Maize Genomics and Genetics 2025, Vol.16, No.4, 229-238 http://cropscipublisher.com/index.php/mgg 234 6.3 Opportunities for stacking multiple resistance traits In the past, it was very complicated to make one crop resistant to multiple herbicides. Now, with base editing, modifications can be made simultaneously or step by step at one or more gene loci. For instance, in wheat, after editing targeting genes such as ALS and ACCase, the crop has developed resistance to various herbicides (Zhang et al., 2020). This strategy can effectively broaden the range of herbicide options and also reduce the risk of resistance failure, which is of great help to weed management. 7 Challenges and Limitations of Base Editing in Maize 7.1 Off-target effects and unintended base conversions Base editing sounds quite precise, but in fact, it is not completely without deviation. Sometimes, even if the editors are well designed, they can still cause some unexpected changes nearby. For instance, cytosine and adenine editors are prone to trigger bystander mutations around the target area. Moreover, if certain regions on the genome are only partially similar to the guide RNA, they may also be "mistakenly damaged". Research has found that adenine editors like ZmAYBEv3 are not inefficient in corn, but chimerism and some unwanted edits can still be observed. This indicates that even if sgRNA and the editor itself are optimized, these risks cannot be completely avoided. Another issue is that the "window" of the editor is relatively small. Some areas cannot be edited at all because the sequence is not appropriate. The current editors can mainly achieve C to T and A to G changes. Other types of base changes are still difficult to achieve (Jeong et al., 2020). 7.2 Efficiency variations across maize genotypes Not all corn varieties are equally "cooperative" with base editing. In other words, some varieties are easy to edit, while others are particularly difficult to handle. The transformation method, the degree of response of tissue culture, and whether chromatin is easily accessible will all affect the editing effect. Especially for some excellent but not easily convertible inbred lines, there are relatively greater limitations in their application. Even if an editor like ZmAYBEv3 performs well in some strains, in others, there may be problems such as incomplete editing or difficulty in passing it on. Some studies have pointed out that in many cases, only a small number of T0 plants exhibit stably inherited target mutations (Fierlej et al., 2022). In addition, whether to use a gene gun or Agrobacterium and other delivery methods, the efficiency will also vary due to the differences between the target genes and the corn itself. 7.3 Regulatory, biosafety, and public acceptance issues Regarding the regulation of base-edited crops, not all national standards are the same. In some places, these plants are not classified as genetically modified because they do not contain exogenous DNA. But elsewhere, they might still be regarded as genetically modified organisms. This makes it very difficult for developers and breeders to determine how to proceed (Rai et al., 2025). In addition to regulations, biosafety issues cannot be ignored either, such as the ecological impact that off-target mutations may bring, and detailed molecular testing is also needed to confirm that there are no hidden exogenous fragments. Finally, there is another point that is often overlooked: how the public views it. Even if the scientific community has proven that these crops are safe, if consumers do not accept them, it will be very difficult to bring these achievements to the market (Namata et al., 2025). 8 Future Perspectives for Base Editing in Maize Herbicide Resistance Improvement 8.1 Integration of base editing with genomic selection and high-throughput phenotyping In the future, corn breeding will no longer rely solely on traditional methods. Base editing technology has begun to enter the toolbox of breeders, especially when it is used in combination with genomic selection and high-throughput phenotypic analysis, the effect is more obvious. In fact, breeders have been constantly seeking methods that can precisely introduce target traits. Now, through genomic prediction combined with base editing, it is possible to screen out plants with the ideal combination of traits more quickly. High-throughput phenotypic analysis systems are not just for show. They can quickly test whether the edited plants have truly developed herbicide resistance and also detect other agronomic traits simultaneously (Andorf et al., 2019). Ultimately, this combination of technologies can significantly shorten the breeding time and make it easier to find truly useful resistant varieties.
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