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Tuning Cyborg Bacteria through Poly(ethylene glycol) variants with distinct structural and molecular weight profiles. A) Flow cytometry analysis of reductive chromogenic dye and fluorescein-PEGDA positive populations of 4arm-PEGDA coupled with I2959 and LAP65 photoinitiators (Q2 = fluorescein-PEGDA and reductive chromogenic dye positive population, n = 2 biological replicates with 2 technical replicates within each biological replicate). B) (Top panels) Geometric mean and percentage of fluorescein-PEGDA positive Cyborg Bacterial populations. (Bottom panels) Geometric mean and percentage of RCD positive Cyborg Bacterial populations. Significantly reduced mean and percent hydrogelation efficiency was observed between 4arm-PEG/I2959 and 4arm-PEG/LAP65 tuned Cyborg EcN populations, yet both groups still showed >90 % hydrogelation yield. Percent and mean metabolic activities were not significantly different between Cyborg EcN tuned with 4arm-PEG/LAP65 and WT EcN, while mean metabolic activity of 4arm-PEG/I2959 was significantly higher than that of WT EcN cells (error bar = SD, n = 2 biological replicates with 2 technical replicates within each biological replicate). C) CFU of sorted and non-sorted Cyborg <t>E.</t> <t>coli</t> Nissle cells, demonstrating non detectable colony counts in sorted Cyborg Bacterial populations (1.5M sorted Cyborg and WT cells, S = sorted, NS = non-sorted, detection limit at 10 3 ). D) Flow cytometry analysis of reductive chromogenic dye and fluorescein-PEGDA positive populations of M n 250 and 575 2arm-PEGDA coupled with I2959 and LAP65 photoinitiators (Q2 = fluorescein-PEGDA and reductive chromogenic dye positive population, n = 2 biological replicates with 2 technical replicates within each biological replicate). E) (Top panels) Geometric mean and percentage of fluorescein-PEGDA positive Cyborg Bacterial populations. (Bottom panels) Geometric mean and percentage of RCD-positive Cyborg Bacterial populations. Results indicate no significant difference in percent positive fluorescein diacrylate bacteria in M n 250 and 575 2arm-PEGDA Cyborg Bacteria with only slight increase (6.22 %) in percent positive RCD bacteria in 575 2arm-PEGDA with I2959 compared to LAP65. There was a notable increase in hydrogelation efficiency in bacteria with M n 575 2arm-PEGDA compared to M n 250 2arm-PEGDA in both I2959 and LAP65 (error bar = SD, n = 2 biological replicates with 2 technical replicates within each biological replicate). F) CFU of sorted and non-sorted Cyborg EcN, demonstrating three-log-fold reduction in colony counts between WT and sorted Cyborg Bacteria (error bar = SD, n = 2 technical replicates, 1.5M sorted Cyborg and WT cells, S = sorted, NS = non-sorted, detection limit at 10 3 ).
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Tuning Cyborg Bacteria through Poly(ethylene glycol) variants with distinct structural and molecular weight profiles. A) Flow cytometry analysis of reductive chromogenic dye and fluorescein-PEGDA positive populations of 4arm-PEGDA coupled with I2959 and LAP65 photoinitiators (Q2 = fluorescein-PEGDA and reductive chromogenic dye positive population, n = 2 biological replicates with 2 technical replicates within each biological replicate). B) (Top panels) Geometric mean and percentage of fluorescein-PEGDA positive Cyborg Bacterial populations. (Bottom panels) Geometric mean and percentage of RCD positive Cyborg Bacterial populations. Significantly reduced mean and percent hydrogelation efficiency was observed between 4arm-PEG/I2959 and 4arm-PEG/LAP65 tuned Cyborg EcN populations, yet both groups still showed >90 % hydrogelation yield. Percent and mean metabolic activities were not significantly different between Cyborg EcN tuned with 4arm-PEG/LAP65 and WT EcN, while mean metabolic activity of 4arm-PEG/I2959 was significantly higher than that of WT EcN cells (error bar = SD, n = 2 biological replicates with 2 technical replicates within each biological replicate). C) CFU of sorted and non-sorted Cyborg <t>E.