Virus induced gene silencing (VIGS) of the RB pathogen R gene for P. infestans resistance in S. bulbocastanum

(Hailing Jin, University of California, Berkeley)
Figure 1. Virus-induced gene silencing of PDS. A) Seed-geminated S. bulbocastanum. B) Tuber-generated S. bulbocastanum. C) Close up of PDS-silenced region of tuber-generated S. bulbocastanum.The focus of current efforts is on developing an efficient silencing system in tuber-generated potato in order to study the function of candidate genes responsible for certain disease resistance and its signaling pathways. S. bulbocastanum PT29, a wild diploid potato species, is highly resistant to all known races of P. infestans. This broad resistance phenotype is determined by a CC-NBS-LRR type of resistance gene, RB (Song et al., 2003). S. bulbocastanum is sexually incompatible and can only be propagated through tuber vegetative growth making it a difficult host to perform genetic studies. VIGS provides a powerful alternative approach to study gene function. The PDS gene in tuber-generated S. bulbocastanum by inoculating TRV-PDS sap propagated from N. benthamiana resulting in 60-70% of the silenced plants exhibiting a PDS silenced photo-bleached phenotype (Figure 1b). Using the same system, the RB gene in PT29 was silenced.

P. infestans strains, US1 and ME980085, were used to infect RB-silenced PT29 resulting in visible disease symptoms and the loss of resistance (Figure 2). S. tuberosum cultivar Katahdin is susceptible to all the P. infestans strains and was used as a susceptible control. PT29 and TRV-R1 treated PT29 were used as the resistance controls. Four individual experiments with a total of 28 tuber-generated PT29 were used for RB silencing and of those, seventeen silenced-PT29s showed susceptible diseased symptom in the pathogen assay.