title: A 454 sequencing approach to dipteran mitochondrial genome research.
abstract: The availability of complete mitochondrial genome (mtgenome) data for Diptera, one of the largest metazoan orders, in public databases is limited. The advent of high throughput sequencing technology provides the potential to generate mtgenomes for many species affordably and quickly. However, these technologies need to be validated for dipterans as the members of this clade play important economic and research roles. Illumina and 454 sequencing platforms are widely used in genomic research involving non-model organisms. The Illumina platform has already been utilized for generating mitochondrial genomes without using conventional long range PCR for insects whereas the power of 454 sequencing for generating mitochondrial genome drafts without PCR has not yet been validated for insects. Thus, this study examines the utility of 454 sequencing approach for dipteran mtgenomic research. We generated complete or nearly complete mitochondrial genomes for Cochliomyia hominivorax, Haematobia irritans, Phormia regina and Sarcophaga crassipalpis using a 454 sequencing approach. Comparisons between newly obtained and existing assemblies for C. hominivorax and H. irritans revealed no major discrepancies and verified the utility of 454 sequencing for dipteran mitochondrial genomes. We also report the complete mitochondrial sequences for two forensically important flies, P. regina and S. crassipalpis, which could be used to provide useful information to legal personnel. Comparative analyses revealed that dipterans follow similar codon usage and nucleotide biases that could be due to mutational and selection pressures. This study illustrates the utility of 454 sequencing to obtain complete mitochondrial genomes for dipterans without the aid of conventional molecular techniques such as PCR and cloning and validates this method of mtgenome sequencing in arthropods.
authors.affiliation: The Institute for Genomics, Biocomputing and Biotechnology, Mississippi State University, Mississippi State, MS 39762, USA; Department of Biochemistry, Molecular Biology, Entomology and Plant Pathology, Mississippi State University, Mississippi State, MS 39762, USA.
authors.forename: Meganathan P
authors.initials: MP
authors.lastname: Ramakodi
authors.affiliation: Department of Biology, West Virginia University, Morgantown, WV 26506, USA.
authors.forename: Baneshwar
authors.initials: B
authors.lastname: Singh
authors.affiliation: Department of Biology, West Virginia University, Morgantown, WV 26506, USA.
authors.forename: Jeffrey D
authors.initials: JD
authors.lastname: Wells
authors.affiliation: USDA-ARS, Knipling Bushland US Livestock Insect Research Laboratory, Kerrville, TX 78028, USA.
authors.forename: Felix
authors.initials: F
authors.lastname: Guerrero
authors.affiliation: The Institute for Genomics, Biocomputing and Biotechnology, Mississippi State University, Mississippi State, MS 39762, USA; Department of Biochemistry, Molecular Biology, Entomology and Plant Pathology, Mississippi State University, Mississippi State, MS 39762, USA. Electronic address: david.4.ray@gmail.com.
authors.forename: David A
authors.initials: DA
authors.lastname: Ray
mesh_terms: D000818:Animals; D004175:Diptera / Q000145:classification / Q000166:cytology / Q000235:genetics*; D019143:Evolution, Molecular; D049750:Genome, Insect; D054629:Genome, Mitochondrial; D059014:High-Throughput Nucleotide Sequencing / Q000379:methods*; D008928:Mitochondria / Q000235:genetics; D008969:Molecular Sequence Data; D010802:Phylogeny; D017422:Sequence Analysis, DNA / Q000379:methods*
keywords: 454 sequencing; Diptera; Evolution; Forensic entomology; Mitochondrial genomes; Phylogenetic analyses