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ARS Home » Northeast Area » Frederick, Maryland » Foreign Disease-Weed Science Research » Research » Publications at this Location » Publication #396974

Research Project: Developing Genomic and Biological Resources to Characterize, Diagnose and Detect Emerging and Invasive Vectored Bacterial and Viral Plant Pathogens for Safeguarding U.S. Agriculture

Location: Foreign Disease-Weed Science Research

Title: Maize rayado fino virus-based tool for high penetrance virus-induced gene silencing

item MLOTSHWA, SIZO - The Ohio State University
item XU, JUNHUAN - The Ohio State University
item Willie, Kristen
item KHATRI, NITIKA - The Ohio State University
item Marty, Deemarie - Dee
item Stewart, Lucy

Submitted to: Mid Atlantic Plant Molecular Biology Society Conference
Publication Type: Abstract Only
Publication Acceptance Date: 8/13/2021
Publication Date: 8/13/2021
Citation: Mlotshwa, S., Xu, J., Willie, K.J., Khatri, N., Marty, D., Stewart, L.R. 2021. Maize rayado fino virus-based tool for high penetrance virus-induced gene silencing. Mid Atlantic Plant Molecular Biology Society Conference .

Interpretive Summary:

Technical Abstract: Maize rayado fino virus (MRFV) is the type species of the phloem-limited genus genus Marafivirus in the family Tymoviridae. It infects maize (Zea mays), its natural host, to which it is transmitted by leafhoppers including Dalbulus maidis and Graminella nigrifrons in a persistent-propagative manner. The MRFV monopartite RNA genome encodes a precursor polyprotein that is processed into replication-associated proteins. The genome is encapsidated by two carboxy co-terminal coat proteins, CP1 and CP2. Cloned MRFV can be readily transmitted to maize by vascular puncture inoculation (VPI), and such virus systems that can be used in maize are valuable to examine plant gene function by gene silencing. However, the efficacy of marafiviruses for virus-induced gene silencing (VIGS) has not been investigated to date. To this end, MRFV genomic loci were tested for their potential to host foreign insertions without attenuating virus viability. This was done using infectious MRFV clones engineered to carry maize phytoene desaturase (PDS) gene fragments (ZmPDS) at various genomic regions. Several MRFV-PDS constructs were generated and tested for infectivity and VIGS in maize. This culminated in identification of the helicase/polymerase (HEL/POL) junction as a viable insertion site that preserved virus infectivity, as well as several sites at which sequence insertion caused loss of virus infectivity. Transcripts of viable constructs, carrying PDS inserts in the HEL/POL junction, induced stable local and systemic MRFV symptoms similar to wild-type infections, and triggered PDS VIGS initiating in veins and spreading into both inoculated and non-inoculated leaves. These constructs were remarkably stable, retaining inserted sequences for at least four VPI passages while maintaining transmissibility by D. maidis. Our data thus identify the MRFV HEL/POL junction as a highly stable insertion site for gene silencing in maize.