M. Borja et al., Restoration of wild-type virus by double recombination of tombusvirus mutants with a host transgene, MOL PL MICR, 12(2), 1999, pp. 153-162
Nicotiana benthamiana plants transformed with the coat protein gene of toma
to bushy stunt virus (TBSV) failed to elicit effective virus resistance whe
n inoculated with wildtype virus. Subsequently, R-1 and R-2 progeny from 13
transgenic lines were inoculated with a TBSV mutant containing a defective
coat protein gene. Mild symptoms typical of those elicited in nontransform
ed plants infected with the TBSV mutant initially appeared. However, within
2 to 4 weeks, up to 20% of the transgenic plants sporadically began to dev
elop the lethal syndrome characteristic of wild-type virus infections. RNA
hybridization and immunoblot analyses of these plants and nontransformed N,
benthamiana inoculated with virus from the transgenic lines indicated that
wild-type virus had been regenerated by a double recombination event betwe
en the defective virus and the coat protein transgene, Similar results were
obtained with a TBSV deletion mutant containing a nucleotide sequence mark
er, and with a chimeric cucumber necrosis virus (CNV) containing the defect
ive TBSV coat protein gene. In both cases, purified virions contained wild-
type TBSV RNA or CNV chimeric RNA derived by recombination with the transge
nic coat protein mRNA, These results thus demonstrate that recombinant tomb
us-viruses can arise frequently from viral genes expressed in transgenic pl
ants.