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A highly selective cell-based fluorescent biosensor for genistein detection

Engineering Microbiology 2023年第2期3卷 72-82页

作者： Lucy Fang-I Chao dany liu Verena Siewers Department of Life Sciences Division of Systems and Synthetic BiologyChalmers University of TechnologyGothenburg SE-41296Sweden

Genistein,an isoflavone found mainly in legumes,has been shown to have numerous health benefits for ***,there is substantial interest in producing it using microbial cell *** aid in screening for high genistein producing microbial strains,a cell-based biosensor for genistein was developed by repurposing the Gal4DBD-ER𝛼-VP16(GEV)transcriptional activator in Saccharomyces *** the presence of genistein,the GEV sensor protein binds to the GAL1 promoter and activates transcription of a downstream GFP *** performance of the biosensor,as measured by fold difference in GFP signal intensity after external genistein induction,was improved by engineering the sensor protein,its promoter and the reporter ***-sor performance increased when the weak promoter REV1 p was used to drive GEV sensor gene expression and the VP16 transactivating domain on GEV was replaced with the tripartite VPR transactivator that had its NLS *** biosensor performance further improved when the binding sites for the inhibitor Mig1 were removed from and two additional Gal4p binding sites were added to the reporter *** genistein induction,our improved biosensor output a GFP signal that was 20 times higher compared to the uninduced *** of the 8 flavonoids tested,the improved biosensor responded only to genistein and in a somewhat linear *** improved biosensor also responded to genistein produced in vivo,with the GFP reporter intensity directly propor-tional to intracellular genistein *** combined with fluorescence-based cell sorting technology,this biosensor could facilitate high-throughput screening of a genistein-producing yeast cell factory.

关键词： Biosensor Genistein Flavonoid Isoflavone

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