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  • 99
    Thermo Fisher od600
    <t>OD600</t> time course of TOP10-rfp-lys grown in 15 ml tubes upon induction with HSL 100 nM (A) and RFP fluorescence time course in the supernatant (B) . Culture absorbance (A) and supernatant fluorescence (B) of TOP10-rfp-lys induced with HSL 100 nM (blue line). Uninduced TOP10-rfp-lys (red line), TOP10 bearing pHC-RFP induced with HSL 100 nM (green line) or uninduced (black line) are the negative controls. Induction was carried out in the exponential phase at OD600~0.55. Error bars represent the 95% confidence interval of the estimated mean.
    Od600, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 4685 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    Beckman Coulter od600
    <t>OD600</t> time course of TOP10-rfp-lys grown in 15 ml tubes upon induction with HSL 100 nM (A) and RFP fluorescence time course in the supernatant (B) . Culture absorbance (A) and supernatant fluorescence (B) of TOP10-rfp-lys induced with HSL 100 nM (blue line). Uninduced TOP10-rfp-lys (red line), TOP10 bearing pHC-RFP induced with HSL 100 nM (green line) or uninduced (black line) are the negative controls. Induction was carried out in the exponential phase at OD600~0.55. Error bars represent the 95% confidence interval of the estimated mean.
    Od600, supplied by Beckman Coulter, used in various techniques. Bioz Stars score: 99/100, based on 981 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    od600  (4Gene)
    86
    4Gene od600
    <t>OD600</t> time course of TOP10-rfp-lys grown in 15 ml tubes upon induction with HSL 100 nM (A) and RFP fluorescence time course in the supernatant (B) . Culture absorbance (A) and supernatant fluorescence (B) of TOP10-rfp-lys induced with HSL 100 nM (blue line). Uninduced TOP10-rfp-lys (red line), TOP10 bearing pHC-RFP induced with HSL 100 nM (green line) or uninduced (black line) are the negative controls. Induction was carried out in the exponential phase at OD600~0.55. Error bars represent the 95% confidence interval of the estimated mean.
    Od600, supplied by 4Gene, used in various techniques. Bioz Stars score: 86/100, based on 8 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    96
    Hitachi Ltd od600
    <t>OD600</t> time course of TOP10-rfp-lys grown in 15 ml tubes upon induction with HSL 100 nM (A) and RFP fluorescence time course in the supernatant (B) . Culture absorbance (A) and supernatant fluorescence (B) of TOP10-rfp-lys induced with HSL 100 nM (blue line). Uninduced TOP10-rfp-lys (red line), TOP10 bearing pHC-RFP induced with HSL 100 nM (green line) or uninduced (black line) are the negative controls. Induction was carried out in the exponential phase at OD600~0.55. Error bars represent the 95% confidence interval of the estimated mean.
    Od600, supplied by Hitachi Ltd, used in various techniques. Bioz Stars score: 96/100, based on 138 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    96
    Biochrom od600
    Growth of various L. paracasei strains in a chemically defined medium with methionine as the sole sulfur source. The bars and black lines represent the mean and standard deviation of the reached <t>OD600</t> after 25 h of growth, respectively, of three independent biologically repeated measurements. Strains that reached an OD600 of one or more were consider as being able to grow in a medium with methionine as sole sulfur source.
    Od600, supplied by Biochrom, used in various techniques. Bioz Stars score: 96/100, based on 233 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    Eppendorf AG od600
    a Phylogenetic tree illustrating the separation of S. mitis NCTC12261 T ]). b Kinetics of transformation in the S. mitis type strain and strain SK321. Pre-cultures at <t>OD600</t> 0.5 were diluted 1:100 in C + Y YB medium and grown until OD600 0.04 at 37 °C in 5% CO 2 . Cultures were treated with 300 nM cognate CSP and distributed into 200 μL aliquots that were further exposed to 1 μg ml − 1 recombinant plasmid pVA838 at indicated times. 20 U ml − 1 DNase I were added after 30 min of exposure to DNA and the culture was incubated in air at 37 °C for additional 30 min. Transformants were recovered in blood agar plates supplemented with Erythromycin. Each line represents results of a single experiment
    Od600, supplied by Eppendorf AG, used in various techniques. Bioz Stars score: 99/100, based on 901 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    94
    Spectronics od600
    a Phylogenetic tree illustrating the separation of S. mitis NCTC12261 T ]). b Kinetics of transformation in the S. mitis type strain and strain SK321. Pre-cultures at <t>OD600</t> 0.5 were diluted 1:100 in C + Y YB medium and grown until OD600 0.04 at 37 °C in 5% CO 2 . Cultures were treated with 300 nM cognate CSP and distributed into 200 μL aliquots that were further exposed to 1 μg ml − 1 recombinant plasmid pVA838 at indicated times. 20 U ml − 1 DNase I were added after 30 min of exposure to DNA and the culture was incubated in air at 37 °C for additional 30 min. Transformants were recovered in blood agar plates supplemented with Erythromycin. Each line represents results of a single experiment
    Od600, supplied by Spectronics, used in various techniques. Bioz Stars score: 94/100, based on 339 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    93
    Carl Zeiss od600
    a Phylogenetic tree illustrating the separation of S. mitis NCTC12261 T ]). b Kinetics of transformation in the S. mitis type strain and strain SK321. Pre-cultures at <t>OD600</t> 0.5 were diluted 1:100 in C + Y YB medium and grown until OD600 0.04 at 37 °C in 5% CO 2 . Cultures were treated with 300 nM cognate CSP and distributed into 200 μL aliquots that were further exposed to 1 μg ml − 1 recombinant plasmid pVA838 at indicated times. 20 U ml − 1 DNase I were added after 30 min of exposure to DNA and the culture was incubated in air at 37 °C for additional 30 min. Transformants were recovered in blood agar plates supplemented with Erythromycin. Each line represents results of a single experiment
    Od600, supplied by Carl Zeiss, used in various techniques. Bioz Stars score: 93/100, based on 87 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    98
    GE Healthcare od600
    Growth curves of P. fluorescens strains cLP6a and cLP6a-1 . Growth of P. fluorescens strains cLP6a and cLP6a-1 at (a) 10°C, (b) 28°C or (c) 35°C determined as <t>OD600</t> Each data point is the mean of three independent cultures, and error bars, where visible, indicate the standard deviation.
    Od600, supplied by GE Healthcare, used in various techniques. Bioz Stars score: 98/100, based on 1227 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    Molecular Devices LLC od600
    Residues on the flat surface of MreB are critical for CbtA-F65S-mediated cell elongation inhibition. (A) Schematic of MreB complementation assay (panels B and C). Depletion strain FB30/pFB174 ( mreBCD :: kan R /pBAD-mreBCD) [ 9 ] was transformed with either an empty vector control (pMLB1113) or a plasmid derived from pFB149 ( plac-mreBCD ) [ 9 ] expressing wild-type mreB (pFB149), mreB-E262G (pDH279), or mreB-S269F (pDH332). (B) Spot dilution assay to assess the ability of each mreB allele to support growth on solid medium. Cultures of each strain grown overnight at 37°C in M9 maltose (0.2% maltose, 0.2% casamino acids, 1 mM MgSO 4 ) supplemented with 0.5% arabinose were back-diluted 1:100 in fresh M9 maltose + 0.5% arabinose and grown at 37°C for several hours until they reached late log phase. Cultures were pelleted and washed to remove arabinose, normalized to the same <t>OD600</t> value, serially diluted, and spotted on LB plates supplemented with either 0.5% arabinose or 250 μM IPTG. Plates were incubated overnight at 37°C. Dilutions 10 −1 to 10 −5 are shown. (C) Cell morphology phenotypes of complemented strains. Aliquots from the same overnight cultures described in (B) were washed once and resuspended in fresh M9 maltose (without arabinose). These washed aliquots were used as inocula for M9 maltose cultures supplemented with 250 μM IPTG. All cultures were grown at 37°C for several hours (maintained in log-phase by back dilution) and cell morphology was monitored periodically by microscopy. Images taken after 5 h of growth in the presence of 250 μM IPTG are shown. (D) Effect of MreB substitution E262G on cell morphology phenotypes in the presence or absence of overproduced CbtA-F65S. Strains DH118/pFB149 (BW27785 mreBCD :: kan R / plac-mreBCD ) and DH118/pDH278 (BW27785 mreBCD :: kan R / plac-mreB-E262G mreCD ) were transformed with either the empty vector pBAD33 control plasmid or pBAD33- cbtA-F65S (pDH212). Transformants were selected on M9 maltose (0.2% maltose 0.2% casamino acids 1 mM MgSO 4 ) plates supplemented with appropriate antibiotics and 250 μM IPTG at 30°C. Overnight cultures were grown in M9 maltose + 250 μM IPTG at 30°C. These overnight cultures were back diluted to a starting OD600 of 0.03 in LB (Cm + Carb) supplemented with 250 μM IPTG, and grown for 1 h at 30°C (reaching an OD600 ~0.08). Cultures were induced by addition of 0.2% arabinose and grown for an additional 2 h at 30°C, at which point microscopic analysis was performed. (E) Effect of MreB substitution E262G on cell viability in the presence or absence of overproduced CbtA-F65S. Aliquots from the same overnight cultures described in (D) were back diluted 1:100 in M9 maltose + 250 μM IPTG and grown at 30°C for ~5 h (until cultures had reached an OD600 of ~0.7). Cultures were normalized to the same OD600 value, serially diluted, and spotted on LB plates supplemented with 250 μM IPTG ± 0.2% arabinose. Plates were incubated for 48 h at RT. Dilutions 10 0 to 10 −4 are shown. (F) Effect of MreB substitution S269F on cell viability in the presence or absence of an overproduced CbtA variant. Strains DH118/pFB149 and DH118/pDH332 (BW27785 mreBCD :: kan R / plac-mreB-S269F mreCD ) were transformed with empty vector pSG369, pDH335 ( ptet-cbtA-F65S ), or pDH337 ( ptet-cbtA-R15C/F65S ). Spot dilution cultures were prepared as described above for panel (E) and spotted on LB (Spec + Strep) plates supplemented with 250 μM IPTG ± 15 ng/mL or 25 ng/mL ATC. Plates were incubated at 37°C for 48 h. Dilutions 10 −1 to 10 −5 are shown. In all microscopy images, scale bars represent 5 μm.
    Od600, supplied by Molecular Devices LLC, used in various techniques. Bioz Stars score: 99/100, based on 967 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    BioTek Instruments od600
    Viability of E. coli D21 andD21f2 cells after treatment with Bm Glvs. Mid-log phase E. coli D21 and D21f2 cells were diluted to <t>OD600</t> = 0.4 in 10 mM phosphate, 100 mMNaCl, pH 5.0, and then incubated with purified recombinant Bm Glvs (final concentration
    Od600, supplied by BioTek Instruments, used in various techniques. Bioz Stars score: 99/100, based on 2204 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    93
    Implen od600
    Viability of E. coli D21 andD21f2 cells after treatment with Bm Glvs. Mid-log phase E. coli D21 and D21f2 cells were diluted to <t>OD600</t> = 0.4 in 10 mM phosphate, 100 mMNaCl, pH 5.0, and then incubated with purified recombinant Bm Glvs (final concentration
    Od600, supplied by Implen, used in various techniques. Bioz Stars score: 93/100, based on 33 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    od600  (UNICO)
    94
    UNICO od600
    Viability of E. coli D21 andD21f2 cells after treatment with Bm Glvs. Mid-log phase E. coli D21 and D21f2 cells were diluted to <t>OD600</t> = 0.4 in 10 mM phosphate, 100 mMNaCl, pH 5.0, and then incubated with purified recombinant Bm Glvs (final concentration
    Od600, supplied by UNICO, used in various techniques. Bioz Stars score: 94/100, based on 38 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Tecan Systems od600
    TM-TMP is active against E. coli and has similar efficacy to TMP in the presence of GSH. In vitro activity of TM-TMP against E. coli 209. (A) <t>OD600</t> of E. coli after 24 h incubation with various concentrations of TM-TMP (black) and TMP (red), in the absence of GSH. (B) OD600 of E. coli after 24 h incubation with various concentration of TM-TMP (black) and TMP (red), in the presence of 500 μ M of GSH.
    Od600, supplied by Tecan Systems, used in various techniques. Bioz Stars score: 99/100, based on 1636 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    96
    Thermo Fisher density od600
    TM-TMP is active against E. coli and has similar efficacy to TMP in the presence of GSH. In vitro activity of TM-TMP against E. coli 209. (A) <t>OD600</t> of E. coli after 24 h incubation with various concentrations of TM-TMP (black) and TMP (red), in the absence of GSH. (B) OD600 of E. coli after 24 h incubation with various concentration of TM-TMP (black) and TMP (red), in the presence of 500 μ M of GSH.
