Strategy of employing plug-and-play vectors and LC-MS screening to facilitate the discovery of natural products using Aspergillus oryzae

Hanliang Shi; Beibei Lin; Mengmeng Zheng; Fengyu Gan; Zhi Lin; Xiujuan Xin; Jian Zhao; Xudong Qu; Faliang An

doi:10.1186/s40643-024-00833-w

Strategy of employing plug-and-play vectors and LC-MS screening to facilitate the discovery of natural products using Aspergillus oryzae

Bioresour Bioprocess. 2025 Jan 8;12(1):2. doi: 10.1186/s40643-024-00833-w.

Authors

Hanliang Shi¹, Beibei Lin¹, Mengmeng Zheng^{2

3

4}, Fengyu Gan¹, Zhi Lin^{2

3

4}, Xiujuan Xin¹, Jian Zhao^{1

5}, Xudong Qu^{2

3

4}, Faliang An^{6

7}

Affiliations

¹ State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, 130 Mei Long Road, Shanghai, 200237, China.
² State Key Laboratory of Microbial Metabolism, School of Life Sciences and Biotechnology, Shanghai Jiao Tong University, Shanghai, 200240, China.
³ Key Laboratory of Combinatorial Biosynthesis and Drug Discovery Ministry of Education, School of Pharmaceutical Sciences, Wuhan University, Wuhan, 430071, China.
⁴ Zhangjiang Institute for Advanced Study, Shanghai Jiao Tong University, Shanghai, 201203, China.
⁵ Department of Applied Biology, East China University of Science and Technology, 130 Mei Long Road, Shanghai, 200237, China.
⁶ State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, 130 Mei Long Road, Shanghai, 200237, China. flan2016@ecust.edu.cn.
⁷ Marine Biomedical Science and Technology Innovation Platform of Lin-Gang Special Area, No. 4, Lane 218, Haiji Sixth Road, Shanghai, 201306, China. flan2016@ecust.edu.cn.

PMID: 39776300
DOI: 10.1186/s40643-024-00833-w

Abstract

Aspergillus oryzae is a widely used host for heterologous expression of fungal natural products. However, the vectors previously developed are not convenient for use and screening positive transformants by PCR and fermentation is time- and effort-consuming. Hence, three plug-and-play vectors were developed here for multi-gene expression and liquid chromatography mass spectrometry detection was introduced to screen positive transformants. Using rug BGC for verification, we demonstrated that the vectors we developed perform well and liquid chromatography mass spectrometry detection is feasible to screen positive transformants. For deleterious gene expression, PxyrA rather than PamyB was employed. Utilizing the toolkit described here to express natural products, dozen days can be saved.

Keywords: Aspergillus oryzae; Heterologous expression; LC–MS detection; PamyB; Plug-and-play vectors.

Grants and funding

2023YFA0914102/National Key R&D Program of China