Bt Research 2025, Vol.16, No.4, 157-167 http://microbescipublisher.com/index.php/bt 157 Research Report Open Access Engineering Bt Strains Using Synthetic Biology Approaches to Enhance Efficacy Xiazhen Huang1, Kaiwen Liang 2 1 Tropical Medicinal Plant Research Center, Hainan Institute of Tropical Agricultural Resources, Sanya, 572025, Hainan, China 2 Agri-Products Application Center, Hainan Institute of Tropical Agricultural Resouces, Sanya, 572025, Hainan, China Corresponding author: kaiwen.liang@hitar.org Bt Research, 2025, Vol.16, No.4 doi: 10.5376/bt.2025.16.0019 Received: 10 Jun., 2025 Accepted: 15 Jul., 2025 Published: 08 Aug., 2025 Copyright © 2025 Huang and Liang, This is an open access article published under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. Preferred citation for this article: Huang X.Z., and Liang K.W., 2025, Engineering Bt strains using synthetic biology approaches to enhance efficacy, Bt Research, 16(4): 157-167 (doi: 10.5376/bt.2025.16.0019) Abstract Bacillus thuringiensis is widely used in crop pest control. This study focuses on reviewing Bt modification strategies driven by synthetic biology, including the design of synthetic novel Cry toxin genes, modular assembly and expression of multigene clusters, and metabolic pathway optimization to enhance the stability and toxin production of Bt strains. Through example analysis of the synthetic design of newCry toxin genes, the gene circuit improves the efficiency of toxin expression, and the field application effect of engineered Bt strains in resistant pest control, the successful practice of synthetic biology is demonstrated. In addition, auxiliary strategies for improving Bt efficacy are discussed, such as combining with RNA interference (RNAi) technology, synergistic with symbiotic bacteria or plant endophytes, and complementary applications with chemical pesticides. Discussions on the ecological safety and risk assessment of transgenic Bt strains, analyzing the impact on non-target organisms and soil microbial communities, as well as the challenges facing synthetic biology in agricultural biosafety regulation. This study aims to show that synthetic biology provides new ideas and means for improving the effectiveness of Bt strains, which is of great significance to the future development of biopesticides. Keywords Bacillus thuringiensis; Insecticidal toxins; Synthetic biology; Genetic engineering; Pest resistance 1 Introduction Bacillus thuringiensis is a soil Bacillus that produces toxin crystal proteins. Its preparations and toxin proteins are widely used in the biological control of agricultural and forestry pests. It is one of the most successfully used and widely used microbial insecticides. Bt preparations are specifically insecticidal and safe for humans and animals, and have significantly reduced the use of chemical pesticides and are environmentally friendly. For example, about 75% of agroforestry pest control in North America relies on spraying Bt bacteria agents (Chakrabarty et al., 2022). In addition, the Bt gene-transformed insect-resistant crops (such as cotton, corn) created by translocating the Bt toxin gene into crops has significantly reduced the harm of pests worldwide since its promotion in the late 1990s (Sansinenea et al., 2010). With the large-scale and long-term application of Bt, some problems have gradually become prominent. The engineering transformation of traditional Bt strains mainly relies on natural strain screening or random mutagenesis, and it is difficult to break through the limitations of the strain's own virulence spectrum. Many Bt biopesticides have limitations such as short field duration and insufficient insecticidal activity for certain pests, and require frequent application. Pest populations are prone to drug resistance under continuous selection pressure. Traditional Bt strain improvement methods show limitations in improving insecticidal spectrum and overcoming resistance. In recent years, the rise of synthetic biology has provided new ideas and tools for the directional transformation of Bt strains. On the one hand, synthetic biology advocates the design of biological systems with an engineering concept, which can modularly transform the gene elements and metabolic pathways of Bt strains (Cheliah et al., 2019). On the other hand, synthetic biology combined with high-throughput sequencing and systematic biological data has given us a deeper understanding of the mechanism of action of Bt toxins and the mechanism of pest resistance, providing a basis for rational transformation (Shikov et al., 2024). Based on the
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