Rapid and efficient in planta genome editing in sorghum using foxtail mosaic virus‐mediated sgRNA delivery
[1];
- DOE Center for Advanced Bioenergy and Bioproducts Innovation St. Paul Minnesota 55108 USA, Department of Genetics, Cell Biology and Development University of Minnesota St. Paul Minnesota 55108 USA, Center for Precision Plant Genomics University of Minnesota St. Paul Minnesota 55108 USA
- Department of Cell and Molecular Biology University of Rhode Island South Kingstown Rhode Island 02881 USA
- Department of Genetics, Cell Biology and Development University of Minnesota St. Paul Minnesota 55108 USA, Center for Precision Plant Genomics University of Minnesota St. Paul Minnesota 55108 USA
- Agronomy Department, Plant Molecular and Cellular Biology Program Genetics Institute University of Florida, IFAS Gainesville Florida USA, DOE Center for Advanced Bioenergy and Bioproducts Innovation Gainesville Florida 32611 USA
SUMMARY The requirement of in vitro tissue culture for the delivery of gene editing reagents limits the application of gene editing to commercially relevant varieties of many crop species. To overcome this bottleneck, plant RNA viruses have been deployed as versatile tools for in planta delivery of recombinant RNA. Viral delivery of single‐guide RNAs (sgRNAs) to transgenic plants that stably express CRISPR‐associated (Cas) endonuclease has been successfully used for targeted mutagenesis in several dicotyledonous and few monocotyledonous plants. Progress with this approach in monocotyledonous plants is limited so far by the availability of effective viral vectors. We engineered a set of foxtail mosaic virus (FoMV) and barley stripe mosaic virus (BSMV) vectors to deliver the fluorescent protein AmCyan to track viral infection and movement in Sorghum bicolor . We further used these viruses to deliver and express sgRNAs to Cas9 and Green Fluorescent Protein (GFP) expressing transgenic sorghum lines, targeting Phytoene desaturase ( PDS ), Magnesium‐chelatase subunit I ( MgCh ), 4‐hydroxy‐3‐methylbut‐2‐enyl diphosphate reductase , orthologs of maize Lemon white1 ( Lw1 ) or GFP . The recombinant BSMV did neither infect sorghum nor deliver or express AmCyan and sgRNAs. In contrast, the recombinant FoMV systemically spread throughout sorghum plants and induced somatic mutations with frequencies reaching up to 60%. This mutagenesis led to visible phenotypic changes, demonstrating the potential of FoMV for in planta gene editing and functional genomics studies in sorghum.
- Research Organization:
- Center for Advanced Bioenergy and Bioproducts Innovation (CABBI), Urbana, IL (United States); University of Florida, Gainesville, FL (United States); University of Minnesota, St. Paul, MN (United States); University of Rhode Island, Kingstown, RI (United States)
- Sponsoring Organization:
- US Department of Energy, Office of Science, Biological and Environmental Research Program; US Department of Energy, Office of Science, Biological and Environmental Research Program under Award Number DE-SC0018420; USDOE; USDOE Office of Science (SC), Biological and Environmental Research (BER); USDOE Office of Science (SC), Biological and Environmental Research (BER). Biological Systems Science (BSS)
- Grant/Contract Number:
- SC0018420
- OSTI ID:
- 2480984
- Journal Information:
- The Plant Journal, Journal Name: The Plant Journal Journal Issue: 2 Vol. 121; ISSN 0960-7412
- Publisher:
- Wiley-BlackwellCopyright Statement
- Country of Publication:
- United Kingdom
- Language:
- English
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