Project Details:

Title:
Non-Transgenic Generation of Herbicide Resistance in Soybean Using CRISPR Base Editing

Parent Project: This is the first year of this project.
Checkoff Organization:North Central Soybean Research Program
Categories:Weed control, Breeding & genetics
NCSRP, USB, QSSB Project Code:NCSRP
Project Year:2019
Lead Principal Investigator:Feng Qu (Ohio Agricultural Research and Development Center)
Co-Principal Investigators:
Keywords:

Contributing Organizations

Funding Institutions

Information and Results

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Project Summary

This is a collaborative project among researchers from four different states (Ohio – Qu and Doohan, Nebraska - Clemente, Missouri - Stacy, and Minnesota – Stupar), with the goals to (i) establish a CRISPR base editing system that generates herbicide-resistant soybean quickly and non-transgenically; and then (ii) use the new system to produce novel herbicide resistance traits, ideally double or triple herbicide resistance.

Project Objectives

• Accelerate base editing in soybean by using germinating soybean seed;
• Streamline the base editing protocol in multiple soybean cultivars by equipping them with the base editing enzyme BE3;
• Generate novel herbicide resistance traits in soybean using the new base editing approach.

Project Deliverables

• A fast, cost-effective, and non-transgenic base editing protocol for accurately modifying soybean genes without disrupting their functions;
• Multiple soybean lines resistant to diverse herbicides generated with the new base editing technology;
• Multiple soybean cultivars equipped with the base editing BE3 gene, ready to be utilized by the soybean research community for editing other soybean gene in order to improve soybean seed quality and yield.

Progress of Work

Updated March 29, 2019:
Project title: Non-transgenic generation of herbicide resistance in soybean using CRISPR base editing

This is a collaborative project among researchers from four different states (Ohio – Qu and Doohan, Nebraska - Clemente, Missouri - Stacy, and Minnesota – Stupar), with the goals to (i) establish a base editing system that generates herbicide-resistant soybean quickly and non-transgenically; and then (ii) use the new system to produce novel herbicide resistance traits. Specifically, we proposed to pursue three specific objectives:
• Accelerate base editing in soybean by using germinating soybean seed;
• Streamline the base editing protocol in multiple soybean cultivars by equipping them with the base editing enzyme BE3;
• Generate novel herbicide resistance traits in soybean using the new base editing approach.

Should the project progresses as planned, it is expected to yield the following deliverables that will directly benefit soybean growers by increasing the profit of growing soybean while reducing inputs:
• A fast, cost-effective, and non-transgenic base editing protocol for accurately modifying soybean genes without disrupting their functions;
• Multiple soybean lines resistant to diverse herbicides generated with the new base editing technology;
• Multiple soybean cultivars equipped with the base editing BE3 gene, ready to be utilized by the soybean research community for editing other soybean gene in order to improve soybean seed quality and yield.

In the proposal we set the following milestones and performance indicators for the first year (October 1, 2018 to September 30, 2019):
(1) Successful base editing of the soybean ALS gene by using germinating soybean seed and particle bombardment DNA delivery, generating Imazapyr-resistant soybean seed;
(2) Successful assembly of DNA constructs for producing BE3 (base editor, third generation)-transgenic soybean;
(3) Initiation of soybean transformation to generate BE3-transgenic soybean in a number of soybean cultivars: Williams 82, Thorne, Maverick, Bert, and Jack.

Progress, accomplishments and deliverables to-date:
By now we have carried out six months of NCSRP-funded research. We are making steady progresses towards meeting all three of the milestones.
• Specifically, we have successfully assembled a construct that contains both the BE3 base-editing Cas9 enzyme, and a guide RNA designed to guide BE3 to a specific position in the soybean ALS gene to mediate the editing of specific bases, leading to herbicide (Imazapyr) resistance. This plasmid is named as pWI-BE3-gGmALS. We also produced another construct that would express the BE3 enzyme only, designated as pWI-BE3 (milestone 2).
• We then used both constructs to transform embryogenic tissues generated from young soybean seed, using a particle bombardment procedure. We obtained five (5) transgenic events for pWI-BE3-gGmALS, and one (1) event for pWI-BE3, from which transgenic seedlings are being induced. At this point at least one event appears to contain the successfully base-edited ALS gene. Other events may also yield base-edited seedlings because the construct continues to carry out the editing while the tissues are being induced for seedling differentiation (milestone 1).
• We will next send both the pWI-BE3-gGmALS and pWI-BE3 constructs to collaborators in Nebraska, Missouri, and Minnesota for producing base-edited plants in other soybean varieties (milestone 3).

View uploaded report Word file

Final Project Results

Benefit to Soybean Farmers

If progressing as anticipated, the results of our research will yield soybean varieties that are resistant to multiple herbicides, allowing farmers to control weeds more effectively, with greater consistency and cost-savings. This would in turn enable the growers to increase profitability from growing soybean.

Performance Metrics

Project Years