Spray-induced gene silencing enables the characterization of gene function during pre-penetration stages in Blumeria graminis f. sp. tritici

Meihui Zhang¹Meijiao Zhou^1,2Yunmeng Jia^1,3Xipeng Li^1,4Aolin Wang¹Tai-Guo Liu¹Yilin Zhou¹Wei Liu^1*Jieru Fan^1*

• ¹Chinese Academy of Agricultural Sciences (CAAS), Beijing, China
• ²College of Plant Protection, Shenyang Agricultural University, Shenyang, China
• ³College of Agriculture and Forestry Science and Technology, Hebei North University, Zhangjiakou, China
• ⁴Heilongjiang Provincial Key Laboratory of Crop-Pest Interaction Biology and Ecological Control, Heilongjiang Bayi Agricultural University, Daqing, China

Blumeria graminis f. sp. tritici (Bgt), the causal agent of wheat powdery mildew, poses a significant threat to global wheat production. As an obligate biotroph, Bgt is recalcitrant to stable genetic manipulation. Although host-induced gene silencing has been used for gene function studies, it remains ineffective for targeting genes active during pre-penetration stages. Consequently, the functional roles of many Bgt genes during pre-penetration stages remain largely unexplored. In this study, the feasibility of spray-induced gene silencing (SIGS) to characterize gene function during pre-penetration stages was evaluated. The results demonstrated that Bgt conidia and germ tubes efficiently took up environmental double-stranded RNA (dsRNA), enabling the targeted silencing of BgtActin. Exogenous application of BgtActin-dsRNA effectively reduced target gene expression and impaired infection of Bgt. BgtActin silencing predominantly induced abnormal appressoria and reduced disease severity when dsRNA was applied at 6 and 10 hours post-inoculation (hpi). In contrast, BgtActin was almost not silenced when dsRNA was applied at 2 hpi. These findings established SIGS as a promising tool for gene functional studies during the pre-penetration stages of Bgt and highlight the potential of RNA-based strategies for the control of wheat powdery mildew.