## 2 Advancement of BEs in Microbes
### Recent Applications of BEs in Nonmodel Microbes
#### 2.2.2 Agricultureally Important Nonmodel Microbes  
_Paenibacillus polymyxa_ (_P. polymyxa_, formerly known as _Bacillus polymyxa_) is an agricurlurally important microbe as it can directly promote crop growth via nitrogen fixation and phosphate solubilization [93]. It naturally produces a large number of valuable compounds like exopolysaccharides (EPS), 2,3-butanediol, antibiotics, and antimicrobial peptides such as polymyxin and fusaricidin [94]. Since the traditional gene editing methods had a low efficiency and required the integration of a selection marker, the discoveries of the unknown characteristics and physiochemical properties of _P. polymyxa_ need a robust and efficient gene editing method. Recently, after editing activities of different CBEs were compared with _B. subtilis_[56], dCas9-PmCDA1-UGI, which has the best editing efficiency, was also applied in _P. polymyna_. There are six BGCs in the _P. polymyna_ genome and five of them are known to produce antibiotics [56]. After the five known BGCs were inactivated by CBE multiplex gene editing, the uncharacterized polyketide produced by the sixth unknown BGC was evaluated. The establishment of CBE in _B. subtilis_ and _P. polymyna_ would greatly accelerate the discovery of hidden antibiotics and could be further applied in other _Bacillus_ and _Paenibacillus_ species. To the best of our knowledge, fusing Cas9 variants with BEs to recognize more PAM sites has not yet been reported in _Bacillus_ and _Paenibacillus_ species, which can be further explored in order to greatly reduce the number of nonetidable genes. AgrobacteriumumSP.Agrobacteriumum, as a remarkable soil phytopathogen, can achieve a stable transformation using any gene of interest in the plant genome via delivery of transferred (T)-DNA [95]. With the help of a _Agrobacteriumum_-mediated transfer, CRISPR/Cas9 and BEs have been successfully established in plants to modify genomes for both biological functional analysis and crop improvement [96; 97]. However, there has been a lack of characterization of genetic parts in _Agrobacteriumum_ and poor exploration of its metabolic and physiological functions [98]. To improve the transformation of _Agrobacteriumum_ in plants, which was previously reported to be inefficient [99; 100], understanding and modifying transformation-related genes in _Agrobacteriumum_ are also important. CBE has recently been established in two widely used plant transformation strains, _A. tumefaciens_ and _A. rhizogenes_[74]. When CRISPR/Cas9 was found to be lethal in _A. tumefaciens_[74], the researchers set their sights on CBE, aiming to avoid the lethality caused by DSBs. The construct of dCas9-PmCDA1-UL, by modulating the length of gRNA from 20 nt to 18 nt, was proven to be available for gene editing with at least 98% efficiency in _A. tumefaciens_ and _A. rhizogenes_. Interestingly, the CBE that was driven by the constitutive promoter was probably lethal to _E. coli_ since no correct clones were obtained. After switching to the AS-activated promoter _virB_, the system could be successfully cloned in _E. coli_ and worked well in _Agrobacteriumum_ with or without AS treatment. The inactivation of _rolB_ and _rolC_ was efficiently achieved to confirm their importance in hairy root development. CBE facilitates the understanding and engineering of nature's engineer, _Agrobacteriumum_. Further applications of BEs can be used in the field of plant-microbe interactions that help deep learning and improvement of agricultural plants and microbes for beneficial use. Sinorhizobium melidiotSinorhizobium melidiotS. melidiot is a soil bacterium that can form nitrogen-fixing nodules on the roots of leguminous plants. The varied and rich metabolic capabilities of _S. melidiot_, such as carbohydrate metabolism, allow it to adapt to very different environmental and nutritional conditions in free-living form or as a plant symbiont [101]. Past studies have reported that _S.