Viral genome sequencing by random priming methods.
BACKGROUND
Most emerging health threats are of zoonotic origin. For the overwhelming majority, their causative agents are RNA viruses which include but are not limited to HIV, Influenza, SARS, Ebola, Dengue, and Hantavirus. Of increasing importance therefore is a better understanding of global viral diversity to enable better surveillance and prediction of pandemic threats; this will require rapid and flexible methods for complete viral genome sequencing.


RESULTS
We have adapted the SISPA methodology 123 to 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 3,000-15,000 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 viral preparations from cell culture supernatant, allantoic fluid and fecal matter.


CONCLUSIONS
The method described is of great utility in generating whole genome assemblies for viruses with little or no available sequence information, viruses from greatly divergent families, previously uncharacterized viruses, or to more fully describe mixed viral infections.
Djikeng
Appolinaire
A
Viral Genomics Group, J, Craig Venter Institute, Rockville, MD 20850, USA. adjikeng@jcvi.org
Halpin
Rebecca
R
Kuzmickas
Ryan
R
Depasse
Jay
J
Feldblyum
Jeremy
J
Sengamalay
Naomi
N
Afonso
Claudio
C
Zhang
Xinsheng
X
Anderson
Norman G
NG
Ghedin
Elodie
E
Spiro
David J
DJ
D001483:Base Sequence; D017931:DNA Primers / Q000737:chemistry; D004267:DNA Viruses / Q000737:chemistry; D016679:Genome, Viral; D023281:Genomics / Q000379:methods*; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction / Q000379:methods; D012328:RNA Viruses / Q000737:chemistry; D017422:Sequence Analysis, DNA / Q000379:methods
D017931:DNA Primers