Performing Department
(N/A)
Non Technical Summary
Fungal pathogens cause crop yield and quality reduction leading to billions of dollars lost annually in the US. Fungicide resistance development and environmental toxicity drive an urgent need for novel fungal control strategies. Sudden Death Syndrome (SDS) in soybeans is caused by the soil borne fungus Fusarium virguliforme which infects young soy seedlings through the roots. Infection most often occurs on cool wet soils and disease symptoms in the adult plant are caused by toxin movement from roots to shoots seen first as yellowing of leaves and later as browning and leaf loss. In severe cases this leaf damage causes 50% crop loss. First observed in 1971 in Arkansas, F. virguliforme is now distributed throughout most soy growing regions of the United States. RNA interference (RNAi) can help plants use their own natural defense systems to protect themselves from soil borne fungi as they emerge from the soil and grow as young seedlings. The dsRNA leading to this response is inherently non-toxic and highly selective, thus environmentally friendly and able to preserve beneficial microbes as opposed to chemical fungicides. GreenLight Biosciences has shown dsRNA can be delivered to germinating soybean seeds and result in protection from this prevalent soil fungus Fusarium virguliforme described above. The goal of this project is to advance our methods to develop a more commercially viable proof-of-concept seed treatment. This will provide farmers with a novel, environmentally friendly mode of action for SDS protection. Our objectives are to 1.) Define a matrix of soy-compatible seed treatment mixes that can 2.) carry dsRNA without significant degradation on the seed prior to germination and 3.) release dsRNA into the seed during germination and thus 4.) result in natural defense pathway regulation to provide protection against F. virguliforme. This commercial soy seed treatment platform of dsRNA delivery could be broadly applicable to provide defense against additional fungi. It may be further extended to create other types of beneficial crop traits providing suites of environmentally sound crop protection and enhancement options for the farmer.
Animal Health Component
100%
Research Effort Categories
Basic
0%
Applied
100%
Developmental
0%
Goals / Objectives
Goal 1.Identify 2 to 5RNA soy seed treatments that activate defenseresponses in plants through RNAi.Objective 1. Discover a panel of 10 to 20 soy seed treatments likely to successfully deliverRNA that are non-phytotoxic in soy.Objective 2. CharacterizeRNA behavior in thetreatments to further select those with best potential forRNA delivery.Objective 3.Identify most effective 2-5 treatments thatactivate plant defense via RNAi.Goal 2.DemonstrateFusarium virguliforme soy protection by the RNA soy seed treatmentin growth chamber assays.Objective 1. Identify at least 1 seed treatment that can provide Fusarium virguliforme protection via RNAi in a chamber setting.
Project Methods
Our goal is to take our currently successful seed soaking method of protecting soy seedings from F. virguliforme and translate it to a proof-of-concept scalable soy seed treatment. To do this we will use our in-house knowledge of formulations chemistry as it relates to RNA delivery into plants and other organisms to devise a large panel of candidate seed treatments likely to deliver RNA to the soy seedling after planting during the germination process.These will be tested for overt toxicity to the soy seed and seedling by tests of germination, emergence from the soil and seedling vigor. Germination and emergence are quantified by scoring root radical emergence from the seed coat (3mm) and emergence from the soil (cotyledons above soil line) respectively. Seedling vigor scores are a typical measure in the field of Agriculture and are done based on visual rating from 1 to 9 against a set of standard images.Those not overtly toxic will be characterized for various physical properties regarding RNA interactions and stability and tested for delivery of RNA to the cytoplasm of the soy seedling by molecular methods testing both target gene downregulation and changes in key genes related to successful fungal defense. The development of the currently successful version of seedling protection used this molecular output as an indicator of successful delivery and we will use the same set of materials for this study.Those formulations that pass the previous test will then be tested for fungal protection in chamber assays provided by an outside academic institution with experience in the assay. This consists of sowing treated soy seeds vs non-target and untreated controls in small pots of soil pre-inoculated with F. virguliforme and measuring shoot height, shoot dry weight, root length, root dry weight and statistically determining the effects of the seed treatment. A commercially available chemical seed treatment will also be used for comparison. Experimental outcomes will be confirmed by repeating the experiment at least one more time and additionally testing for the expected gene expression changes.