Tree Genetics and Molecular Breeding 2024, Vol.14, No.2, 81-94 http://genbreedpublisher.com/index.php/tgmb 86 Eucalyptus lines. For example, the functional validation of the NRT2.5 gene, identified through comparative genomics, demonstrated its involvement in the resistance to various biotic stressors, including pathogens like Phytophthora cinnamomi (du Toit et al., 2020). Additionally, the modification of lignin biosynthesis pathways through CRISPR/Cas9-mediated gene editing has resulted in Eucalyptus variants with not only improved resistance to pathogens but also enhanced industrial traits, such as reduced lignin content for better pulping efficiency. These dual benefits highlight the value of integrating functional genomics with breeding programs to achieve both agricultural and industrial objectives (Dai et al., 2020). Overall, the successful functional verification of disease resistance genes in Eucalyptus provides a powerful foundation for the development of disease-resistant cultivars. By leveraging genome editing and other biotechnological tools, researchers can continue to enhance the resilience of Eucalyptus species, ensuring their sustainability in the face of evolving pathogen pressures. 5 Integration of Genome Editing with Traditional Breeding The integration of genome editing technologies with traditional breeding approaches represents a powerful strategy to enhance disease resistance in Eucalyptus (Van Vu et al., 2022). By combining the precision of molecular breeding with the broad genetic diversity harnessed through conventional breeding, researchers can develop more resilient Eucalyptus varieties faster and more efficiently. 5.1 Combining molecular breeding and biotechnological approaches Molecular breeding, which involves the use of genetic markers to select for desirable traits, has already transformed traditional breeding practices. Genome editing technologies like CRISPR/Cas9, TALENs, and ZFNs have further expanded the toolkit available to breeders, allowing for precise modifications of target genes associated with disease resistance (Nerkar et al., 2022). The combination of these biotechnological approaches with traditional breeding methods enables the rapid incorporation of resistance genes into elite Eucalyptus lines. For example, genome editing can be used to introduce resistance genes identified through marker-assisted selection into high-yielding Eucalyptus varieties. This approach allows breeders to combine the broad-spectrum resistance conferred by traditional breeding with the specific, targeted improvements made possible by genome editing. The result is a more effective and efficient breeding process, capable of producing Eucalyptus varieties that are both high-yielding and disease-resistant (Mushtaq et al., 2019). 5.2 Enhancing the efficacy of conventional breeding with genetic insights The integration of genetic insights into conventional breeding programs significantly enhances their efficacy by enabling the selection of traits that would be difficult or impossible to achieve through traditional methods alone. By using genetic markers and genome editing, breeders can overcome the limitations of conventional breeding, such as long generation times and the complex inheritance patterns of certain traits. For instance, the use of genomic selection—where the entire genome is analyzed to predict the performance of specific traits-allows breeders to make more informed decisions when selecting parent plants for breeding. This approach can be complemented by genome editing, which can be used to directly modify or enhance specific genes that contribute to disease resistance. As a result, the breeding cycle is shortened, and the likelihood of success in developing resistant Eucalyptus varieties is increased (Candotti et al., 2022). 5.3 Examples of integrated breeding programs for disease resistance Several integrated breeding programs have successfully combined traditional breeding with genome editing to develop disease-resistant Eucalyptus varieties. One notable example is the use of CRISPR/Cas9 in conjunction with conventional breeding to enhance resistance to Mycosphaerella leaf disease. Researchers first identified resistance-associated loci through traditional breeding and marker-assisted selection, and then used CRISPR/Cas9 to introduce targeted mutations in these loci, resulting in Eucalyptus lines with enhanced disease resistance (Dai et al., 2020).
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