The era of precision breeding is coming! Bayer opens the era of flexible editing of soybean and corn genotypes - TREDMIL

Recently, Dr. Yu Rong Chen and Dr. Edward Cargill of Bayer Crop Science Research Headquarters in Missouri, USA, are co-corresponding authors of the paper entitled "Simultaneous Genetic Transformation and Genome Editing" published in aBIOTECH. The research paper "of Mixed Lines in Soybean (Glycine max) and Maize (Zea mays)" reports a method of mixed line transformation and editing, which can simultaneously transform and edit many excellent genotypes in soybean and maize.

The study reports a method for simultaneous Transformation and Editing of multiple genotypes by Mixed lines (genotypes) prior to production of seed embryo explants, called Transformation and Editing of Mixed lines (TREDMIL). In this approach, many lines are mixed before producing seed embryo explants, and then, using genome editing technology, the mixed lines are transformed to produce insertions and deletions in the target gene region by co-expressing soybean Dt1 specific or corn BM3-specific CRISPR RNAs (crRNAs) and nuclease Cas12a. After regeneration, the strain identity was decoded by genotype identification, and the edited results were verified by amplicon sequencing.

▲ Fig.1  Simultaneous editing of multiple elite soybean and maize with no obvious bias

 By genotypic identification of regeneration events, we found that 97% (101) of the 104 good soybean genotypes were simultaneously transformed, and the recovered inverters were distributed in different mature groups (MG) from 00 to VII. Similarly, 55%(22) of the 40 superior maize inbred lines were simultaneously transformed, and these converters were distributed in different relative maturity (RM) from 92 to 117. Amplification sequencing showed that 94% of the 101 soybean transformed lines were edits at the target Dt1, and more than 80% of the transformed soybean lines produced more than 90% of the editing events for that line. Similarly, in maize conversion lines, the editing of the target Bm3 reached 69%, involving 17 of the 22 converted female inbred lines. These results suggest that seed embryo transformation systems can be used for efficient, genotypically flexible transformation and genome editing.

▲ Fig.2  Distribution of distinct edits across maize and soybean

Extend 

CRISPR/Cas genome editing technology has become a powerful tool for crop breeding due to its powerful functions, bringing revolutionary changes to the agricultural field. This technique not only enables precise modification of genes, but also combines with flexible plant transformation methods of genotypes to theoretically create targeted variations at any location in different germplasm of a species. However, despite the rapid development of genome editing technology itself, the development of plant transformation technology is relatively backward, which limits the widespread application of gene editing technology in crop breeding to a certain extent.

The development of plant transformation technology 

The technology has advanced significantly since scientists first successfully transformed plants in the mid-1980s. From initial suspension culture to callus culture to the evolution of immature embryos, plant transformation techniques are continuously optimized. In recent years, by using developmental regulators such as BBM and WUS, as well as growth regulators GRFs and GRF-interaction factors GIF and GRF-GIF chimeras, scientists have been able to promote somatic embryogenesis, improve the regenerative ability of some monocotyledonous and dicotyledonous species, and make the transformation of difficult-to-transform genotypes more successful. Nevertheless, these methods still have certain limitations and require further innovation and improvement.

TREDMIL technology innovation and breakthrough 

In a research paper published in aBIOTECH, a research team from Bayer Crop Science Research Headquarters in Missouri, USA, introduces an innovative hybrid line transformation and editing (TREDMIL) method. In this method, several plants of different genotypes are mixed together for transformation and editing before seed embryo explants are produced. Insertion deletions in the target gene region are generated by co-expression of soybean Dt1 specific or corn BM3-specific CRISPR RNAs (crRNAs) and nuclease Cas12a. After regeneration, the strain identity was decoded by genotype identification, and the edited results were verified by amplicon sequencing.

The successful implementation of TREDMIL technology shows its many advantages in crop breeding. First, it can transform and edit multiple genotypes at the same time, which greatly improves the efficiency of breeding. Second, the method is suitable for a variety of different genotypes, including those that are difficult to transform. In addition, through genotype identification and amplicon sequencing, it is possible to ensure that each strain can be accurately identified and verify the accuracy of the edited results.

Application results of TREDMIL technology 

By genotyping regeneration events, the researchers found that 97 percent (101) of 104 good soybean genotypes were simultaneously transformed, and 55 percent (22) of 40 good maize inbred lines were simultaneously transformed. These transformants were distributed in different maturity groups and relative maturity, showing genotype flexibility. In all 101 soybean conversion lines, the editing of target Dt1 reached 94%, while in maize conversion lines, the editing of target Bm3 reached 69%. These results show that TREDMIL technology can achieve efficient, genotypic flexible transformation and genome editing.

Diversity of editing characteristics and breeding significance 

Further studies have shown that large edited spectra generated in different germplasm resources can facilitate genomic discovery. For example, 45 percent of editing events in 98 different genotypes in soybeans detected seven base deletions at the Dt-1389 site, while 65 percent of editing events in corn had significant three base deletions at Bm3-2070. This diversity of editing characteristics provides breeders with valuable information that helps assess germplasm-editing event interactions early in the breeding program, leading to more informed breeding decisions.

Future prospects for precision breeding 

The application of TREDMIL technology indicates the coming of the era of precision breeding. With this approach, breeders can accelerate the development of new varieties, address global food security and environmental challenges, and improve crop yields and quality. As technology continues to improve and optimize, we have reason to believe that future crop breeding will be more efficient and precise, and better able to meet human needs for healthy and sustainable food.

Conclusion 

The introduction of CRISPR/Cas genome editing technology, especially the TREDMIL method, has revolutionized crop breeding. It not only improves the efficiency and precision of breeding, but also provides new tools for addressing global food security and environmental challenges. As technology continues to advance and optimize, we expect to see more innovations and breakthroughs in crop breeding in the future to achieve more efficient and precise crop improvement to meet humanity's growing demand for healthy and sustainable food.