Database Research SummariesImproving CRISPR gene editing in soybean
Research Focus
The focus of this project is to lay a foundation for improving gene editing in soybeans. Researchers have been building up new tools and resources for gene editing in soybeans and initiating tests of their feasibility.
Objectives
- Modification of acetolactate synthase 1 (Als1) and acetolactate synthase 2 (Als2) genes.
- Modification of eIF4E and elFiso4e genes.
- Characterization of edited plants.
Results
- We have built up a CRISPR/Cas9 vector system based on an Arabidopsis egg cell specific promoter and two U6 promoters to express Cas9 and its associated guide RNAs, respectively, in a single binary plasmid vector for Agrobacterium-mediated soybean transformation. The CRISPR construct also contains the 35S promoter driven bar gene suitable for transformation selection with the herbicide bialaphos and a GFP gene under a soybean ubiquitin promoter as a visual marker of transformants.
- Researchers tested the feasibility of the CRISPR system for genome editing in stable soybean transformant, two genes (AGO7a and AGO7b, argonaute genes involved in small RNA mediated gene regulation in soybean) were chosen for targeted mutagenesis.
- We built up a CRISPR/Cpf1 vector system based on an Arabidopsis ubiquitin promoter and two U3 promoters for expression of Cpf1 and its associated guide RNA, respectively, in a single binary vector for Agrobacterium-mediated soybean transformation. To test the feasibility of this CRISPR system for genome editing in soybean, the same two genes (AGO7a and AGO7b) were chosen for targeted mutagenesis with two constructs.
- The two constructs were transferred into Agrobacterium strain EHA101 for stable soybean transformation and Agrobacteriumrhizogenes for hairy root transformation for a quick assessment of activities of these two constructs before stored at ISU.
Importance
- This research will help to reduce CRISPR gene editing to practice and make the technology practical for more quickly breeding new traits into soybean plants.
- This research could also potentially lead to a new virus resistance trait for soybean plants.
For more information about this research project, please visit the National Soybean Checkoff Research Database.
Funded in part by the soybean checkoff.
