Abstract
Downy mildew of grapevine (Vitis vinifera L.), caused by the oomycete pathogen Plasmopara viticola, is one of the most serious concerns for grape production worldwide. It has been widely reported that the pathogenesis-related 4 (PR4) protein plays important roles in plant resistance to diseases. However, little is known about the role of PR4 in the defense of grapevine against P. viticola. In this study, we engineered loss-of-function mutations in the VvPR4b gene from the cultivar “Thompson Seedless” using the CRISPR/Cas9 system and evaluated the consequences for downy mildew resistance. Sequencing results showed that deletions were the main type of mutation introduced and that no off-target events occurred. Infection assays using leaf discs showed that, compared to wild-type plants, the VvPR4b knockout lines had increased susceptibility to P. viticola. This was accompanied by reduced accumulation of reactive oxygen species around stomata. Measurement of the relative genomic abundance of P. viticola in VvPR4b knockout lines also demonstrated that the mutants had increased susceptibility to the pathogen. Our results confirm that VvPR4b plays an active role in the defense of grapevine against downy mildew.
Highlights
Grapevine is one of the most economically important fruit crops worldwide and is widely used for producing wine, juice, and dried and fresh fruit[1]
The expression cassette was cloned into the linearized vector by homologous recombination to obtain the VvPR4b knockout vector, which contains a single guide RNA (Fig. 1b)
There have been many studies on the pathology of the causative agent, P. viticola, few studies related to functional analyses of resistance genes in grapevine have been investigated
Summary
Grapevine is one of the most economically important fruit crops worldwide and is widely used for producing wine, juice, and dried and fresh fruit[1]. Et Curt.) Berlese and De Toni, is among the most serious fungal diseases of grapevine worldwide[2]. Grapevine downy mildew, caused by the oomycete P. viticola This pathogen infects green parts such as leaves, petioles, new shoots, and young fruit[3]. Pathogenesis-related (PR) proteins comprise a large and heterogeneous class of plant disease defense proteins. Transgenic plants expressing PR proteins at high levels show significantly higher resistance to various pathogens
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