The infectivity of dimeric potato yellow mosaic geminivirus clones in different hosts

Abstract

Head-to-tail dimeric clones of both DNA A and DNA B of potato yellow mosaic geminivirus (PYMV) were constructed. These constructs were infectious when inoculated onto Nicotiana benthamiana plants either as DNA or by agroinoculation and were also infectious for tomato plants by agroinoculation. The dimers were not infectious for potato plants following inoculation by either method. Symptom induction required both DNA A and DNA B but agroinoculation with DNA A alone resulted in virus spread in 30% of the inoculated N. benthamiana plants. Leaf disc explants of N. benthamiana, tomato and potato could all be infected by agroinoculation indicating that the method of delivery of the DNA to intact potato plants was unsuitable for successful inoculation rather than an inherent inability of the virus to replicate/spread in potato per se.

Mutational analysis of potato yellow mosaic geminivirus

“…The nucleotide sequences of the mutants were determined by the dideoxy nucleotide chain termination procedure of Sanger et aL (t977) using appropriate primers. Maintenance of viable PYMV DNA A mutants was assessed using total nucleic acid extracts from infected plants and PCR amplification or, for those mutations which created restriction sites, digestion with the appropriate restriction endonuclease and Southern analysis (Buragohain et al, 1994). For PCR amplification, leaf extracts were produced by grinding nitrogen pulverized tissue (0.5 g) in a buffer (1 ml: 10 mM-Tris-HC1 pH 7'0, 100 mM-NaC1, 10 mM EDTA, 1% SDS) prior to three extractions with 50 mM-Tris-HC1 pH 7.0, saturated phenol, nucleic acid precipitation and resuspension in water.…”

“…Dimeric constructs of all the DNA A mutants were created after excision with HindIII and ligation into HindIII restricted pBinl9 as previously described (Buragohain et al, 1994).…”

“…Following agroinoculation of N. benthamiana plants with dimers of the PYMV genome we demonstrated that both components were required to cause systemic symptoms of * Author for correspondence. Fax +44 17l 584 2056. e-mail r.coutts @ ic.ac.uk infection (Buragohain et al, 1994) but that similar to both African cassava mosaic geminivirus (ACMV) and AbMV (Klinkenberg & Stanley, 1990;Evans & Jeske, 1993a) the PYMV A component replicated and spread independently in a limited fashion in N. benthamiana plants. Mixtures of the monomers or dimers of the complete PYMV genome were not infectious for potato, the natural host of the virus (Buragohain et al, 1994), including the same potato cultivar (Desiree) that was successfully infected by grafting from systemically infected N. benthamiana plants (Roberts et al, 1988).…”

“…Fax +44 17l 584 2056. e-mail r.coutts @ ic.ac.uk infection (Buragohain et al, 1994) but that similar to both African cassava mosaic geminivirus (ACMV) and AbMV (Klinkenberg & Stanley, 1990;Evans & Jeske, 1993a) the PYMV A component replicated and spread independently in a limited fashion in N. benthamiana plants. Mixtures of the monomers or dimers of the complete PYMV genome were not infectious for potato, the natural host of the virus (Buragohain et al, 1994), including the same potato cultivar (Desiree) that was successfully infected by grafting from systemically infected N. benthamiana plants (Roberts et al, 1988). This observation suggested that the inoculum was not delivered to the correct site(s) to initiate infection and confirmed difficulties often encountered when re-introducing cloned geminivirus DNAs back into their original hosts (Gilbertson et al, 1991;Garz6n-Tinado et al, 1993).…”

Potato yellow mosaic geminivirus AC2 protein is a sequence non‐specific DNA binding protein

“…Potato yellow mosaic virus (PYMV) belongs to the bipartite (,eminivirus genus in subgroup III [2] and is the only geminiv'rus known to infect potato naturally [3]. The PYMV geno~ne consists of two DNA components designated A and B [2] both of which are required for infectivity [4,5]. The PYMV g~:nome contains six or seven open reading frames (ORFs) hich are transcribed bidirectionally and are separated by al intergenic region, termed the common region (CR), which is almost identical in both DNAs [2].…”