    Density Od600, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 96/100, based on 68 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    88
    Chrom Tech spectrophotometer od600
    TM-TMP is active against E. coli and has similar efficacy to TMP in the presence of GSH. In vitro activity of TM-TMP against E. coli 209. (A) <t>OD600</t> of E. coli after 24 h incubation with various concentrations of TM-TMP (black) and TMP (red), in the absence of GSH. (B) OD600 of E. coli after 24 h incubation with various concentration of TM-TMP (black) and TMP (red), in the presence of 500 μ M of GSH.
    Spectrophotometer Od600, supplied by Chrom Tech, used in various techniques. Bioz Stars score: 88/100, based on 5 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    94
    Implen od600 diluphotometer
    TM-TMP is active against E. coli and has similar efficacy to TMP in the presence of GSH. In vitro activity of TM-TMP against E. coli 209. (A) <t>OD600</t> of E. coli after 24 h incubation with various concentrations of TM-TMP (black) and TMP (red), in the absence of GSH. (B) OD600 of E. coli after 24 h incubation with various concentration of TM-TMP (black) and TMP (red), in the presence of 500 μ M of GSH.
    Od600 Diluphotometer, supplied by Implen, used in various techniques. Bioz Stars score: 94/100, based on 3 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    86
    Beckman Coulter od600 centrifuge
    TM-TMP is active against E. coli and has similar efficacy to TMP in the presence of GSH. In vitro activity of TM-TMP against E. coli 209. (A) <t>OD600</t> of E. coli after 24 h incubation with various concentrations of TM-TMP (black) and TMP (red), in the absence of GSH. (B) OD600 of E. coli after 24 h incubation with various concentration of TM-TMP (black) and TMP (red), in the presence of 500 μ M of GSH.
    Od600 Centrifuge, supplied by Beckman Coulter, used in various techniques. Bioz Stars score: 86/100, based on 5 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    88
    Thermo Fisher od600 data
    Representation of <t>OD600</t> measurements vs NPs number/μL. The data were analysed by GraphPad software according to a linear regression straight line equation. ( A ) Calibration curve of 100 nm NPs. ( B ) Calibration curve of 200 nm NPs. ( C ) Calibration curve of 460 nm NPs. 1OD600 value, line equation and correlation factor (r2) are indicated for each NP size. Nine replicates were measured per NPs concentration.
    Od600 Data, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 88/100, based on 6 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    93
    Shimadzu Corporation od600 value
    Representation of <t>OD600</t> measurements vs NPs number/μL. The data were analysed by GraphPad software according to a linear regression straight line equation. ( A ) Calibration curve of 100 nm NPs. ( B ) Calibration curve of 200 nm NPs. ( C ) Calibration curve of 460 nm NPs. 1OD600 value, line equation and correlation factor (r2) are indicated for each NP size. Nine replicates were measured per NPs concentration.
    Od600 Value, supplied by Shimadzu Corporation, used in various techniques. Bioz Stars score: 93/100, based on 27 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    96
    Jenway od600
    Representation of <t>OD600</t> measurements vs NPs number/μL. The data were analysed by GraphPad software according to a linear regression straight line equation. ( A ) Calibration curve of 100 nm NPs. ( B ) Calibration curve of 200 nm NPs. ( C ) Calibration curve of 460 nm NPs. 1OD600 value, line equation and correlation factor (r2) are indicated for each NP size. Nine replicates were measured per NPs concentration.
    Od600, supplied by Jenway, used in various techniques. Bioz Stars score: 96/100, based on 93 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    Shimadzu Corporation od600
    a The <t>OD600</t> values of strains Po1f, Po1f-LN-000, Po1f-LN-004, and Po1f-LN-051 cultured in YPD medium, measured at 0, 8, 14, 24, 32, 48, 72, and 96 h. b Squalene production in strains Po1f, Po1f-LN-000, Po1f-LN-004, Po1f-LN-051 cultured in YPD medium for 5 days
    Od600, supplied by Shimadzu Corporation, used in various techniques. Bioz Stars score: 99/100, based on 689 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Bio-Rad od600
    Effect of nicotine on initial attachment of S. epidermidis . Primary attachment of the SE1457 to polystyrene surfaces. Overnight cultures grown to an <t>OD600</t> of 0.6 were diluted with TSB (Control) and TSB supplemented with 4000 μg/ml nicotine (Nicotine) to an OD 600 of 0.1 in PBS and inoculated into a six-well polyethylene plate for 1 h at 37°C. The attached cells were photographed under a light microscope (A) and the cell numbers were counted by Image J software (B) .
    Od600, supplied by Bio-Rad, used in various techniques. Bioz Stars score: 99/100, based on 895 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    PerkinElmer od600
    Effect of nicotine on initial attachment of S. epidermidis . Primary attachment of the SE1457 to polystyrene surfaces. Overnight cultures grown to an <t>OD600</t> of 0.6 were diluted with TSB (Control) and TSB supplemented with 4000 μg/ml nicotine (Nicotine) to an OD 600 of 0.1 in PBS and inoculated into a six-well polyethylene plate for 1 h at 37°C. The attached cells were photographed under a light microscope (A) and the cell numbers were counted by Image J software (B) .
    Od600, supplied by PerkinElmer, used in various techniques. Bioz Stars score: 99/100, based on 815 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    od600  (Difco)
    94
    Difco od600
    SPI-1 induction varies in M9 minimal medium. A, C, E. P prgH activity of the SPI-1 reporter strain (P prgH -gfp [LVA]) from one representative experiment. B, D, F. Comparing LB-M to glucose M9, glucose M9 containing different brands of Casamino acids, and 3 alternative carbon sources in M9, respectively. % promoter activity was calculated from n = 3 at maximum promoter activity of <t>GFP/OD600;</t> normalized to LB-Miller grown bacteria (B and F) or those grown with CAA3 (D). The Casamino acids used are ‘existing’ Bacto (CAA1), Bacto (CAA2), Bacto Technical (CAA3), Sigma (CAA4), Remel (CAA5). Statistical analyses were unpaired t-test with Welch’s correction (B) or a one-way ANOVA with Dunnett’s multiple comparisons test (D, F). Data shown are mean ±SD (* p
    Od600, supplied by Difco, used in various techniques. Bioz Stars score: 94/100, based on 226 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    94
    Formedium od600
    SPI-1 induction varies in M9 minimal medium. A, C, E. P prgH activity of the SPI-1 reporter strain (P prgH -gfp [LVA]) from one representative experiment. B, D, F. Comparing LB-M to glucose M9, glucose M9 containing different brands of Casamino acids, and 3 alternative carbon sources in M9, respectively. % promoter activity was calculated from n = 3 at maximum promoter activity of <t>GFP/OD600;</t> normalized to LB-Miller grown bacteria (B and F) or those grown with CAA3 (D). The Casamino acids used are ‘existing’ Bacto (CAA1), Bacto (CAA2), Bacto Technical (CAA3), Sigma (CAA4), Remel (CAA5). Statistical analyses were unpaired t-test with Welch’s correction (B) or a one-way ANOVA with Dunnett’s multiple comparisons test (D, F). Data shown are mean ±SD (* p
    Od600, supplied by Formedium, used in various techniques. Bioz Stars score: 94/100, based on 66 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    od600  (ibidi)
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    ibidi od600
    SPI-1 induction varies in M9 minimal medium. A, C, E. P prgH activity of the SPI-1 reporter strain (P prgH -gfp [LVA]) from one representative experiment. B, D, F. Comparing LB-M to glucose M9, glucose M9 containing different brands of Casamino acids, and 3 alternative carbon sources in M9, respectively. % promoter activity was calculated from n = 3 at maximum promoter activity of <t>GFP/OD600;</t> normalized to LB-Miller grown bacteria (B and F) or those grown with CAA3 (D). The Casamino acids used are ‘existing’ Bacto (CAA1), Bacto (CAA2), Bacto Technical (CAA3), Sigma (CAA4), Remel (CAA5). Statistical analyses were unpaired t-test with Welch’s correction (B) or a one-way ANOVA with Dunnett’s multiple comparisons test (D, F). Data shown are mean ±SD (* p
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    SPI-1 induction varies in M9 minimal medium. A, C, E. P prgH activity of the SPI-1 reporter strain (P prgH -gfp [LVA]) from one representative experiment. B, D, F. Comparing LB-M to glucose M9, glucose M9 containing different brands of Casamino acids, and 3 alternative carbon sources in M9, respectively. % promoter activity was calculated from n = 3 at maximum promoter activity of <t>GFP/OD600;</t> normalized to LB-Miller grown bacteria (B and F) or those grown with CAA3 (D). The Casamino acids used are ‘existing’ Bacto (CAA1), Bacto (CAA2), Bacto Technical (CAA3), Sigma (CAA4), Remel (CAA5). Statistical analyses were unpaired t-test with Welch’s correction (B) or a one-way ANOVA with Dunnett’s multiple comparisons test (D, F). Data shown are mean ±SD (* p
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    od600  (Roche)
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    Actin mutant alleles affecting the activation of ExoY. ( a ) Drop tests of serial dilutions of S. cerevisiae strains expressing wild-type (wt) actin (SC489), D25Y/D222G (SC690) or D25N (SC691) and p1593 for galactose-induced expression of ExoY or the corresponding vector control YEpGal555. Cell suspensions were normalized to an <t>OD600</t> of 1.0 and 5-fold serial dilutions were applied as 3 μl drops on SD or SG-agar plates. ( b ) Western blot analysis to verify expression of Myc-ExoY in SC690 and SC691 with anti-Myc or anti-RPS9 (loading control). ( c ) Western blot analysis to verify expression of actin in SC489 (1), SC690 (2) and SC691 (3) with anti-actin or anti-RPS9 (loading control). Uncropped images of ( b , c ) are shown in Supplementary Fig. 10 .
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    Actin mutant alleles affecting the activation of ExoY. ( a ) Drop tests of serial dilutions of S. cerevisiae strains expressing wild-type (wt) actin (SC489), D25Y/D222G (SC690) or D25N (SC691) and p1593 for galactose-induced expression of ExoY or the corresponding vector control YEpGal555. Cell suspensions were normalized to an <t>OD600</t> of 1.0 and 5-fold serial dilutions were applied as 3 μl drops on SD or SG-agar plates. ( b ) Western blot analysis to verify expression of Myc-ExoY in SC690 and SC691 with anti-Myc or anti-RPS9 (loading control). ( c ) Western blot analysis to verify expression of actin in SC489 (1), SC690 (2) and SC691 (3) with anti-actin or anti-RPS9 (loading control). Uncropped images of ( b , c ) are shown in Supplementary Fig. 10 .
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    Actin mutant alleles affecting the activation of ExoY. ( a ) Drop tests of serial dilutions of S. cerevisiae strains expressing wild-type (wt) actin (SC489), D25Y/D222G (SC690) or D25N (SC691) and p1593 for galactose-induced expression of ExoY or the corresponding vector control YEpGal555. Cell suspensions were normalized to an <t>OD600</t> of 1.0 and 5-fold serial dilutions were applied as 3 μl drops on SD or SG-agar plates. ( b ) Western blot analysis to verify expression of Myc-ExoY in SC690 and SC691 with anti-Myc or anti-RPS9 (loading control). ( c ) Western blot analysis to verify expression of actin in SC489 (1), SC690 (2) and SC691 (3) with anti-actin or anti-RPS9 (loading control). Uncropped images of ( b , c ) are shown in Supplementary Fig. 10 .
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    Actin mutant alleles affecting the activation of ExoY. ( a ) Drop tests of serial dilutions of S. cerevisiae strains expressing wild-type (wt) actin (SC489), D25Y/D222G (SC690) or D25N (SC691) and p1593 for galactose-induced expression of ExoY or the corresponding vector control YEpGal555. Cell suspensions were normalized to an <t>OD600</t> of 1.0 and 5-fold serial dilutions were applied as 3 μl drops on SD or SG-agar plates. ( b ) Western blot analysis to verify expression of Myc-ExoY in SC690 and SC691 with anti-Myc or anti-RPS9 (loading control). ( c ) Western blot analysis to verify expression of actin in SC489 (1), SC690 (2) and SC691 (3) with anti-actin or anti-RPS9 (loading control). Uncropped images of ( b , c ) are shown in Supplementary Fig. 10 .
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    Image Search Results


