The detection of recombinant, tuber necrosing isolates of Potato virus Y (PVY(NTN)) using a three-primer PCR based in the coat protein gene.
A simple and reliable procedure for reverse transcription-polymerase chain reaction (RT-PCR) detection and strain differentiation of Potato virus Y (PVY) was developed. Three primers were designed within the coat protein (CP) and nuclear inclusion protein b (NIb) region, exploiting a single base polymorphism identified as being present in all the recombinant PVY(NTN) isolates published. Samples infected with PVY produce a single band of 569 bp, while isolates belonging to PVY(NTN) strain give an additional band of 334 bp. The technique was tested on a collection of well-characterised isolates of PVY from a range of strains and was found to detect all of the isolates reported as belonging to the PVY(NTN) strain. All of the isolates detected possess a recombination event within the coat protein. Further sequence analysis revealed that all the recombinant PVY(NTN) isolates reported thus far would be detected using this assay, whilst isolates thought to be PVY(NTN) that do not possess the coat protein recombination event would not be detected.
Moravec
T
T
Institute of Experimental Botany, Czech Academy of Science, Na Karlovce 1, 16000 Prague, Czech Republic. cerovska@ueb.cas.cz
Cerovská
N
N
Boonham
N
N
D001483:Base Sequence; D036022:Capsid Proteins / Q000235:genetics*; D017931:DNA Primers; D012321:DNA-Directed RNA Polymerases; D008969:Molecular Sequence Data; D010935:Plant Diseases / Q000821:virology*; D016133:Polymerase Chain Reaction / Q000379:methods*; D011110:Polymorphism, Genetic; D017800:Potyvirus / Q000235:genetics / Q000302:isolation & purification*; D011995:Recombination, Genetic; D017422:Sequence Analysis, DNA; D011198:Solanum tuberosum / Q000821:virology; D014764:Viral Proteins / Q000235:genetics
D036022:Capsid Proteins; D017931:DNA Primers; D014764:Viral Proteins; C061396:nuclear inclusion protein b, mosaic viruses; D012321:DNA-Directed RNA Polymerases