Journal: Advanced Science

Article Title: START: A Versatile Platform for Bacterial Ligand Sensing with Programmable Performances

doi: 10.1002/advs.202402029

Figure Lengend Snippet: Design and characterization of STARTs for tetracycline and MS2 coat protein. a) Detailed schematics of the tetracycline Apta‐trigger, Tetra_B3. The lower stem region of tetracycline aptamer subject to engineering is marked by a gray box. b) GFP fluorescence of toehold switch B and trigger RNA pairs in the absence (gray bars) and presence (green bars) of 12 µм tetracycline. Positive control (PC) used the toehold switch trigger B without the aptamer sequence, while negative control (NC) used the decoy RNA without any predicted interactions. c) GFP fluorescence output for switch B and Tetra_B3 pair for different concentrations of tetracycline inputs. d) Detailed schematics of the MS2 coat protein Apta‐trigger, MS2_C1. The lower stem region of MS2 aptamer subject to engineering is marked by a gray box. e) GFP fluorescence of toehold switch C and trigger RNA pairs without (gray bars) and with (green bars) MS2 induction, by 10 ng mL −1 anhydrotetracycline (aTc) treatment. PC used the toehold switch trigger C, and NC used the decoy RNA. f) GFP fluorescence output for switch C and MS2_C1 pair for different induction levels for MS2 expression. P‐values were determined by an unpaired t ‐test, with P > 0.05 designated by “ns”, P ≤ 0.05 designated by “*”, and P ≤ 0.0001 designated by “****”. Error bars are the SD from three biological replicates.

Article Snippet: The MS2 protein coding sequence was PCR amplified using the bacteriophage MS2 coat protein expression plasmid from Addgene (pHMM, #67717) as a template and inserted into a pACYCDuet plasmid with an aTc inducible promoter sequence via Gibson assembly.

Techniques: Fluorescence, Positive Control, Sequencing, Negative Control, Expressing

Journal: Advanced Science

Article Title: START: A Versatile Platform for Bacterial Ligand Sensing with Programmable Performances

doi: 10.1002/advs.202402029

Figure Lengend Snippet: Construction of Boolean logic circuits. a) Design of the two‐input OR gate for theophylline and MS2. b) GFP fluorescence output of the two‐input OR gate for different input combinations. c) Design of the two‐input AND gate for theophylline and MS2. d) GFP fluorescence output of the two‐input AND gate for different input combinations. e) Design of the NOT gate for theophylline using a 3WJ repressor. f) GFP fluorescence of the NOT gate in the absence and presence of theophylline input. Theophylline was treated at a concentration of 10 mм, and aTc was treated at a concentration of 10 ng mL −1 . Error bars represent the SD from three biological replicates.

Article Snippet: The MS2 protein coding sequence was PCR amplified using the bacteriophage MS2 coat protein expression plasmid from Addgene (pHMM, #67717) as a template and inserted into a pACYCDuet plasmid with an aTc inducible promoter sequence via Gibson assembly.

Techniques: Fluorescence, Concentration Assay