    OD600 time course of TOP10-rfp-lys grown in 15 ml tubes upon induction with HSL 100 nM (A) and RFP fluorescence time course in the supernatant (B) . Culture absorbance (A) and supernatant fluorescence (B) of TOP10-rfp-lys induced with HSL 100 nM (blue line). Uninduced TOP10-rfp-lys (red line), TOP10 bearing pHC-RFP induced with HSL 100 nM (green line) or uninduced (black line) are the negative controls. Induction was carried out in the exponential phase at OD600~0.55. Error bars represent the 95% confidence interval of the estimated mean.

    Journal: Journal of Biological Engineering

    Article Title: Characterization of a synthetic bacterial self-destruction device for programmed cell death and for recombinant proteins release

    doi: 10.1186/1754-1611-5-8

    Figure Lengend Snippet: OD600 time course of TOP10-rfp-lys grown in 15 ml tubes upon induction with HSL 100 nM (A) and RFP fluorescence time course in the supernatant (B) . Culture absorbance (A) and supernatant fluorescence (B) of TOP10-rfp-lys induced with HSL 100 nM (blue line). Uninduced TOP10-rfp-lys (red line), TOP10 bearing pHC-RFP induced with HSL 100 nM (green line) or uninduced (black line) are the negative controls. Induction was carried out in the exponential phase at OD600~0.55. Error bars represent the 95% confidence interval of the estimated mean.

    Article Snippet: Three of the six replicates of TOP10-rfp-lys and TOP10 with pHC-RFP were induced with HSL at a final concentration of 100 nM and all the 13 tubes were incubated under the same conditions as before for 125 min. Every 25 min, the OD600 was measured with the NanoDrop ND-1000 and a 300 μl aliquot was taken from each culture.

    Techniques: Fluorescence

    Lysis profile of TOP10 bearing the lysis device in low copy number when induced in different growth phases in microplate reader . OD600 of TOP10 with pLC-T4LysHSL induced with HSL 100 nM (blue line) and uninduced (red line), pLC-T4Lys - induced with HSL 100 nM (green line) and uninduced (black line). Induction was performed in exponential phase (OD600 = 0.2) at t = 0 (A), early stationary phase (OD600~1.3) at t = 4 h (B) and late stationary phase (OD600~2) at t = 20 h (C). Error bars represent the 95% confidence interval of the estimated mean. For clarity of presentation, data points shown here are resampled with a 1-hour sampling time.

    Journal: Journal of Biological Engineering

    Article Title: Characterization of a synthetic bacterial self-destruction device for programmed cell death and for recombinant proteins release

    doi: 10.1186/1754-1611-5-8

    Figure Lengend Snippet: Lysis profile of TOP10 bearing the lysis device in low copy number when induced in different growth phases in microplate reader . OD600 of TOP10 with pLC-T4LysHSL induced with HSL 100 nM (blue line) and uninduced (red line), pLC-T4Lys - induced with HSL 100 nM (green line) and uninduced (black line). Induction was performed in exponential phase (OD600 = 0.2) at t = 0 (A), early stationary phase (OD600~1.3) at t = 4 h (B) and late stationary phase (OD600~2) at t = 20 h (C). Error bars represent the 95% confidence interval of the estimated mean. For clarity of presentation, data points shown here are resampled with a 1-hour sampling time.

