bl21 de3 competent e coli cells  (New England Biolabs)

Journal: iScience

Article Title: Engineering electron conduits in bacteria for selective biointerfacing and enhanced energy transfer

doi: 10.1016/j.isci.2026.114805

Figure Lengend Snippet: Functional modification of MtrC with a SpyTag bioconjugation domain in S. oneidensis and E. coli (A) Fusion of a SpyTag domain to the C-terminus of MtrC enables the bioconjugation of a SpyCatcher-fused fluorescent protein, facilitating selective surface labeling of modified cells. (B) The fluorescence of S. oneidensis cells was quantified by spectrofluorometry (arbitrary units) and shown as the mean of 3 separate experiments performed in triplicate with subtracted background fluorescence of unreacted control cells. Error bars represent standard deviation between experiments ( n = 3, significance calculated using a parametric, unpaired, one-tailed t test). Expression of MtrC or MtrC-SpyTag in S. oneidensis cells was regulated using either a weak or strong RBS. (C) The fluorescence of E. coli cells expressing CymA-MtrCAB or CymA-MtrCAB-SpyTag shown as the mean of 3 separate experiments performed in triplicate, with subtracted background fluorescence of unreacted control cells. Error bars represent standard deviation between experiments ( n = 3, significance calculated using a parametric, unpaired, one-tailed t test). (D) Fluorescence microscopy revealed observable mCerulean3 fluorescence for E. coli cells expressing CymA-MtrCAB-SpyTag compared with cells expressing CymA-MtrCAB following incubation with mCerulean3-SpyCatcher. Scale bars, 100 μm.

Article Snippet: E. coli BL21 (DE3) , NEB , Cat#C2527.

Techniques: Functional Assay, Modification, Labeling, Fluorescence, Control, Standard Deviation, One-tailed Test, Expressing, Microscopy, Incubation

Journal: iScience

Article Title: Engineering electron conduits in bacteria for selective biointerfacing and enhanced energy transfer

doi: 10.1016/j.isci.2026.114805

Figure Lengend Snippet: Extracellular substrate reduction by S. oneidensis and E. coli cells that express engineered MtrC proteins (A) Reduction of iron(III) to iron(II) by wild-type S. oneidensis , compared to a Δ mtrC /Δ mtrF /Δ omcA knockout strain. The Δ mtrC /Δ mtrF /Δ omcA knockout strain was subsequently engineered to express MtrC or MtrC-SpyTag. The formation of iron(II) after 16 h of anaerobic culture was quantified using a ferrozine assay and shown as a mean of three experiments with three biological repeats. Error bars represent standard deviation between experiments ( n = 3, significance calculated using a parametric, unpaired, two-tailed t test). (B) Reduction of methyl orange with electrons from E. coli BL21(DE3) cells containing only the pEC86 plasmid, or cells co-expressing either the CymA-MtrCAB complex, CymA-MtrCAB with SpyTag, or CymA-MtrCAB with GrBP5. The change in methyl orange absorbance at 465 nm upon the reduction of the azo group was measured after 24 h of anaerobic culture, and is shown as a mean of three experiments with three biological repeats ( n = 3, significance calculated using a parametric, unpaired, two-tailed t test). Error bars represent the standard deviation between experiments.

Article Snippet: E. coli BL21 (DE3) , NEB , Cat#C2527.

Techniques: Knock-Out, Ferrozine Assay, Standard Deviation, Two Tailed Test, Plasmid Preparation, Expressing

Journal: iScience

Article Title: Engineering electron conduits in bacteria for selective biointerfacing and enhanced energy transfer

doi: 10.1016/j.isci.2026.114805

Figure Lengend Snippet: Improved graphite binding by S. oneidensis and E. coli cells expressing MtrC-GrBP5 (A) Illustration shows the C-terminal modification of MtrC with a graphite binding domain to improve the affinity of S. oneidensis cells to a graphite electrode. (B) SEM images of graphite felt fibers with no cells, as well as with bound S. oneidensis Δ mtrC /Δ mtrF /Δ omcA cells expressing either MtrC or MtrC-GrBP5 (scale bars, 5 μm). (C) SEM images of graphite felt fibers with no cells, as well as with bound E. coli BL21(DE3) expressing either CymA-MtrCAB or CymA-MtrCAB-GrBP5 (scale bars, 5 μm). (D) Lysed protein content from a graphite felt electrode aerobically incubated with S. oneidensis or E. coli cells expressing MtrC with or without the GrBP5 binding domain. The data represent the mean protein per cm 2 of geometric surface area of graphite felt electrode (mg/cm 2 ) in cell lysate from three biological replicates from three separate experiments of S. oneidensis Δ mtrC /Δ mtrF /Δ omcA cells expressing recombinant MtrC or MtrC-GrBP5, or of E. coli BL21(DE3) cells expressing CymA-MtrCAB-GrBP5 compared to CymA-MtrCAB. Error bars represent standard deviation between experiments ( n = 3, significance calculated using a parametric, unpaired, one-tailed t test).

Article Snippet: E. coli BL21 (DE3) , NEB , Cat#C2527.

Techniques: Binding Assay, Expressing, Modification, Incubation, Recombinant, Standard Deviation, One-tailed Test

Journal: bioRxiv

Article Title: Large-scale exploration of protein space by automated NMR

doi: 10.64898/2026.02.16.706194

Figure Lengend Snippet: Design pipeline (top); de novo backbones were generated using either RFdiffusion, or Proteína, followed by sequence design with ProteinMPNN and filtering with Boltz-1. Experimental validation (bottom); designs were ordered as synthetic genes, cloned into expression vectors, and produced in E. coli from 15 N-labelled autoinduction media in 96-deepwell plates (left). Samples were purified by immobilized metal affinity chromatography in 96-well plate format, followed by high-throughput size-exclusion chromatography (middle). NMR fingerprints are obtained in 45 min for all produced samples.

Article Snippet: Assembled plasmids were transformed into lab-made chemically competent BL21(DE3) E. coli cells (NEB #C2527) obtained with an adapted protocol from Yang et al. To each 1 μL Golden Gate reaction, 8 μL of ice-cold KCM (500 mM KCl, 250 mM MgCl 2 , 150 mM CaCl 2 ) and 8 μL of ice-cold competent cells were added, followed by incubation on ice for 30 minutes.

Techniques: Generated, Sequencing, Biomarker Discovery, Clone Assay, Expressing, Produced, Purification, Affinity Chromatography, High Throughput Screening Assay, Size-exclusion Chromatography