PVX virus-induced gene silencing (VIGS) of NbIMPα1 and NbIMPα2 genes.

(A) Alignment of AtIMPα1 and two IMP orthologues (NbIMPα1 and NbIMPα2) of N . benthamiana . The amino acid positions are indicated above the alignment. (B and C) Real-time RT-PCR quantification of NbIMPα1 and NbIMPα2 transcripts in the silenced N . benthamiana plants. A partial sequence of either NbIMPα1 or NbIMPα2 was cloned into the PVX vector in a reverse orientation (PVX-IMPα1 or PVX-IMPα2). N . benthamiana plants were co-inoculated with PVX-IMPα1 and PVX-IMPα2 (PVX-IMPα1+2) to simultaneously silence NbIMPα1 and NbIMPα2. Total RNA extracts from the infected leaves were analyzed at 15 dpi. Mean values (±SE) ( n = 4) are fold-changes calculated when the value of PVX is set to 1.0, and the values were analyzed on log-transformed data for a significant difference using Student’s t -test (** P < 0.01, **** P < 0.0001). (D) The PVX RNA levels were quantified by real-time RT-PCR. Mean values (±SE) ( n = 5) are fold-changes calculated when the value of PVX is set to 1.0, and the values were analyzed on log-transformed data for a significant difference using Student’s t -test ( P > 0.05). (TIF)