Agroinfection as a rapid method for propagating Cowpea mosaic virus-based constructs.

To increase the efficiency of infections with Cowpea mosaic virus (CPMV)-based constructs, clones suitable for agroinfection were constructed. Full-length copies of RNA-1 and RNA-2 were inserted between the sequence of a Cauliflower mosaic virus (CaMV) 35S promoter and a nos terminator and were introduced into the Agrobacterium tumefaciens plasmid, pBINPLUS. Infiltration of leaves of either Nicotiana benthamiana or cowpea (Vigna unguiculata) with a bacterial suspension containing a mixture of the RNA-1- and RNA-2-based plasmids resulted in the plants developing typical CPMV symptoms. To confirm the utility of this approach for use with CPMV-based vectors, a GFP construct based on RNA-2 was adapted for agroinfection. Infiltration of N. benthamiana leaves with a mixture of Agrobacteria containing this construct and the RNA-1 plasmid resulted in high levels of GFP expression. The results demonstrate that agroinfection is a suitable method for the propagation of CPMV-based derivatives.