    Article Snippet: Three of the six replicates of TOP10-rfp-lys and TOP10 with pHC-RFP were induced with HSL at a final concentration of 100 nM and all the 13 tubes were incubated under the same conditions as before for 125 min. Every 25 min, the OD600 was measured with the NanoDrop ND-1000 and a 300 μl aliquot was taken from each culture.

    Techniques: Lysis, Low Copy Number, Planar Chromatography, Sampling

    Transfer function, rise time and delay time of the HSL-inducible lysis device in low copy plasmid in early stationary phase in microplate reader . Lysis entity of TOP10 cells with pLC-T4LysHSL induced with different concentrations of HSL in early stationary phase at OD600 = 0.9 (A). The experimental data (circles) were fitted with a Hill function (line, Vmax = 76, K 50 = 0.37, n = 1.3). For each concentration, the rise time, i.e. the time to rise from the 10% to 90% of the lysis entity (B) and the delay time before the OD600 drop after induction (C) are also shown. Error bars represent the 95% confidence interval of the estimated mean.

    Journal: Journal of Biological Engineering

    Article Title: Characterization of a synthetic bacterial self-destruction device for programmed cell death and for recombinant proteins release

    doi: 10.1186/1754-1611-5-8

    Figure Lengend Snippet: Transfer function, rise time and delay time of the HSL-inducible lysis device in low copy plasmid in early stationary phase in microplate reader . Lysis entity of TOP10 cells with pLC-T4LysHSL induced with different concentrations of HSL in early stationary phase at OD600 = 0.9 (A). The experimental data (circles) were fitted with a Hill function (line, Vmax = 76, K 50 = 0.37, n = 1.3). For each concentration, the rise time, i.e. the time to rise from the 10% to 90% of the lysis entity (B) and the delay time before the OD600 drop after induction (C) are also shown. Error bars represent the 95% confidence interval of the estimated mean.

    Article Snippet: Three of the six replicates of TOP10-rfp-lys and TOP10 with pHC-RFP were induced with HSL at a final concentration of 100 nM and all the 13 tubes were incubated under the same conditions as before for 125 min. Every 25 min, the OD600 was measured with the NanoDrop ND-1000 and a 300 μl aliquot was taken from each culture.

    Techniques: Lysis, Plasmid Preparation, Planar Chromatography, Concentration Assay

    Lysis dynamics of TOP10 bearing the thermoinducible lysis device in low copy plasmid grown in microplate reader . OD600 of TOP10 with pLC-T4LysHeat induced with a temperature shift from 30°C to 42°C in the microplate reader (blue line). Heat-induced pLC-T4Lys - (green line) is shown as the negative control. Induction was performed in exponential phase at OD600 = 0.3. Error bars represent the 95% confidence interval of the estimated mean. For clarity of presentation, data points shown here are resampled with a 30-minute sampling time.

    Journal: Journal of Biological Engineering

    Article Title: Characterization of a synthetic bacterial self-destruction device for programmed cell death and for recombinant proteins release

    doi: 10.1186/1754-1611-5-8

    Figure Lengend Snippet: Lysis dynamics of TOP10 bearing the thermoinducible lysis device in low copy plasmid grown in microplate reader . OD600 of TOP10 with pLC-T4LysHeat induced with a temperature shift from 30°C to 42°C in the microplate reader (blue line). Heat-induced pLC-T4Lys - (green line) is shown as the negative control. Induction was performed in exponential phase at OD600 = 0.3. Error bars represent the 95% confidence interval of the estimated mean. For clarity of presentation, data points shown here are resampled with a 30-minute sampling time.

    Article Snippet: Three of the six replicates of TOP10-rfp-lys and TOP10 with pHC-RFP were induced with HSL at a final concentration of 100 nM and all the 13 tubes were incubated under the same conditions as before for 125 min. Every 25 min, the OD600 was measured with the NanoDrop ND-1000 and a 300 μl aliquot was taken from each culture.

    Techniques: Lysis, Plasmid Preparation, Planar Chromatography, Negative Control, Sampling

    Growth of various L. paracasei strains in a chemically defined medium with methionine as the sole sulfur source. The bars and black lines represent the mean and standard deviation of the reached OD600 after 25 h of growth, respectively, of three independent biologically repeated measurements. Strains that reached an OD600 of one or more were consider as being able to grow in a medium with methionine as sole sulfur source.

    Journal: Frontiers in Microbiology

    Article Title: Conversion of Methionine to Cysteine in Lactobacillus paracasei Depends on the Highly Mobile cysK-ctl-cysE Gene Cluster

    doi: 10.3389/fmicb.2018.02415

    Figure Lengend Snippet: Growth of various L. paracasei strains in a chemically defined medium with methionine as the sole sulfur source. The bars and black lines represent the mean and standard deviation of the reached OD600 after 25 h of growth, respectively, of three independent biologically repeated measurements. Strains that reached an OD600 of one or more were consider as being able to grow in a medium with methionine as sole sulfur source.

    Article Snippet: Optical density at 600 nm (OD600) was determined with a spectrophotometer (LKB Biochrom 4050 Ultrospec II).

    Techniques: Standard Deviation

    a Phylogenetic tree illustrating the separation of S. mitis NCTC12261 T ]). b Kinetics of transformation in the S. mitis type strain and strain SK321. Pre-cultures at OD600 0.5 were diluted 1:100 in C + Y YB medium and grown until OD600 0.04 at 37 °C in 5% CO 2 . Cultures were treated with 300 nM cognate CSP and distributed into 200 μL aliquots that were further exposed to 1 μg ml − 1 recombinant plasmid pVA838 at indicated times. 20 U ml − 1 DNase I were added after 30 min of exposure to DNA and the culture was incubated in air at 37 °C for additional 30 min. Transformants were recovered in blood agar plates supplemented with Erythromycin. Each line represents results of a single experiment

    Journal: BMC Genomics

    Article Title: High-resolution profiles of the Streptococcus mitis CSP signaling pathway reveal core and strain-specific regulated genes

    doi: 10.1186/s12864-018-4802-y

    Figure Lengend Snippet: a Phylogenetic tree illustrating the separation of S. mitis NCTC12261 T ]). b Kinetics of transformation in the S. mitis type strain and strain SK321. Pre-cultures at OD600 0.5 were diluted 1:100 in C + Y YB medium and grown until OD600 0.04 at 37 °C in 5% CO 2 . Cultures were treated with 300 nM cognate CSP and distributed into 200 μL aliquots that were further exposed to 1 μg ml − 1 recombinant plasmid pVA838 at indicated times. 20 U ml − 1 DNase I were added after 30 min of exposure to DNA and the culture was incubated in air at 37 °C for additional 30 min. Transformants were recovered in blood agar plates supplemented with Erythromycin. Each line represents results of a single experiment

    Article Snippet: Pre-cultures of NCTC12261T and SK321 at OD600 of 0.5 were centrifuged at 8000 g, 4 °C, for 10 min, resuspendend 100-fold in TSB or C + YYB and the diluted cultures were incubated at 37 °C 5% CO2 until an OD600 of 0.04 was reached.

    Techniques: Transformation Assay, Recombinant, Plasmid Preparation, Incubation

    Growth characteristics and hemolytic activities of the A. pleuropneumoniae mutants. a The growth curves of the S-8, S-8 △clp and S-8 △clp△apxIIC strains. Overnight cultures were inoculated into fresh TSB and then incubated at 37 °C. Growth was monitored by OD600 at an interval of 1 h. b Hemolytic activity test for the S-8, S-8 △clp and S-8 △clp△apxIIC strains. Section 1, S-8; section 2, S-8 △clp ; section 3, S-8 △clp△apxIIC

    Journal: BMC Veterinary Research

    Article Title: Attenuated Actinobacillus pleuropneumoniae double-deletion mutant S-8∆clpP/apxIIC confers protection against homologous or heterologous strain challenge

    doi: 10.1186/s12917-016-0928-9

    Figure Lengend Snippet: Growth characteristics and hemolytic activities of the A. pleuropneumoniae mutants. a The growth curves of the S-8, S-8 △clp and S-8 △clp△apxIIC strains. Overnight cultures were inoculated into fresh TSB and then incubated at 37 °C. Growth was monitored by OD600 at an interval of 1 h. b Hemolytic activity test for the S-8, S-8 △clp and S-8 △clp△apxIIC strains. Section 1, S-8; section 2, S-8 △clp ; section 3, S-8 △clp△apxIIC

    Article Snippet: The OD600 values were recorded at an interval of 1 h using the Eppendorf BioPhotometer (Eppendorf, Germany).

    Techniques: Incubation, Activity Assay

    Growth curves of P. fluorescens strains cLP6a and cLP6a-1 . Growth of P. fluorescens strains cLP6a and cLP6a-1 at (a) 10°C, (b) 28°C or (c) 35°C determined as OD600 Each data point is the mean of three independent cultures, and error bars, where visible, indicate the standard deviation.

    Journal: BMC Microbiology

    Article Title: An alternative physiological role for the EmhABC efflux pump in Pseudomonas fluorescens cLP6a

    doi: 10.1186/1471-2180-11-252

    Figure Lengend Snippet: Growth curves of P. fluorescens strains cLP6a and cLP6a-1 . Growth of P. fluorescens strains cLP6a and cLP6a-1 at (a) 10°C, (b) 28°C or (c) 35°C determined as OD600 Each data point is the mean of three independent cultures, and error bars, where visible, indicate the standard deviation.

