CRISPR/Cas genome editing in soybean: challenges and new insights to overcome existing bottlenecks

• Published in: Journal of advanced research
• Main Authors: Freitas-Alves, Nayara Sabrina, Moreira-Pinto, Clidia E., Távora, Fabiano T.P.K., Paes-de-Melo, Bruno, Arraes, Fabricio B.M., Lourenço-Tessutti, Isabela T., Moura, Stéfanie M., Oliveira, Antonio C., Morgante, Carolina V., Qi, Yiping, Fatima Grossi-de-Sa, Maria
• Format: Journal Article
• Language: English
• Published: Egypt Elsevier B.V 18.08.2024
• Subjects: Base editors; Cas12a; CRISPR/Cas9; Crops; Genetic transformation; Glycine max; Base editors; Genetic transformation; Cas12a; CRISPR/Cas9; Glycine max; Crops; Plant crop
• ISSN: 2090-1232; 2090-1224; 2090-1224
• DOI: 10.1016/j.jare.2024.08.024

[Display omitted] •Soybean genome editing has been mainly achieved by the nucleases Cas9, nCas9 derived base editors, and Cas12a.•Bottlenecks in soybean genome editing are related to transformation, editing efficiency and traits improvement in the field.•Challenges in soybean genetic manipulation can be overcome by improving CRISPR cassette design and plant transformation.•RNP-based systems have promising applications since they generate transgene-free mutant plants.•CRISPR/dCas9 is an efficient system to modulate the expression of soybean target genes. Soybean is a worldwide-cultivated crop due to its applications in the food, feed, and biodiesel industries. Genome editing in soybean began with ZFN and TALEN technologies; however, CRISPR/Cas has emerged and shortly became the preferable approach for soybean genome manipulation since it is more precise, easy to handle, and cost-effective. Recent reports have focused on the conventional Cas9 nuclease, Cas9 nickase (nCas9) derived base editors, and Cas12a (formally Cpf1) as the most commonly used genome editors in soybean. Nonetheless, several challenges in the complex plant genetic engineering pipeline need to be overcome to effectively edit the genome of an elite soybean cultivar. These challenges include (1) optimizing CRISPR cassette design (i.e., gRNA and Cas promoters, gRNA design and testing, number of gRNAs, and binary vector), (2) improving transformation frequency, (3) increasing the editing efficiency ratio of targeted plant cells, and (4) improving soybean crop production. This review provides an overview of soybean genome editing using CRISPR/Cas technology, discusses current challenges, and highlights theoretical (insights) and practical suggestions to overcome the existing bottlenecks. The CRISPR/Cas system was discovered as part of the bacterial innate immune system. It has been used as a biotechnological tool for genome editing and efficiently applied in soybean to unveil gene function, improve agronomic traits such as yield and nutritional grain quality, and enhance biotic and abiotic stress tolerance. To date, the efficiency of gRNAs has been validated using protoplasts and hairy root assays, while stable plant transformation relies on Agrobacterium-mediated and particle bombardment methods. Nevertheless, most steps of the CRISPR/Cas workflow require optimizations to achieve a more effective genome editing in soybean plants.