</t> <t>coli</t> Nissle cells, demonstrating non detectable colony counts in sorted Cyborg Bacterial populations (1.5M sorted Cyborg and WT cells, S = sorted, NS = non-sorted, detection limit at 10 3 ). D) Flow cytometry analysis of reductive chromogenic dye and fluorescein-PEGDA positive populations of M n 250 and 575 2arm-PEGDA coupled with I2959 and LAP65 photoinitiators (Q2 = fluorescein-PEGDA and reductive chromogenic dye positive population, n = 2 biological replicates with 2 technical replicates within each biological replicate). E) (Top panels) Geometric mean and percentage of fluorescein-PEGDA positive Cyborg Bacterial populations. (Bottom panels) Geometric mean and percentage of RCD-positive Cyborg Bacterial populations. Results indicate no significant difference in percent positive fluorescein diacrylate bacteria in M n 250 and 575 2arm-PEGDA Cyborg Bacteria with only slight increase (6.22 %) in percent positive RCD bacteria in 575 2arm-PEGDA with I2959 compared to LAP65. There was a notable increase in hydrogelation efficiency in bacteria with M n 575 2arm-PEGDA compared to M n 250 2arm-PEGDA in both I2959 and LAP65 (error bar = SD, n = 2 biological replicates with 2 technical replicates within each biological replicate). F) CFU of sorted and non-sorted Cyborg EcN, demonstrating three-log-fold reduction in colony counts between WT and sorted Cyborg Bacteria (error bar = SD, n = 2 technical replicates, 1.5M sorted Cyborg and WT cells, S = sorted, NS = non-sorted, detection limit at 10 3 ).
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Tuning Cyborg Bacteria through Poly(ethylene glycol) variants with distinct structural and molecular weight profiles. A) Flow cytometry analysis of reductive chromogenic dye and fluorescein-PEGDA positive populations of 4arm-PEGDA coupled with I2959 and LAP65 photoinitiators (Q2 = fluorescein-PEGDA and reductive chromogenic dye positive population, n = 2 biological replicates with 2 technical replicates within each biological replicate). B) (Top panels) Geometric mean and percentage of fluorescein-PEGDA positive Cyborg Bacterial populations. (Bottom panels) Geometric mean and percentage of RCD positive Cyborg Bacterial populations. Significantly reduced mean and percent hydrogelation efficiency was observed between 4arm-PEG/I2959 and 4arm-PEG/LAP65 tuned Cyborg EcN populations, yet both groups still showed >90 % hydrogelation yield. Percent and mean metabolic activities were not significantly different between Cyborg EcN tuned with 4arm-PEG/LAP65 and WT EcN, while mean metabolic activity of 4arm-PEG/I2959 was significantly higher than that of WT EcN cells (error bar = SD, n = 2 biological replicates with 2 technical replicates within each biological replicate). C) CFU of sorted and non-sorted Cyborg E. coli Nissle cells, demonstrating non detectable colony counts in sorted Cyborg Bacterial populations (1.5M sorted Cyborg and WT cells, S = sorted, NS = non-sorted, detection limit at 10 3 ). D) Flow cytometry analysis of reductive chromogenic dye and fluorescein-PEGDA positive populations of M n 250 and 575 2arm-PEGDA coupled with I2959 and LAP65 photoinitiators (Q2 = fluorescein-PEGDA and reductive chromogenic dye positive population, n = 2 biological replicates with 2 technical replicates within each biological replicate). E) (Top panels) Geometric mean and percentage of fluorescein-PEGDA positive Cyborg Bacterial populations. (Bottom panels) Geometric mean and percentage of RCD-positive Cyborg Bacterial populations. Results indicate no significant difference in percent positive fluorescein diacrylate bacteria in M n 250 and 575 2arm-PEGDA Cyborg Bacteria with only slight increase (6.22 %) in percent positive RCD bacteria in 575 2arm-PEGDA with I2959 compared to LAP65. There was a notable increase in hydrogelation efficiency in bacteria with M n 575 2arm-PEGDA compared to M n 250 2arm-PEGDA in both I2959 and LAP65 (error bar = SD, n = 2 biological replicates with 2 technical replicates within each biological replicate). F) CFU of sorted and non-sorted Cyborg EcN, demonstrating three-log-fold reduction in colony counts between WT and sorted Cyborg Bacteria (error bar = SD, n = 2 technical replicates, 1.5M sorted Cyborg and WT cells, S = sorted, NS = non-sorted, detection limit at 10 3 ).