    Article Snippet: Growth was measured as optical density at 600 nm (OD600 ) using an Ultrospec 3100 pro UV/Visible spectrophotometer (GE Healthcare Bio-Sciences), diluting the TSB blank and culture sample with distilled water as necessary.

    Techniques: Standard Deviation

    Residues on the flat surface of MreB are critical for CbtA-F65S-mediated cell elongation inhibition. (A) Schematic of MreB complementation assay (panels B and C). Depletion strain FB30/pFB174 ( mreBCD :: kan R /pBAD-mreBCD) [ 9 ] was transformed with either an empty vector control (pMLB1113) or a plasmid derived from pFB149 ( plac-mreBCD ) [ 9 ] expressing wild-type mreB (pFB149), mreB-E262G (pDH279), or mreB-S269F (pDH332). (B) Spot dilution assay to assess the ability of each mreB allele to support growth on solid medium. Cultures of each strain grown overnight at 37°C in M9 maltose (0.2% maltose, 0.2% casamino acids, 1 mM MgSO 4 ) supplemented with 0.5% arabinose were back-diluted 1:100 in fresh M9 maltose + 0.5% arabinose and grown at 37°C for several hours until they reached late log phase. Cultures were pelleted and washed to remove arabinose, normalized to the same OD600 value, serially diluted, and spotted on LB plates supplemented with either 0.5% arabinose or 250 μM IPTG. Plates were incubated overnight at 37°C. Dilutions 10 −1 to 10 −5 are shown. (C) Cell morphology phenotypes of complemented strains. Aliquots from the same overnight cultures described in (B) were washed once and resuspended in fresh M9 maltose (without arabinose). These washed aliquots were used as inocula for M9 maltose cultures supplemented with 250 μM IPTG. All cultures were grown at 37°C for several hours (maintained in log-phase by back dilution) and cell morphology was monitored periodically by microscopy. Images taken after 5 h of growth in the presence of 250 μM IPTG are shown. (D) Effect of MreB substitution E262G on cell morphology phenotypes in the presence or absence of overproduced CbtA-F65S. Strains DH118/pFB149 (BW27785 mreBCD :: kan R / plac-mreBCD ) and DH118/pDH278 (BW27785 mreBCD :: kan R / plac-mreB-E262G mreCD ) were transformed with either the empty vector pBAD33 control plasmid or pBAD33- cbtA-F65S (pDH212). Transformants were selected on M9 maltose (0.2% maltose 0.2% casamino acids 1 mM MgSO 4 ) plates supplemented with appropriate antibiotics and 250 μM IPTG at 30°C. Overnight cultures were grown in M9 maltose + 250 μM IPTG at 30°C. These overnight cultures were back diluted to a starting OD600 of 0.03 in LB (Cm + Carb) supplemented with 250 μM IPTG, and grown for 1 h at 30°C (reaching an OD600 ~0.08). Cultures were induced by addition of 0.2% arabinose and grown for an additional 2 h at 30°C, at which point microscopic analysis was performed. (E) Effect of MreB substitution E262G on cell viability in the presence or absence of overproduced CbtA-F65S. Aliquots from the same overnight cultures described in (D) were back diluted 1:100 in M9 maltose + 250 μM IPTG and grown at 30°C for ~5 h (until cultures had reached an OD600 of ~0.7). Cultures were normalized to the same OD600 value, serially diluted, and spotted on LB plates supplemented with 250 μM IPTG ± 0.2% arabinose. Plates were incubated for 48 h at RT. Dilutions 10 0 to 10 −4 are shown. (F) Effect of MreB substitution S269F on cell viability in the presence or absence of an overproduced CbtA variant. Strains DH118/pFB149 and DH118/pDH332 (BW27785 mreBCD :: kan R / plac-mreB-S269F mreCD ) were transformed with empty vector pSG369, pDH335 ( ptet-cbtA-F65S ), or pDH337 ( ptet-cbtA-R15C/F65S ). Spot dilution cultures were prepared as described above for panel (E) and spotted on LB (Spec + Strep) plates supplemented with 250 μM IPTG ± 15 ng/mL or 25 ng/mL ATC. Plates were incubated at 37°C for 48 h. Dilutions 10 −1 to 10 −5 are shown. In all microscopy images, scale bars represent 5 μm.

    Journal: PLoS Genetics

    Article Title: CbtA toxin of Escherichia coli inhibits cell division and cell elongation via direct and independent interactions with FtsZ and MreB

    doi: 10.1371/journal.pgen.1007007

    Figure Lengend Snippet: Residues on the flat surface of MreB are critical for CbtA-F65S-mediated cell elongation inhibition. (A) Schematic of MreB complementation assay (panels B and C). Depletion strain FB30/pFB174 ( mreBCD :: kan R /pBAD-mreBCD) [ 9 ] was transformed with either an empty vector control (pMLB1113) or a plasmid derived from pFB149 ( plac-mreBCD ) [ 9 ] expressing wild-type mreB (pFB149), mreB-E262G (pDH279), or mreB-S269F (pDH332). (B) Spot dilution assay to assess the ability of each mreB allele to support growth on solid medium. Cultures of each strain grown overnight at 37°C in M9 maltose (0.2% maltose, 0.2% casamino acids, 1 mM MgSO 4 ) supplemented with 0.5% arabinose were back-diluted 1:100 in fresh M9 maltose + 0.5% arabinose and grown at 37°C for several hours until they reached late log phase. Cultures were pelleted and washed to remove arabinose, normalized to the same OD600 value, serially diluted, and spotted on LB plates supplemented with either 0.5% arabinose or 250 μM IPTG. Plates were incubated overnight at 37°C. Dilutions 10 −1 to 10 −5 are shown. (C) Cell morphology phenotypes of complemented strains. Aliquots from the same overnight cultures described in (B) were washed once and resuspended in fresh M9 maltose (without arabinose). These washed aliquots were used as inocula for M9 maltose cultures supplemented with 250 μM IPTG. All cultures were grown at 37°C for several hours (maintained in log-phase by back dilution) and cell morphology was monitored periodically by microscopy. Images taken after 5 h of growth in the presence of 250 μM IPTG are shown. (D) Effect of MreB substitution E262G on cell morphology phenotypes in the presence or absence of overproduced CbtA-F65S. Strains DH118/pFB149 (BW27785 mreBCD :: kan R / plac-mreBCD ) and DH118/pDH278 (BW27785 mreBCD :: kan R / plac-mreB-E262G mreCD ) were transformed with either the empty vector pBAD33 control plasmid or pBAD33- cbtA-F65S (pDH212). Transformants were selected on M9 maltose (0.2% maltose 0.2% casamino acids 1 mM MgSO 4 ) plates supplemented with appropriate antibiotics and 250 μM IPTG at 30°C. Overnight cultures were grown in M9 maltose + 250 μM IPTG at 30°C. These overnight cultures were back diluted to a starting OD600 of 0.03 in LB (Cm + Carb) supplemented with 250 μM IPTG, and grown for 1 h at 30°C (reaching an OD600 ~0.08). Cultures were induced by addition of 0.2% arabinose and grown for an additional 2 h at 30°C, at which point microscopic analysis was performed. (E) Effect of MreB substitution E262G on cell viability in the presence or absence of overproduced CbtA-F65S. Aliquots from the same overnight cultures described in (D) were back diluted 1:100 in M9 maltose + 250 μM IPTG and grown at 30°C for ~5 h (until cultures had reached an OD600 of ~0.7). Cultures were normalized to the same OD600 value, serially diluted, and spotted on LB plates supplemented with 250 μM IPTG ± 0.2% arabinose. Plates were incubated for 48 h at RT. Dilutions 10 0 to 10 −4 are shown. (F) Effect of MreB substitution S269F on cell viability in the presence or absence of an overproduced CbtA variant. Strains DH118/pFB149 and DH118/pDH332 (BW27785 mreBCD :: kan R / plac-mreB-S269F mreCD ) were transformed with empty vector pSG369, pDH335 ( ptet-cbtA-F65S ), or pDH337 ( ptet-cbtA-R15C/F65S ). Spot dilution cultures were prepared as described above for panel (E) and spotted on LB (Spec + Strep) plates supplemented with 250 μM IPTG ± 15 ng/mL or 25 ng/mL ATC. Plates were incubated at 37°C for 48 h. Dilutions 10 −1 to 10 −5 are shown. In all microscopy images, scale bars represent 5 μm.

    Article Snippet: Growth was monitored every 30–60 min by transferring 200 μL to a sterile microtitre plate and taking OD600 measurements on a plate reader (Molecular Devices).

