (average length of 182 bp). De novo assembly generated 18,103 contigs with average length of 253 bp. The average depth of coverage of these contigs was 8.3. In total 20.8% of all reads were novel when compared to reference databases. The sequencing of the field strain generated 204,758 reads compared to 170,861 from the insecticide resistant laboratory strain. The contigs most differentially represented in the resistant strain belong to the P450 gene family and cuticular genes which correlates with previous studies implicating both of these gene families in pyrethroid resistance. qPCR carried out on six contigs indicates that these ESTs could be suitable for gene expression studies such as microarray. 31,000 sites were estimated to contain Single Nucleotide Polymorphisms (SNPs) and analysis of SNPs from 20 contigs suggested that most of these SNPs are likely to be true SNPs. Gene conservation analysis confirmed the close phylogenetic relationship between An. funestus and An. gambiae.

Conclusion: This study represents a significant advance for the genetics and genomics of An. funestus since it provides an extensive set of both Expressed Sequence Tags (ESTs) and SNPs which can be readily adopted for the design of new genomic tools such as microarray or SNP platforms.

Show MeSH
Related in: MedlinePlus