Journal: Materials Today Bio

Article Title: Architectural engineering of Cyborg Bacteria with intracellular hydrogel ☆

doi: 10.1016/j.mtbio.2024.101226

Figure Lengend Snippet: Tuning Cyborg Bacteria through Poly(ethylene glycol) variants with distinct structural and molecular weight profiles. A) Flow cytometry analysis of reductive chromogenic dye and fluorescein-PEGDA positive populations of 4arm-PEGDA coupled with I2959 and LAP65 photoinitiators (Q2 = fluorescein-PEGDA and reductive chromogenic dye positive population, n = 2 biological replicates with 2 technical replicates within each biological replicate). B) (Top panels) Geometric mean and percentage of fluorescein-PEGDA positive Cyborg Bacterial populations. (Bottom panels) Geometric mean and percentage of RCD positive Cyborg Bacterial populations. Significantly reduced mean and percent hydrogelation efficiency was observed between 4arm-PEG/I2959 and 4arm-PEG/LAP65 tuned Cyborg EcN populations, yet both groups still showed >90 % hydrogelation yield. Percent and mean metabolic activities were not significantly different between Cyborg EcN tuned with 4arm-PEG/LAP65 and WT EcN, while mean metabolic activity of 4arm-PEG/I2959 was significantly higher than that of WT EcN cells (error bar = SD, n = 2 biological replicates with 2 technical replicates within each biological replicate). C) CFU of sorted and non-sorted Cyborg E. coli Nissle cells, demonstrating non detectable colony counts in sorted Cyborg Bacterial populations (1.5M sorted Cyborg and WT cells, S = sorted, NS = non-sorted, detection limit at 10 3 ). D) Flow cytometry analysis of reductive chromogenic dye and fluorescein-PEGDA positive populations of M n 250 and 575 2arm-PEGDA coupled with I2959 and LAP65 photoinitiators (Q2 = fluorescein-PEGDA and reductive chromogenic dye positive population, n = 2 biological replicates with 2 technical replicates within each biological replicate). E) (Top panels) Geometric mean and percentage of fluorescein-PEGDA positive Cyborg Bacterial populations. (Bottom panels) Geometric mean and percentage of RCD-positive Cyborg Bacterial populations. Results indicate no significant difference in percent positive fluorescein diacrylate bacteria in M n 250 and 575 2arm-PEGDA Cyborg Bacteria with only slight increase (6.22 %) in percent positive RCD bacteria in 575 2arm-PEGDA with I2959 compared to LAP65. There was a notable increase in hydrogelation efficiency in bacteria with M n 575 2arm-PEGDA compared to M n 250 2arm-PEGDA in both I2959 and LAP65 (error bar = SD, n = 2 biological replicates with 2 technical replicates within each biological replicate). F) CFU of sorted and non-sorted Cyborg EcN, demonstrating three-log-fold reduction in colony counts between WT and sorted Cyborg Bacteria (error bar = SD, n = 2 technical replicates, 1.5M sorted Cyborg and WT cells, S = sorted, NS = non-sorted, detection limit at 10 3 ).

Article Snippet: E. coli Top-10 cells (ThermoFisher) were used primarily for plasmid cloning and maintenance, as well as a diverse host for dsDNA-PEG hydrogel formation.

Techniques: Bacteria, Molecular Weight, Flow Cytometry, Activity Assay