    Techniques: Inhibition, Transformation Assay, Plasmid Preparation, Derivative Assay, Expressing, Dilution Assay, Incubation, Microscopy, Variant Assay

    CbtA interacts with Bsu FtsZ chimera containing Eco H6/H7 loop sequence. (A) Amino acid sequence alignment of the H6/H7 loop sequences from E . coli (residues 168–182) and B . subtilis (residues 169–183) is shown. Identical residues are shown in black; similar residues are shown in gray. Alignment was prepared using Boxshade. (B) Two-hybrid analysis shows that a Bsu FtsZ chimera containing the H6/H7 loop sequence from Eco FtsZ (residues 169–183 of Bsu FtsZ are replaced with residues 168–182 of Eco FtsZ) can interact with CbtA. Reporter strain cells containing compatible plasmids encoding the indicated λCI-FtsZ variant (or λCI) and α-CbtA (or wild-type α) were grown in the presence of 100 μM IPTG and assayed for β-galactosidase. Bars represent the average β-galactosidase activity from three independent measurements; error bars represent standard deviations. Dashed line designates highest basal lacZ expression, i.e. the Miller Unit value of the highest empty vector control. (C) Spot dilution assay was used to measure CbtA toxicity in B . subtilis strains containing the indicated ftsZ allele and the indicated his 6 -cbtA-gfp allele (or his 6 - gfp only). A single colony of each strain was grown in LB at 37°C until late-log phase. All cultures were normalized to the same OD600 value, serially diluted, and spotted on LB plates with or without 1mM IPTG. Plates were incubated at 37°C overnight. Dilutions 10 −1 to 10 −5 are shown. (D) Growth curve analysis was performed on B . subtilis strains containing the indicated ftsZ allele and the indicated his 6 -cbtA-gfp allele (or his 6 - gfp only). Triplicate cultures of each strain were grown in LB ± 1 mM IPTG at 37°C over several hours. Each point represents the average of triplicate values; error bars represent standard deviation.

    Journal: PLoS Genetics

    Article Title: CbtA toxin of Escherichia coli inhibits cell division and cell elongation via direct and independent interactions with FtsZ and MreB

    doi: 10.1371/journal.pgen.1007007

    Figure Lengend Snippet: CbtA interacts with Bsu FtsZ chimera containing Eco H6/H7 loop sequence. (A) Amino acid sequence alignment of the H6/H7 loop sequences from E . coli (residues 168–182) and B . subtilis (residues 169–183) is shown. Identical residues are shown in black; similar residues are shown in gray. Alignment was prepared using Boxshade. (B) Two-hybrid analysis shows that a Bsu FtsZ chimera containing the H6/H7 loop sequence from Eco FtsZ (residues 169–183 of Bsu FtsZ are replaced with residues 168–182 of Eco FtsZ) can interact with CbtA. Reporter strain cells containing compatible plasmids encoding the indicated λCI-FtsZ variant (or λCI) and α-CbtA (or wild-type α) were grown in the presence of 100 μM IPTG and assayed for β-galactosidase. Bars represent the average β-galactosidase activity from three independent measurements; error bars represent standard deviations. Dashed line designates highest basal lacZ expression, i.e. the Miller Unit value of the highest empty vector control. (C) Spot dilution assay was used to measure CbtA toxicity in B . subtilis strains containing the indicated ftsZ allele and the indicated his 6 -cbtA-gfp allele (or his 6 - gfp only). A single colony of each strain was grown in LB at 37°C until late-log phase. All cultures were normalized to the same OD600 value, serially diluted, and spotted on LB plates with or without 1mM IPTG. Plates were incubated at 37°C overnight. Dilutions 10 −1 to 10 −5 are shown. (D) Growth curve analysis was performed on B . subtilis strains containing the indicated ftsZ allele and the indicated his 6 -cbtA-gfp allele (or his 6 - gfp only). Triplicate cultures of each strain were grown in LB ± 1 mM IPTG at 37°C over several hours. Each point represents the average of triplicate values; error bars represent standard deviation.

    Article Snippet: Growth was monitored every 30–60 min by transferring 200 μL to a sterile microtitre plate and taking OD600 measurements on a plate reader (Molecular Devices).

    Techniques: Sequencing, Variant Assay, Activity Assay, Expressing, Plasmid Preparation, Dilution Assay, Incubation, Standard Deviation

    CbtA homologs YpjF and YkfI inhibit cell elongation and cell division in a conserved manner. (A) Spot dilution analysis shows that overproduction of His 6 -YpjF-GFP and His 6 -YkfI-GFP from pCA24N-derived plasmids pMT138 and p3-37, respectively, is toxic. Overnight cultures of BW27785/pMT136 (directing the synthesis of His 6 -GFP), BW27785/pMT138, and BW27785/p3-37 were back diluted to a starting OD600 of 0.05 in fresh LB (Cm) and grown until late-log phase at 37°C. Cultures were normalized, serially diluted, and spotted on LB (Cm) with or without 100 μM IPTG. Plates were incubated at 37°C overnight. Dilutions 10 −1 to 10 −5 are shown. (B) Cell morphology phenotypes. Strains BW27785/pMT138 ( his 6 -ypjF-gfp) , BW27785/pMT188 ( his 6 -ypjF-F65S-gfp ), BW27785/p3-37 ( his 6 -ykfI-gfp) , or BW27785/pMT144 ( his 6 -ykfI-F65S-gfp ) were imaged after 1.5 h induction at 37°C with 100 μM IPTG. (C, D) Bacterial two-hybrid assay detects interactions of YpjF and YkfI with FtsZ or MreB. Results of β-galactosidase assays performed with reporter strain cells containing compatible plasmids encoding the indicated λCI fusion protein and the indicated α fusion protein (α-FtsZ in (C) and α-MreB in (D)) or wild-type α. Cells were grown in the presence of 25 μM IPTG (C) and 100 μM IPTG (D). (E) Two-hybrid analysis shows that the Bsu FtsZ chimera containing the Eco H6/H7 loop (fused to λCI) can interact with both YkfI and YpjF (fused to α). Cells were grown in the presence of 100 μM IPTG. (F) Effect of MreB substitutions E262G and S269F on the two-hybrid interaction between MreB and YpjF. Cells were grown in the presence of 100 μM IPTG. In (C-F), bars represent averages of triplicate values, error bars represent standard deviation, and dashed lines designate highest basal lacZ expression. (G) Strains DH118/pFB149 and DH118/pDH278 (see legend to Fig 9D ) were transformed with either the empty vector pBAD33 control plasmid or pBAD33- ypjF-F65S (pDH289). Transformants were selected on M9 maltose (0.2% maltose, 0.2% casamino acids, 1 mM MgSO 4 ) plates supplemented with appropriate antibiotics and 250 μM IPTG at 30°C. Overnight cultures were grown in M9 maltose + 250 μM IPTG at 30°C. The next morning, cultures were back diluted to a starting OD600 of 0.03 in LB (Cm + Carb) supplemented with 250 μM IPTG, and grown for 1 h at 30°C (reaching an OD600 ~0.08). Cultures were induced by addition of 0.2% arabinose and grown for an additional 2 h at 30°C, at which point microscopic analysis was performed. Scale bars represent 5 μm in (B) and (G).

    Journal: PLoS Genetics

    Article Title: CbtA toxin of Escherichia coli inhibits cell division and cell elongation via direct and independent interactions with FtsZ and MreB

    doi: 10.1371/journal.pgen.1007007

    Figure Lengend Snippet: CbtA homologs YpjF and YkfI inhibit cell elongation and cell division in a conserved manner. (A) Spot dilution analysis shows that overproduction of His 6 -YpjF-GFP and His 6 -YkfI-GFP from pCA24N-derived plasmids pMT138 and p3-37, respectively, is toxic. Overnight cultures of BW27785/pMT136 (directing the synthesis of His 6 -GFP), BW27785/pMT138, and BW27785/p3-37 were back diluted to a starting OD600 of 0.05 in fresh LB (Cm) and grown until late-log phase at 37°C. Cultures were normalized, serially diluted, and spotted on LB (Cm) with or without 100 μM IPTG. Plates were incubated at 37°C overnight. Dilutions 10 −1 to 10 −5 are shown. (B) Cell morphology phenotypes. Strains BW27785/pMT138 ( his 6 -ypjF-gfp) , BW27785/pMT188 ( his 6 -ypjF-F65S-gfp ), BW27785/p3-37 ( his 6 -ykfI-gfp) , or BW27785/pMT144 ( his 6 -ykfI-F65S-gfp ) were imaged after 1.5 h induction at 37°C with 100 μM IPTG. (C, D) Bacterial two-hybrid assay detects interactions of YpjF and YkfI with FtsZ or MreB. Results of β-galactosidase assays performed with reporter strain cells containing compatible plasmids encoding the indicated λCI fusion protein and the indicated α fusion protein (α-FtsZ in (C) and α-MreB in (D)) or wild-type α. Cells were grown in the presence of 25 μM IPTG (C) and 100 μM IPTG (D). (E) Two-hybrid analysis shows that the Bsu FtsZ chimera containing the Eco H6/H7 loop (fused to λCI) can interact with both YkfI and YpjF (fused to α). Cells were grown in the presence of 100 μM IPTG. (F) Effect of MreB substitutions E262G and S269F on the two-hybrid interaction between MreB and YpjF. Cells were grown in the presence of 100 μM IPTG. In (C-F), bars represent averages of triplicate values, error bars represent standard deviation, and dashed lines designate highest basal lacZ expression. (G) Strains DH118/pFB149 and DH118/pDH278 (see legend to Fig 9D ) were transformed with either the empty vector pBAD33 control plasmid or pBAD33- ypjF-F65S (pDH289). Transformants were selected on M9 maltose (0.2% maltose, 0.2% casamino acids, 1 mM MgSO 4 ) plates supplemented with appropriate antibiotics and 250 μM IPTG at 30°C. Overnight cultures were grown in M9 maltose + 250 μM IPTG at 30°C. The next morning, cultures were back diluted to a starting OD600 of 0.03 in LB (Cm + Carb) supplemented with 250 μM IPTG, and grown for 1 h at 30°C (reaching an OD600 ~0.08). Cultures were induced by addition of 0.2% arabinose and grown for an additional 2 h at 30°C, at which point microscopic analysis was performed. Scale bars represent 5 μm in (B) and (G).

