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United States Department of Agriculture

Agricultural Research Service

Research Project: NEWCASTLE DISEASE EPIDEMIOLOGY, PATHOGENESIS, AND CONTROL

Location: Exotic and Emerging Avian Viral Diseases Research Unit

Title: Viral genome sequencing bt random priming methods

Authors
item Djikeng, A - THE INST GENOMIC RESEARCH
item Halpin, R - THE INST GENOMIC RESEARCH
item Kuzmickas, R - THE INST GENOMIC RESEARCH
item Sengamalay, N - THE INST GENOMIC RESEARCH
item Afonso, Claudio
item Xang, X - THE OHIO STATE UNIV
item Anderson, N - VIRAL DEFENSE FOUNDATION
item Ghedin, E - UNIVERSITY OF PITTSBURGH
item Spiro, D - THE INST GENOMIC RESEARCH

Submitted to: Biomed Central (BMC) Genomics
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: January 8, 2008
Publication Date: January 30, 2008
Citation: Djikeng, A., Halpin, R., Kuzmickas, R., Sengamalay, N., Afonso, C.L., Xang, X., Anderson, N.G., Ghedin, E., Spiro, D.J. 2008. Viral genome sequencing by random priming methods. Biomed Central (BMC) Genomics. 9:5.

Interpretive Summary: The causative agents of many important diseases are viruses. Of increasing importance is an understanding of the viral diversity to enable better surveillance and prediction of pandemic threats. Increased understanding of viral diversity will require rapid and flexible methods for complete genome sequencing. Here a technology for rapid sequencing has been adapted for the complete genome sequencing of several RNA and DNA viruses. obtaining near complete genome sequence of viruses ranging in size from 3000-15,000 kb with a median depth of coverage of 14.33. The utility of this technique to generate full viral genome sequence in the presence of host contaminants, using cell culture purified viruses and viral preps from cell culture supernatant, allantoic fluid and fecal matter is demonstrated. The method described is potentially of great utility in generating whole genome assemblies for viruses with little or no available sequence information, viruses greatly divergent from sequenced family members, and previously uncharacterized viruses.

Technical Abstract: Most emerging health threats are of zoonotic origin. For the overwhelming majority, their causative agents are viruses which include but are not limited to HIV, Influenza, SARS, Ebola, Dengue, and Hantavirus. Of increasing importance therefore is an understanding of the viral diversity to enable better surveillance and prediction of pandemic threats. Increased understanding of viral diversity will require rapid and flexible methods for complete genome sequencing. Results: We have adapted the SISPA-DNase I methodology (Allander et al. 12891-96) to complete genome sequencing of RNA and DNA viruses. We have demonstrated the utility of the method on various types and sources of viruses, obtaining near complete genome sequence of viruses ranging in size from 3000-15,000 kb with a median depth of coverage of 14.33. We used this technique to generate full viral genome sequence in the presence of host contaminants, using cell culture purified viruses and viral preps from cell culture supernatant, allantoic fluid and fecal matter. Conclusion: The method described is potentially of great utility in generating whole genome assemblies for viruses with little or no available sequence information, viruses greatly divergent from sequenced family members, and previously uncharacterized viruses.

Last Modified: 4/16/2014
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