Bt Research 2025, Vol.16, No.5, 194-203 http://microbescipublisher.com/index.php/bt 200 regulate the expression of defense genes in the plant body, but may also exudate the Bt community attached to it (Dezhabad et al., 2018). Research points out that plant-derived salicylic acid may inhibit the movement and colonization-related gene expression of certain Bts, thereby limiting their overproliferation. Figure 2 Schematic representation of the crossing of two Bt genotypes with two non-Bt genotypes in a reciprocal manner and their F1 hybrids (Adopted from Noor et al., 2022) In a two-biological interaction study, researchers provided Bt-t-treated corn root secretions to Bt cultures and found that the reduced sugar contained in it significantly promoted Bt growth and induced upregulation of NprR-related genes in the population sensing system, suggesting that Bt might "sniff" the abundant nutrients of plants and accelerate colonization and spore formation. In addition, the plant surface microenvironment also plays a role in shaping Bt gene networks. Bt activates SOS repair and DNA protection genes under strong UV irradiation, while inhibiting growth and toxin synthesis genes to enter a tolerant state. 7 Application of Gene Editing and Synthetic Biology in The study of Bt Regulatory Network 7.1 Application of CRISPR/Cas in the analysis of Bt gene regulatory elements Traditional Bt genetic modification has always been difficult due to difficulties such as many natural plasmids and low homologous recombination efficiency. The rise of CRISPR/Cas gene editing technology has brought breakthrough progress to the research of Bt regulatory networks. Using the CRISPR/Cas9 system, researchers can efficiently knock out, insert or point mutations at specific sites in the Bt genome to verify the function of a certain regulatory gene or modify regulatory elements to observe network responses. A typical application is knockout analysis of global regulatory genes. Taking the RNA chaperone Hfq as an example, it was very cumbersome to construct hfq mutant strains in the past by using conventional methods. With the help of CRISPR/Cas9, Bt hfq single, double and triple knockout strains were quickly constructed and the phenotypes were systematically compared. The results greatly deepened the understanding of the role of Hfq in the global regulation of Bt, such as multiple effects such as movement, spore cytosis and virulence reduction (Wang et al., 2020; Zhao et al., 2020). 7.2 Case analysis: research on enhanced Bt toxin yield through gene editing Gene editing technology is not only used to analyze network functions, but also used to modify Bt strains to improve their useful performance. In this regard, enhancing Bt toxin yield and insecticidal activity is one of the primary goals. Researchers have tried a variety of gene editing strategies to achieve this goal, including deleting negative regulatory factors, increasing toxin gene copy, and modifying metabolic pathways. A successful case is to improve the strain UV tolerance and field effectiveness by knocking out the hmgAgene in Bt. CRISPR/Cas9 was used to accurately knock out hmgA in the industrial high-virulence Bt HD-1 strain. This gene encodes a homogeneous Ketoate dioxygenase. After knockout, the strain accumulated a large amount of melanin.
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