The use of degenerate-primed random amplification of polymorphic DNA (DP-RAPD) for strain-typing and inferring the genetic similarity among closely related viruses.
Often it is necessary to distinguish among strains of closely related viruses, as well as infer the genetic relatedness within large groups of viruses. Current methods for strain-typing viruses are time-consuming, require significant quantities of extracted DNA, and/or may require a priori genetic information. In this study we modified random amplification of polymorphic DNA (RAPD) by using a degenerate primer to produce unique and reproducible banding patterns from viral genomes. In the degenerate-primed RAPD analysis (DP-RAPD), a selection of algal virus and bacteriophage strains were profiled that encompassed an array of genome sizes and virus families. Closely related viruses (e.g. strains infecting Micromonas pusilla) generated similar, yet unique DP-RAPD patterns that could be readily distinguished from viruses within the same family (Phycodnaviridae) infecting a Chlorella-like alga. As well, marine vibriophage from the families Myoviridae, Siphoviridae, and Podoviridae showed high diversity and were distinct from coliphage and cyanophage. Contamination of host DNA, even at levels above those that would normally be encountered, did not interfere with the viral patterns. These findings describe a rapid, PCR-based tool for strain-typing viral isolates that allows inferences to be made on genetic relatedness within groups of closely related viruses.
Comeau
André M
AM
Department of Microbiology and Immunology, University of British Columbia, Vancouver, Canada V6T 1Z4.
Short
Steven
S
Suttle
Curtis A
CA
D017899:Caudovirales / Q000145:classification* / Q000235:genetics; D016172:DNA Fingerprinting / Q000379:methods*; D017931:DNA Primers; D016679:Genome, Viral; D006090:Gram-Negative Bacteria / Q000821:virology; D017862:Phycodnaviridae / Q000145:classification* / Q000235:genetics; D019105:Random Amplified Polymorphic DNA Technique
D017931:DNA Primers