Cotton Genomics and Genetics 2025, Vol.16, No.1, 12-20 http://cropscipublisher.com/index.php/cgg 18 7 Future Directions in Gene Editing for Cotton Improvement 7.1 Integration of multi-omics data for target gene discovery Now, scientists are combining different research methods such as genomics, transcriptomics, proteomics and metabolomics to find important genes related to cotton drought resistance, salt resistance, etc. more quickly. These genes may also control cotton fiber quality and yield. With the continuous progress of cotton whole genome sequencing, researchers can more accurately locate target genes that can be edited, thereby providing better ideas and strategies for improving cotton traits (Peng et al., 2020). Through these omics data, people can discover new editing targets and verify whether they are indeed related to stress resistance or yield (Huang et al., 2021). 7.2 Use of AI and machine learning to optimize guide RNA design How to design guide RNA (gRNA) has always been an unavoidable problem in CRISPR editing. In the early days, it mainly relied on manual experience, but now it is different. Artificial intelligence (AI) and machine learning have also begun to intervene in this field and have done a good job. These algorithms are not omnipotent, but they are very practical in one respect - they can predict in advance which places may be "off-target" and help select the most suitable target sites. Especially when facing a "complex player" like cotton with many genes and many repetitive sequences, it is really not easy to manually design gRNA (Kumar et al., 2024). Of course, this type of AI tool is not without a threshold to use, but once it runs through, the editing process can indeed become more convenient. Not only is the accuracy improved, but the failure rate can also be reduced. For the entire breeding process, it is equivalent to adding an "accelerator". 7.3 Development of transgene-free editing techniques for regulatory acceptance The goal of non-GMO editing technology is to make the final cotton variety free of foreign DNA. Such methods are more likely to pass regulatory approval and be more acceptable to consumers. For example, transient expression of CRISPR/Cas or direct delivery of protein complexes into cells will not leave the GMO fragments in the plant (Mubarik et al., 2021). This will not only reduce concerns about GMOs, but also increase people's trust in gene-edited cotton (Khan et al., 2023). In the future, gene editing research in cotton is likely to rely more and more on the integration of multi-omics methods, the help of artificial intelligence, and non-GMO editing methods to enhance cotton's stress resistance while meeting strict regulatory requirements. 8 Concluding Remarks Now, gene editing, especially CRISPR/Cas9 technology, has become an important tool for improving cotton's drought resistance, salt resistance, heat resistance, cold resistance and other abilities. It can directly modify regulatory genes and functional genes, making the improvement faster and more targeted. This method breaks through the limitations of traditional breeding and transgenic methods. Moreover, it can also breed cotton varieties without foreign genes, which makes it more acceptable and more likely to pass regulation. The next research should focus more on combining different omics data, such as genomes and transcriptomes. This will find new key genes and regulatory networks, and give a clearer understanding of how cotton responds to stress. Artificial intelligence and machine learning are also useful. They can help optimize the design of guide RNAs, improve the success rate of editing, and minimize off-target situations. In addition, if gene editing is to be widely used in cotton breeding, it is also necessary to improve the transformation efficiency and the success rate of tissue regeneration. Researchers, breeders and government departments should communicate more and work together to solve regulatory problems and make more people understand and accept gene-edited crops. Although the goals of stress resistance, stable yield, and high-quality fiber sound ideal, it is becoming increasingly difficult to achieve them using conventional methods. At present, gene editing technology is constantly advancing, and many traits that were originally difficult to deal with can now be accurately "cut". It's not to say that the problem has been solved all at once, but at least the direction is clearer. The tools have been updated, the data has been integrated, and the algorithms are smarter - these new means together make cotton planting less passive in responding to climate change. After all, it's not just as simple as increasing yields. For future food and fiber security, this technical path may be one of the few that still has hope. If the cotton industry wants to be stable, it can't just rely on the old methods. These current advances may be the key to moving forward.
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