Bt Research 2025, Vol.16, No.2, 79-85 http://microbescipublisher.com/index.php/bt 79 Feature Review Open Access High-throughput Screening Technology for Novel Bt Toxins Yinghua Chen, Zhongqi Wu Institute of Life Science, Jiyang College of Zhejiang A&F University, Zhuji, 311800, Zhejiang, China Corresponding author: zhongqi.wu@jicat.org Bt Research, 2025, Vol.16, No.2 doi: 10.5376/bt.2025.16.0010 Received: 23 Feb., 2025 Accepted: 08 Apr., 2025 Published: 25 Apr., 2025 Copyright © 2025 Chen and Wu, 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: Chen Y.H., and Wu Z.Q., 2025, High-throughput screening technology for novel Bt toxins, Bt Research, 16(2): 79-85 (doi: 10.5376/bt.2025.16.0010) Abstract This study evaluated the progress of high-throughput screening (HTS) technology in the discovery of novel Bt toxins, introduced the core principles and main parameters of HTS, and compared it with traditional screening methods. It also discussed the application of genomics and metagenomics, high-throughput bioassays, etc. in the research of Bt toxins The strategies of combining HTS with transcriptomics, proteomics, machine learning, etc. were analyzed, and its potential in the functional research of novel toxins, resistance management and precise pest control was explored. This study aims to provide a theoretical basis and technical reference for the efficient discovery and long-term application of Bt toxins. Keywords High-throughput screening; Bt toxins; Metagenomics; Machine learning; Synthetic biology 1 Introduction The effectiveness of traditional Bt (Bacillus thuringiensis) toxins in pest control is declining as the resistance of agricultural pests becomes increasingly severe. Nowadays, it is necessary to develop new Bt toxins with different mechanisms of action to delay the development of resistance and ensure crop safety. The discovery of new Bt toxins can enrich the types of biopesticides and provide more pest management tools for sustainable agriculture. High-throughput screening (HTS) technology, with its automated equipment and rapid detection methods, can screen the activity of a large number of candidate molecules in a very short time, significantly enhancing the efficiency of discovering new bioactive substances. HTS has been widely applied in drug development and industrial biotechnology. It relies on diverse screening libraries and sensitive detection methods, such as fluorescence, energy transfer, polarization, etc., to effectively find molecules with target activity (Blay et al., 2020; Zeng et al., 2020; Cheng and Li, 2021). In the screening of Bt toxins, HTS is expected to accelerate the discovery and functional verification of new toxins and promote the innovation of pest management strategies. This study summarizes the application progress of high-throughput screening technology in the discovery of novel Bt toxins, analyzes the advantages and disadvantages of current screening strategies, and explores the future directions of technology integration and innovation. This study aims to provide a theoretical basis and technical reference for the efficient development of Bt toxins and the management of pest resistance. 2 Principles of High-throughput Screening (HTS) 2.1 Concept and workflow of HTS in microbial and toxin research High-throughput screening (HTS) is a technology that uses automated equipment and miniaturized detection systems to simultaneously analyze a large number of samples in a short period of time. It first constructs diverse microbial or toxin libraries, then uses an automated system to allocate samples into microplates, then conducts activity screening with sensitive detection methods, and finally identifies candidate molecules or strains with target characteristics through data analysis (Leavell et al., 2020; O’Connor et al., 2024). HTS can rapidly identify functional genes, screen efficient strains or novel active molecules in microbial and toxin research, improving research efficiency and the possibility of discovery (Sarnaik et al., 2020; Zeng et al., 2020; Huberman, 2025). 2.2 Key parameters: sensitivity, throughput, and reproducibility The effect of HTS mainly depends on three key parameters. The first one is sensitivity. The detection method
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