Rapid identification of myxoma virus variants by long-range PCR and restriction fragment length polymorphism analysis.
A long-range PCR method directed at the Myxoma virus (MV) left hand and right hand terminal inverted repeats (TIRs) for rapid amplification of genomic DNA and MV isolate differentiation by restriction fragment length polymorphism (RFLP) analysis is described. The efficacy of this method was tested by comparing the results from full genome RFLPs with those from TIRs amplified separately using reference strain Lausanne (Lu) and a field MV strain characterised previously for its virulence in rabbits. The usefulness of this method was also demonstrated by amplifying MV DNA directly from the eyelid tissue of an infected rabbit and comparative RFLP analysis with respect to Lu. The results proved the long-range PCR technique to be a simple highly efficient method for identifying mutations between MV genomes by RFLP analyses of the amplified TIRs and may be used in future studies to identify variable regions for phylogenetic studies.
Dalton
Kevin P
KP
Instituto Universitario de Biotecnología de Asturias, Departamento de Bioquímica y Biología Molecular, Edificio Santiago Gascón, Campus El Cristo, Universidad de Oviedo, Oviedo, Spain. daltonkevin@uniovi.es
Ringleb
Franziska
F
Martín Alonso
Jose Manuel
JM
Parra
Francisco
F
D000818:Animals; D004279:DNA, Viral / Q000032:analysis / Q000235:genetics; D014644:Genetic Variation; D009233:Myxoma virus / Q000235:genetics / Q000302:isolation & purification; D009234:Myxomatosis, Infectious / Q000453:epidemiology / Q000821:virology*; D016133:Polymerase Chain Reaction / Q000379:methods; D012150:Polymorphism, Restriction Fragment Length; D011817:Rabbits; D012680:Sensitivity and Specificity; D020079:Terminal Repeat Sequences
D004279:DNA, Viral