Maize Genomics and Genetics 2025, Vol.16, No.4, 229-238 http://cropscipublisher.com/index.php/mgg 233 is quite good, with the highest editing efficiency within the body reaching up to 13.8%. It is worth mentioning that this method can also directly obtain homozygous mutants without exogenous DNA. These plants only carry one or two point mutations on the ALS gene. 5.3 Outcomes in resistance phenotype and agronomic performance Later observations revealed that these plants with the ZmALS1 mutation, or those with both ZmALS1 and ZmALS2 mutations, demonstrated remarkable resistance when confronted with the sulfonylurea herbicide chlorosulfonon. Even when the dosage is increased to 15 times the usual field dosage, they can still withstand it. Moreover, these mutated corns grew normally without showing any growth or development issues, which indicates that the introduced mutations did not affect their agronomic traits. This research shows that base editing technology can indeed be used to precisely breed herbicide-resistant corn varieties, and it is also a non-GMO route, which is quite meaningful for the future direction of breeding. Figure 2 Targeted base editing in two non-allelic ALS genes, ZmALS1 and ZmALS2, conferred high-dose chlorsulfuron resistance in maize plants. (A) Sequencing profiles identified a T3 line, 54-42-34-26, with homologous thymine mutations in both ZmALS1 and ZmALS2 at target site 7. Blue arrows indicate the T/G SNP discriminating ZmALS1 and ZmALS2; red arrows indicate the induced target mutation of cytosine to thymine. (B) Three chlorsulfuron concentration level treatments in both ZmALS1 and ZmALS2 targeted CT base edited mutants (on left in pods) and wild-type control (on right in pods) of ZC01. Photographs were made 30 days after herbicide (chlorsulfuron) treatments. Treatments with 500 mL of 0 (empty control), 100, 200, and 300 mg/L chlorsulfuron were applied in each pot of this test. Each pot contained 6 mutant plants on the left and 6 WT plants on the right. 5×, 10×, and 15×indicate the received chlorsulfuron dosage for plants: 5, 10, and 15 times the recommended field application dose, respectively. Scale bar, 10 cm (Adopted from Li et al., 2020) 6 Advantages and Potential of Base Editing in Herbicide Resistance Breeding 6.1 Precision and predictability of trait introduction Not all herbicide-resistant breeding requires breaking DNA strands. Base editing technology can actually directly modify a single base at a specific position without cutting DNA or requiring a donor template. In this way, herbicide-resistant mutations can be introduced more quickly and accurately. Such changes are very small and look almost no different from natural mutations, and the risk of off-target is also lower. At present, crops such as corn, wheat, soybeans and rapeseed have developed some herbicide-resistant varieties that can be inherited without genetically modified labels through this method (Dong et al., 2021; Shi et al., 2023). In addition, with the addition of specific allele markers, the screening process becomes more convenient (Wu et al., 2020; Wei et al., 2023). 6.2 Shorter breeding cycles compared to conventional methods In the past, breeding for herbicide resistance relied on mutagenesis or backcrossing generation after generation, and slowness was the norm. Now it's different. With base editing, point mutations can be directly applied to superior varieties, and the speed is very fast. It doesn't take too many generations to obtain homozygous mutants without genetically modified components. This approach significantly saves time and is also easier to promote (Shimatani et al., 2017; Zhang et al., 2019b).
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