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Arch Virol. 2015 Feb;160(2):517-21. doi: 10.1007/s00705-014-2266-1. Epub 2014 Nov 23.

Construction of infectious clones of tomato torrado virus and their delivery by agroinfiltration.

Author information

1
Interdepartmental Laboratory of Molecular Biology, Institute of Plant Protection, National Research Institute, Władysława Węgorka 20 St, 60-318, Poznań, Poland.

Abstract

The first biologically active infectious clones of tomato torrado virus (ToTV) were generated and delivered into Nicotiana benthamiana and Solanum lycopersicum plants via Agrobacterium tumefaciens. The engineered constructs consisted of PCR-amplified complementary DNAs derived from the ToTV RNA1 and RNA2 components, individually inserted into an engineered pGreen binary vector between the CaMV 35S promoter and nopaline synthase terminator. These constructs were introduced into the plant hosts by means of A. tumefaciens-mediated infiltration. In the presence of the progeny virus, typical symptoms of ToTV infection developed in N. benthamiana and S. lycopersicum. Moreover, the virus was sap-transmissible when isolated from agroinfiltrated plants and induced symptoms similar to those caused by the wild-type virus. The presence of viral particles and viral genetic material was confirmed by electron microscopy and re-inoculation to S. lycopersicum and N. benthamiana, as well as by reverse transcription polymerase chain reaction and high-resolution melt analysis.

PMID:
25416854
PMCID:
PMC4315485
DOI:
10.1007/s00705-014-2266-1
[Indexed for MEDLINE]
Free PMC Article

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