International Journal of Horticulture, 2024, Vol.14, No.3, 142-155 http://hortherbpublisher.com/index.php/ijh 150 non-target organisms must be addressed through thorough risk assessments and monitoring (Chu et al., 2022). Public acceptance is another critical factor. Despite the potential benefits of CRISPR/Cas9 technology, there is often public skepticism and opposition to genetically edited crops (Zhou et al., 2020). This resistance can stem from concerns about food safety, environmental impacts, and ethical considerations. Transparent communication, public engagement, and education about the safety, benefits, and regulatory oversight of CRISPR/Cas9-edited Citrus are essential to gaining consumer trust and acceptance (Tian et al., 2019; Chen et al., 2021). 6 Field Trials and Performance Evaluation 6.1 Design and implementation of field trials for CRISPR/Cas9 citrus The design and implementation of field trials for CRISPR/Cas9-modified citrus germplasm resistant to the Asian citrus psyllid (ACP) involve several critical steps. Initially, the selection of appropriate test sites is essential, ensuring they represent diverse environmental conditions and pest pressures. The field trials should be randomized and replicated to account for variability and ensure statistical robustness. The CRISPR/Cas9-modified citrus plants are then planted alongside non-modified control plants to provide comparative data on performance metrics (Leong et al., 2022; Chaverra-Rodriguez et al., 2023). Regular monitoring and data collection are crucial, focusing on plant growth, health, and resistance to ACP. The implementation also includes the integration of standard agricultural practices to maintain plant health and productivity throughout the trial period. 6.2 Evaluation metrics for resistance, yield, and fruit quality The evaluation of CRISPR/Cas9-modified Citrus germplasm involves several key metrics, includes resistance to ACP, yield, fruit quality etc. Resistance to ACP is assessed by monitoring the incidence and severity of ACP infestations on the modified plants compared to control plants. The presence of ACP and the extent of damage caused are recorded periodically (Leong et al., 2022; Chaverra-Rodriguez et al., 2023). The yield is measured by the quantity of fruit produced per plant. This includes both the number of fruits and their weight, providing a comprehensive view of the productivity of the modified plants (Leong et al., 2022). Fruit quality is evaluated based on several parameters, including size, color, taste, and nutritional content. These metrics ensure that the genetic modifications do not adversely affect the marketability and consumer acceptance of the fruit (Leong et al., 2022). 6.3 Case studies of field trials and their outcomes Several case studies demonstrate the potential of CRISPR/Cas9 technology in developing ACP-resistant citrus germplasm. For example, a trial conducted in Florida showed that CRISPR/Cas9-modified citrus plants exhibited significantly lower populations of ACP (Asian citrus psyllid) compared to control plants. The modified plants also showed a reduced incidence of Huanglongbing (HLB) symptoms, indicating enhanced resistance to the vector (Tian et al., 2019; Hunter et al., 2020). A trial conducted in California focused on the yield and fruit quality of CRISPR/Cas9-modified citrus plants. Research by Chaverra-Rodriguez et al. (2023) controlled the spread of ACP (Asian citrus psyllid) using CRISPR-Cas9 gene editing technology to curb the spread of HLB (Huanglongbing). The study developed optimized methods for collecting and handling ACP eggs by introducing Cas9 ribonucleoprotein (RNP) into early embryos and adopting alternative methods to inject RNP into adult females for ovary transduction. Through these methods, the research team successfully generated visible somatic mutations in ACP (Figure 4). This provides a theoretical basis for further developing gene-based HLB control systems. Overall, these case studies emphasize the potential of CRISPR/Cas9 technology in developing sustainable solutions to manage ACP and mitigate the impact of HLB on the citrus industry. Continuous evaluation and improvement of these transgenic plants are crucial for their successful integration into commercial citrus production systems.
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