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ARS Home » Southeast Area » Gainesville, Florida » Center for Medical, Agricultural and Veterinary Entomology » Imported Fire Ant and Household Insects Research » Research » Publications at this Location » Publication #233333
Title: Gene discovery using massively parallel pyrosequencing to develop ESTs for the flesh fly Sarcophaga crassipalpis

Author
item HAHN, DANIEL - DEPT OF ENTOMOL. UF
item RAGLAND, GREGORY - University Of Florida
item Shoemaker, David
item DENLINGER, DAVID - The Ohio State University

Submitted to: BMC Genomics
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 5/19/2009
Publication Date: 5/19/2009
Citation: Hahn, D.A., Ragland, G., Shoemaker, D.D., Denlinger, D.L. 2009. Gene discovery using massively parallel pyrosequencing to develop ESTs for the flesh fly Sarcophaga crassipalpis. Biomed Central (BMC) Genomics. 10:234.

Interpretive Summary: Flesh flies have long been important models in insect physiology and biochemistry, particularly with respect to the study of diapause, reproduction, and immunity. In addition, many flesh fly species can cause myasis in humans and livestock with significant wounds and are therefore of importance to medical and veterinary entomology. A scientist at the Center for Medical, Agricultural, and Veterinary Entomology, USDA-ARS, Gainesville, Florida and scientists from the University of Florida and Ohio State University describe here the results of a study using massively parallel pyrosequencing to produce a substantial EST sequence dataset for the flesh fly. From these data, 11,757 unique gene elements (E<0.0001) representing approximately 9,000 independent transcripts were identified, many of which share high sequence similarity to genes involved in diapause. These EST data provide the foundation for a proliferation of functional work on flesh flies such as the development of microarrays for large scale gene expression studies as well as serving as a foundation for identifying previously unrecognizable proteins in proteomics studies. Future studies of the putative genes involved in diapause may lead to innovative new methods of insect pest control by disrupting the diapause program.

Technical Abstract: Flesh flies in the genus Sarcophaga are important models for investigating endocrinology, diapause, cold hardiness, reproduction, and immunity. Despite the prominence of Sarcophaga flesh flies as models for insect physiology and biochemistry, and in forensic studies, little genomic or transcriptomic data are available for members of this genus. We used massively parallel pyrosequencing on the Roche 454-FLX platform to produce a substantial EST dataset for the flesh fly Sarcophaga crassipalpis. To maximize sequence diversity, we pooled RNA extracted from whole bodies of all life stages and normalized the cDNA pool after reverse transcription. We obtained 207,110 ESTs with an average read length of 241 bp. These reads assembled into 20,995 contigs and 31,047 singletons. Using BLAST searches of the NR and NT databases we were able to identify 11,757 unique gene elements (E<0.0001) representing approximately 9,000 independent transcripts. Comparison of the distribution of S. crassipalpis unigenes among GO Biological Process functional groups with that of the Drosophila melanogaster transcriptome suggests that our ESTs are broadly representative of the flesh fly transcriptome. In addition, we identified over 500 potential microsatellite loci among our ESTs that could be used in population and systematic studies of S. crassipalpis and other flesh flies.