    Article Snippet: Growth was monitored every 30–60 min by transferring 200 μL to a sterile microtitre plate and taking OD600 measurements on a plate reader (Molecular Devices).

    Techniques: Derivative Assay, Incubation, Two Hybrid Assay, Standard Deviation, Expressing, Transformation Assay, Plasmid Preparation

    Residues in the H6/H7 loop are necessary for CbtA-FtsZ interaction. (A) Graphs show the results of β-galactosidase assays performed with two-hybrid reporter strain cells containing two compatible plasmids: one that encoded the indicated α-FtsZ variant or wild-type α (all three panels) and another that encoded the indicated λCI fusion protein (λCI-CbtA, top; λCI-ZipA CTD (residues 186–328), middle; λCI-FtsZ, bottom) or λCI. The cells were grown in the presence of 100 μM IPTG (top), 25 μM IPTG (middle), and 100 μM IPTG (bottom). Bars represent the average β-galactosidase activity from three independent measurements, and error bars represent standard deviations. Dashed lines designate highest basal lacZ expression, i.e. the Miller Unit value of the highest empty vector control. (B) Crystal structure depicting two Methanococcus jannaschii FtsZ monomers (space-filled in wheat and purple, PDB 1W5B [ 64 ]) in head-to-tail arrangement. The T7 loop (shown in orange) contacts the GTP nucleotide (green) bound within the GTP-binding pocket. The loop connecting helix 6 and helix 7 (H6/H7 loop) is shown in yellow. (C) Phase contrast images were taken of either wild-type BW27785 or BW27785 ftsZ-L169P (DH73) cells overproducing (or not) His 6 -CbtA-GFP (encoded on plasmid pMT139). Overnight cultures grown at 30°C in M9 maltose (0.4% maltose, 1mM MgSO 4 , 0.01% casamino acids) were back-diluted 1:3,000 into fresh medium and grown at 30°C for ~14 h until they reached an OD600 of 0.3. Cultures were then induced with 100 μM IPTG (or not) and grown for 8 h at 30°C. Scale bars represent 5 μm. (D) Quantification of cell roundness (cell width/ cell length) from cells in (C). Histograms include compiled measurements from three-independent experiments ( n = 725, wild-type without IPTG; n = 706, wild-type + IPTG; n = 739, L169P without IPTG; n = 813, L169P +IPTG). Width and length measurements were made manually in ImageJ[ 65 ] using the ObjectJ plugin.

    Journal: PLoS Genetics

    Article Title: CbtA toxin of Escherichia coli inhibits cell division and cell elongation via direct and independent interactions with FtsZ and MreB

    doi: 10.1371/journal.pgen.1007007

    Figure Lengend Snippet: Residues in the H6/H7 loop are necessary for CbtA-FtsZ interaction. (A) Graphs show the results of β-galactosidase assays performed with two-hybrid reporter strain cells containing two compatible plasmids: one that encoded the indicated α-FtsZ variant or wild-type α (all three panels) and another that encoded the indicated λCI fusion protein (λCI-CbtA, top; λCI-ZipA CTD (residues 186–328), middle; λCI-FtsZ, bottom) or λCI. The cells were grown in the presence of 100 μM IPTG (top), 25 μM IPTG (middle), and 100 μM IPTG (bottom). Bars represent the average β-galactosidase activity from three independent measurements, and error bars represent standard deviations. Dashed lines designate highest basal lacZ expression, i.e. the Miller Unit value of the highest empty vector control. (B) Crystal structure depicting two Methanococcus jannaschii FtsZ monomers (space-filled in wheat and purple, PDB 1W5B [ 64 ]) in head-to-tail arrangement. The T7 loop (shown in orange) contacts the GTP nucleotide (green) bound within the GTP-binding pocket. The loop connecting helix 6 and helix 7 (H6/H7 loop) is shown in yellow. (C) Phase contrast images were taken of either wild-type BW27785 or BW27785 ftsZ-L169P (DH73) cells overproducing (or not) His 6 -CbtA-GFP (encoded on plasmid pMT139). Overnight cultures grown at 30°C in M9 maltose (0.4% maltose, 1mM MgSO 4 , 0.01% casamino acids) were back-diluted 1:3,000 into fresh medium and grown at 30°C for ~14 h until they reached an OD600 of 0.3. Cultures were then induced with 100 μM IPTG (or not) and grown for 8 h at 30°C. Scale bars represent 5 μm. (D) Quantification of cell roundness (cell width/ cell length) from cells in (C). Histograms include compiled measurements from three-independent experiments ( n = 725, wild-type without IPTG; n = 706, wild-type + IPTG; n = 739, L169P without IPTG; n = 813, L169P +IPTG). Width and length measurements were made manually in ImageJ[ 65 ] using the ObjectJ plugin.

    Article Snippet: Growth was monitored every 30–60 min by transferring 200 μL to a sterile microtitre plate and taking OD600 measurements on a plate reader (Molecular Devices).

    Techniques: Variant Assay, Activity Assay, Expressing, Plasmid Preparation, Binding Assay

    Viability of E. coli D21 andD21f2 cells after treatment with Bm Glvs. Mid-log phase E. coli D21 and D21f2 cells were diluted to OD600 = 0.4 in 10 mM phosphate, 100 mMNaCl, pH 5.0, and then incubated with purified recombinant Bm Glvs (final concentration

    Journal: Insect biochemistry and molecular biology

    Article Title: Gloverins of the silkworm Bombyx mori: Structural and binding properties and activities

    doi: 10.1016/j.ibmb.2013.03.013

    Figure Lengend Snippet: Viability of E. coli D21 andD21f2 cells after treatment with Bm Glvs. Mid-log phase E. coli D21 and D21f2 cells were diluted to OD600 = 0.4 in 10 mM phosphate, 100 mMNaCl, pH 5.0, and then incubated with purified recombinant Bm Glvs (final concentration

    Article Snippet: OD600 was measured every hour by Powerwave XS plate reader (BioTek, VT, US).

    Techniques: Incubation, Purification, Recombinant, Concentration Assay

    Dose-dependent activity of Bm Glvs against E. coli D21 and D21f2 mutant strains. Mid-log phase E. coli D21 and D21f2 cells were diluted to OD600 = 0.4 in 10 mM phosphate, 100 mM NaCl, pH 5.0, and then incubated with increasing concentrations of purified

    Journal: Insect biochemistry and molecular biology

    Article Title: Gloverins of the silkworm Bombyx mori: Structural and binding properties and activities

    doi: 10.1016/j.ibmb.2013.03.013

    Figure Lengend Snippet: Dose-dependent activity of Bm Glvs against E. coli D21 and D21f2 mutant strains. Mid-log phase E. coli D21 and D21f2 cells were diluted to OD600 = 0.4 in 10 mM phosphate, 100 mM NaCl, pH 5.0, and then incubated with increasing concentrations of purified

    Article Snippet: OD600 was measured every hour by Powerwave XS plate reader (BioTek, VT, US).

    Techniques: Activity Assay, Mutagenesis, Incubation, Purification

    TM-TMP is active against E. coli and has similar efficacy to TMP in the presence of GSH. In vitro activity of TM-TMP against E. coli 209. (A) OD600 of E. coli after 24 h incubation with various concentrations of TM-TMP (black) and TMP (red), in the absence of GSH. (B) OD600 of E. coli after 24 h incubation with various concentration of TM-TMP (black) and TMP (red), in the presence of 500 μ M of GSH.

    Journal: Bioconjugate chemistry

    Article Title: A Trimethoprim Conjugate of Thiomaltose Has Enhanced Antibacterial Efficacy In Vivo

    doi: 10.1021/acs.bioconjchem.8b00177

    Figure Lengend Snippet: TM-TMP is active against E. coli and has similar efficacy to TMP in the presence of GSH. In vitro activity of TM-TMP against E. coli 209. (A) OD600 of E. coli after 24 h incubation with various concentrations of TM-TMP (black) and TMP (red), in the absence of GSH. (B) OD600 of E. coli after 24 h incubation with various concentration of TM-TMP (black) and TMP (red), in the presence of 500 μ M of GSH.

    Article Snippet: The plate was incubated at 37 °C, shaken at 190 rpm for 24 h, and the OD600 was measured using a Tecan Infinite 200 plate reader.

    Techniques: In Vitro, Activity Assay, Incubation, Concentration Assay

    The un-evolved co-culture of strains K and L. (A) Nutrient exchange and dependence in co-culture of two E. coli strains L and K. Strain L is incapable of synthesizing leucine, while strain K is unable to synthesize lysine. In the co-culture, if exchange of leucine and lysine occurs then both strains can grow in glucose minimal medium. Panel (B) shows the concentration profiles of glucose (red x) and optical density (black squares) during batch growth of the co-culture. The error bars indicate the standard deviations across replicates. (C) Genomic DNA from the two mutants were extracted from the co-culture at several time points during batch growth of the co-culture and analyzed by qPCR. Blue circles and red diamonds represent the K and L strains, respectively. The error bars were calculated by the error propagation method described in File S2 . (D) The ratio of K to L was calculated from the qPCR results. The K to L ratio measured using qPCR was 1.59±0.18 for a 1∶1 mixture of un-evolved cells based on OD600. The error bars indicate standard deviations.

