Molecular Microbiology Research, 2025, Vol.15, No.1, 1-9 http://microbescipublisher.com/index.php/mmr 7 improvement tool worth referring to. In addition to cucumber itself, it may actually be used for the production of vaccines or certain biological materials, and there is a lot of room for expansion. Of course, this is not the only success story. The cucumber variety "Xintai Mischi" in northern China has also made progress. This time it was replaced by the GV3101 strain, and the researchers mainly optimized the use of antibiotics and ferulic acid ketone, and finally achieved an 8.1% conversion efficiency (Chai et al., 2020a; 2020b). Although not as high as Poinsett 76, this is already one of the best levels that can be found in the "New Tai Misty" variety. More importantly, this method also shows good application potential in improving stress resistance and improving nutritional quality, and is easier to implement in actual cultivation. 7.2 Comparative analysis of transformation techniques There have been many "versions" of cucumber gene transformation technology in recent years. Once there are many methods, the effects will naturally be high and low. The most commonly used old method is to use the EHA105 strain combined with cotyledon explants. This combination is most widely used because of its stable transformation efficiency (Wang et al., 2015). But that doesn't mean other methods are not worth a try. Such as vacuum penetration combined with filter paper core co-culture, it sounds not traditional, but it really improves the conversion efficiency. On the one hand, it can enhance the infection effect of Agrobacterium rhizogenes, and on the other hand, it also reduces explant necrosis, with an average conversion rate of 11.9% (Nanasato et al., 2012). This idea obviously solves the infection bottleneck more "treat" with it. There is also a more "quick knife" approach, which is to use Agrobacterium rhizogenes to generate a complex plant with a genetically modified root, a stem or a wild type. This single-step method saves a lot of trouble. The success rate of transgenic roots can exceed 90%, which is particularly convenient for functional gene analysis or studying root traits (Fan et al., 2020). As for CRISPR/Cas9, it is still in the exploratory stage on cucumbers. Its efficiency is not as good as the traditional method for the time being, but its advantage is that it is more directional. For example, by regulating the penetration intensity, researchers have made plants with shorter internodes and more compact structures, which shows that this technology does have some potential in the cucurbitae (Xin et al., 2022). 8 Concluding Remarks In plant genetic engineering, the transformation system of Agrobacterium rhizogenes can be said to be unavoidable, especially when studying cucumber plants, it is used very much. But its advantages are not outstanding from the beginning. Many improvements have actually been accumulated slowly in recent years. For example, the transient transformation method was not favored by many people in the early stage, but later because it was fast, step-saving, and easy to scale, it has now been widely used in the research of functional genomes and recombinant proteins. In terms of efficiency, traditional transformation has not been improved, especially after choosing the right genotype, explant and strain, some solutions can even achieve a conversion rate of 23% - on crops like cucumbers, this achievement is not low. In addition, there is a one-step approach that has also attracted attention: directly using the root strain bacteria to generate a composite plant of genetically modified roots and wild-type stems, which does not require staged processing, the process is simple, and the effect is quite stable. In recent years, technical details like this have been continuously optimized, making Agrobacterium rhizogenes more and more like a "standard configuration in the toolbox" in crop improvement and genetic research. Although transformation technology has made a lot in recent years, the problem has not been solved. Some old problems are stuck there. For example, some cucumber varieties cannot improve their efficiency no matter how they are done - this is directly related to the genotype. In other words, it is not a general solution that can handle all the materials, and targeted optimization still needs to be done. Another direction that is attracting great attention now is combining CRISPR/Cas9 with Agrobacterium rhizogenes transformation technology. Although traditional methods are stable, their accuracy is limited; while CRISPR can be edited in a fixed location. If used in conjunction, it may better control the conversion effect and reduce some biosafety concerns. But then again, whether the gene can be inserted stably is not enough. It also needs to figure out where it lands in the genome, whether it causes mutations or large-scale rearrangements. If you don’t understand this kind of
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