Genomics and Applied Biology 2024, Vol.15, No.1, 47-53 http://bioscipublisher.com/index.php/gab 50 Researchers used the CRISPR/Cas9 system to target and edit the plant enolpyruvate dehydrogenase (MePDS) gene in cassava. The MePDS gene is involved in the synthesis pathway of chlorophyll. When the gene is knocked out or its function is impaired, plants will exhibit a clear albino phenotype, that is, the leaves lose their green color, which is due to the inhibition of chlorophyll synthesis. This easily recognizable phenotype has become an intuitive indicator for evaluating gene editing efficiency (Gomez et al., 2023). Through this approach, researchers have observed a high frequency of gene mutation events, including insertion, deletion, and point mutations of gene sequences. This not only demonstrates the high efficiency of CRISPR/Cas9 technology in cassava, but also provides the possibility for subsequent screening of mutant plants with disease resistance or other excellent traits. More importantly, although MePDS editing itself does not directly confer disease resistance, it demonstrates how to precisely manipulate the genome of cassava through the CRISPR/Cas9 system. Based on this, scientists can further target genes directly associated with pathogen resistance, such as genes encoding disease resistant proteins or regulatory factors involved in plant immune responses. By precisely knocking out, modifying, or introducing new gene fragments, new varieties that can resist specific diseases can be created, such as those that can effectively combat cassava mosaic virus or African cassava mosaic virus (Tussipkan and Manabayeva, 2021). The flexibility and accuracy of CRISPR/Cas9 technology also means that solutions can be customized for major diseases in different regions, thereby improving cassava's disease resistance and overall productivity on a global scale. With the continuous progress of gene editing technology and the expansion of its application scope, it is expected to see more gene edited cassava varieties in the future. Not only have strong disease resistance, but they may also have better nutritional value, stress tolerance, and higher yields, making significant contributions to ensuring food security in tropical regions and even globally. 3.3 Potential and challenges of improving cassava disease resistance through CRISPR/Cas technology While CRISPR/Cas technology offers a promising avenue for enhancing disease resistance in cassava, there are several challenges and limitations to consider. The precision and robustness of CRISPR/Cas9 make it a valuable tool for crop improvement, but the development of transgene-free disease-resistant crops is still in its infancy (Cao et al., 2020). Resistance against CRISPR/Cas9 gene drive, a method to spread genetic modifications, is also a concern, as resistance can evolve almost inevitably in natural populations. This highlights the need for strategies to suppress resistance mechanisms or to use resistance as a control method. Despite these challenges, CRISPR/Cas technology holds great potential for the rapid and precise genetic improvement of cassava disease resistance, which is crucial for sustainable agricultural production and global food security (Cao et al., 2020). 4 Case Studies: Specific Application of CRISPR/Cas in Cassava Disease Resistance Breeding 4.1 A case study on using CRISPR/Cas technology to improve the disease resistance of cassava The application of CRISPR/Cas technology in cassava disease resistance breeding has shown promising results. One specific case involves the targeting of susceptibility genes in cassava that are exploited by pathogens. By using CRISPR/Cas systems, researchers can knock out these genes, thereby disrupting the life cycle of the pathogen and conferring resistance to the plant (Schenke and Cai, 2020). Genes that facilitate the infection of cassava by viruses can be edited to create loss-of-function mutations, which in turn prevent the virus from successfully infecting the plant. This approach has been particularly effective against DNA viruses, where CRISPR/Cas9 can directly target viral sequences, and RNA viruses, where the technology is used to modify host plant genes to confer resistance (Robertson et al., 2022). Another case study involves the use of CRISPR/Cas9 for the development of transgene-free disease-resistant crops. By targeting and editing endogenous genes within the cassava genome, researchers can enhance the plant's natural defense mechanisms without the introduction of foreign DNA, thus avoiding the regulatory hurdles associated with genetically modified organisms (GMOs) (Ahmad et al., 2020). This is particularly important for cassava, which is a staple food crop in many developing countries where GMOs may not be readily accepted.
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