    Journal: PLoS ONE

    Article Title: Adaptive Evolution of Synthetic Cooperating Communities Improves Growth Performance

    doi: 10.1371/journal.pone.0108297

    Figure Lengend Snippet: The un-evolved co-culture of strains K and L. (A) Nutrient exchange and dependence in co-culture of two E. coli strains L and K. Strain L is incapable of synthesizing leucine, while strain K is unable to synthesize lysine. In the co-culture, if exchange of leucine and lysine occurs then both strains can grow in glucose minimal medium. Panel (B) shows the concentration profiles of glucose (red x) and optical density (black squares) during batch growth of the co-culture. The error bars indicate the standard deviations across replicates. (C) Genomic DNA from the two mutants were extracted from the co-culture at several time points during batch growth of the co-culture and analyzed by qPCR. Blue circles and red diamonds represent the K and L strains, respectively. The error bars were calculated by the error propagation method described in File S2 . (D) The ratio of K to L was calculated from the qPCR results. The K to L ratio measured using qPCR was 1.59±0.18 for a 1∶1 mixture of un-evolved cells based on OD600. The error bars indicate standard deviations.

    Article Snippet: OD600 values were measured in a Tecan microplate reader and the changes in OD600 values and growth rates for the co-culture were calculated.

    Techniques: Co-Culture Assay, Concentration Assay, Real-time Polymerase Chain Reaction

    Computational model predictions of co-culture composition and growth rates. The model was constrained using either amino acid uptake (panels A and C) or release rates (panels B and D). Panels A and B display the predicted K:L ratio at a co-culture OD≈0.2. The color map indicates the value of K:L ratio. Panels C and D show the predicted average growth rate of co-culture, indicated by the color map. The evolutionary trajectory of co-culture 4 is shown on panels A through D, where the open circles indicate passages 1,4,7,10,12,15,19 and 21. The estimated uptake or release rates for evolved K ev and L ev strains in each passage were then used to constrain the model. Panel E compares the model predicted K:L ratio and average growth rate of the co-culture near OD600≈0.2 to the estimated experimental values. Blue diamonds and red triangles denote the predictions when the model was constrained by the estimated uptake rates for co-culture 4 and 6, respectively. Green squares and purple circles denote the predictions when the model was constrained by the estimated release rates for the two co-cultures for co-culture 4 and 6, respectively.

    Journal: PLoS ONE

    Article Title: Adaptive Evolution of Synthetic Cooperating Communities Improves Growth Performance

    doi: 10.1371/journal.pone.0108297

    Figure Lengend Snippet: Computational model predictions of co-culture composition and growth rates. The model was constrained using either amino acid uptake (panels A and C) or release rates (panels B and D). Panels A and B display the predicted K:L ratio at a co-culture OD≈0.2. The color map indicates the value of K:L ratio. Panels C and D show the predicted average growth rate of co-culture, indicated by the color map. The evolutionary trajectory of co-culture 4 is shown on panels A through D, where the open circles indicate passages 1,4,7,10,12,15,19 and 21. The estimated uptake or release rates for evolved K ev and L ev strains in each passage were then used to constrain the model. Panel E compares the model predicted K:L ratio and average growth rate of the co-culture near OD600≈0.2 to the estimated experimental values. Blue diamonds and red triangles denote the predictions when the model was constrained by the estimated uptake rates for co-culture 4 and 6, respectively. Green squares and purple circles denote the predictions when the model was constrained by the estimated release rates for the two co-cultures for co-culture 4 and 6, respectively.

    Article Snippet: OD600 values were measured in a Tecan microplate reader and the changes in OD600 values and growth rates for the co-culture were calculated.

    Techniques: Co-Culture Assay

    Comparisons between un-evolved co-cultures L+K and hybrid co-cultures containing L+K ev or L ev +K. Cells from 10 colonies of K ev (or L ev ) at each selected passage were grown individually in co-culture with the un-evolved partner strain (L or K). The growth rate and change in OD600 for each hybrid co-culture was normalized to the growth rate and change in OD600 of the un-evolved co-culture grown on the same microplate. The resulting growth rate ratios and change in OD600 ratios are shown as blue diamonds (L+K ev ) and red squares (L ev +K), respectively, in panels A and B (isolates from co-culture 4) and panels D and E (isolates from co-culture 6). The error bars indicate the standard deviations based on 10 separate hybrid co-cultures each with four replicates (n = 40). The dashed lines indicate the behavior of the un-evolved co-culture (L+K). Panels C and F shows the K:L ratio in L+K ev and L ev +K in hybrid co-cultures and the un-evolved co-culture. The hybrid co-cultures contained evolved isolates from co-culture 4 (panel C) or co-culture 6 (panel F). The error bars indicate the standard deviations based on hybrid co-cultures using three different isolates and three measurements for each passage (n = 9), see File S2 for details. The shaded bands in C and F show the mean ± the standard deviation for the K:L ratio in the un-evolved co-culture at an OD600 of 0.2 when grown in 96 well plates (1.62±0.14).

    Journal: PLoS ONE

    Article Title: Adaptive Evolution of Synthetic Cooperating Communities Improves Growth Performance

    doi: 10.1371/journal.pone.0108297

    Figure Lengend Snippet: Comparisons between un-evolved co-cultures L+K and hybrid co-cultures containing L+K ev or L ev +K. Cells from 10 colonies of K ev (or L ev ) at each selected passage were grown individually in co-culture with the un-evolved partner strain (L or K). The growth rate and change in OD600 for each hybrid co-culture was normalized to the growth rate and change in OD600 of the un-evolved co-culture grown on the same microplate. The resulting growth rate ratios and change in OD600 ratios are shown as blue diamonds (L+K ev ) and red squares (L ev +K), respectively, in panels A and B (isolates from co-culture 4) and panels D and E (isolates from co-culture 6). The error bars indicate the standard deviations based on 10 separate hybrid co-cultures each with four replicates (n = 40). The dashed lines indicate the behavior of the un-evolved co-culture (L+K). Panels C and F shows the K:L ratio in L+K ev and L ev +K in hybrid co-cultures and the un-evolved co-culture. The hybrid co-cultures contained evolved isolates from co-culture 4 (panel C) or co-culture 6 (panel F). The error bars indicate the standard deviations based on hybrid co-cultures using three different isolates and three measurements for each passage (n = 9), see File S2 for details. The shaded bands in C and F show the mean ± the standard deviation for the K:L ratio in the un-evolved co-culture at an OD600 of 0.2 when grown in 96 well plates (1.62±0.14).

    Article Snippet: OD600 values were measured in a Tecan microplate reader and the changes in OD600 values and growth rates for the co-culture were calculated.

    Techniques: Co-Culture Assay, Standard Deviation

    Biofilm formation of Ea1189, Ea1189Δ hfq , and Ea1189Δ arcZ . (A) Biofilm formation of Ea1189, Ea1189Δ hfq , Ea1189Δ arcZ , and Ea1189Δ arcZ carrying complementation plasmid pMLarcZ on glass cover slips. Bacterial strains were incubated with glass cover slips in static cultures of 0.5X LB broth. The biofilm formed on the cover slips was stained with crystal violet, and quantified by measuring light absorbance at OD600. Asterisks indicate significant difference ( P

    Journal: BMC Genomics

    Article Title: Genome-wide identification of Hfq-regulated small RNAs in the fire blight pathogen Erwinia amylovora discovered small RNAs with virulence regulatory function

    doi: 10.1186/1471-2164-15-414

    Figure Lengend Snippet: Biofilm formation of Ea1189, Ea1189Δ hfq , and Ea1189Δ arcZ . (A) Biofilm formation of Ea1189, Ea1189Δ hfq , Ea1189Δ arcZ , and Ea1189Δ arcZ carrying complementation plasmid pMLarcZ on glass cover slips. Bacterial strains were incubated with glass cover slips in static cultures of 0.5X LB broth. The biofilm formed on the cover slips was stained with crystal violet, and quantified by measuring light absorbance at OD600. Asterisks indicate significant difference ( P

    Article Snippet: The solubilized crystal violet in the elusion solution was quantified by measuring the OD600 absorbance using a Safire microplate reader (Tecan; Research Triangle Park, NC).

    Techniques: Plasmid Preparation, Incubation, Staining

    Representation of OD600 measurements vs NPs number/μL. The data were analysed by GraphPad software according to a linear regression straight line equation. ( A ) Calibration curve of 100 nm NPs. ( B ) Calibration curve of 200 nm NPs. ( C ) Calibration curve of 460 nm NPs. 1OD600 value, line equation and correlation factor (r2) are indicated for each NP size. Nine replicates were measured per NPs concentration.

    Journal: Scientific Reports

    Article Title: Number of Nanoparticles per Cell through a Spectrophotometric Method - A key parameter to Assess Nanoparticle-based Cellular Assays

    doi: 10.1038/srep10091

    Figure Lengend Snippet: Representation of OD600 measurements vs NPs number/μL. The data were analysed by GraphPad software according to a linear regression straight line equation. ( A ) Calibration curve of 100 nm NPs. ( B ) Calibration curve of 200 nm NPs. ( C ) Calibration curve of 460 nm NPs. 1OD600 value, line equation and correlation factor (r2) are indicated for each NP size. Nine replicates were measured per NPs concentration.

    Article Snippet: The optical density at 600 nm (OD600) was recorded three times in three different asssays per suspension using two different spectrophotometers: EppendorfBioPhotometer (with path length of 10 mm cuvettes), and NanoDrop Spectrophotometer ND-1000, Thermo Fisher Scientific (1 mm path length system), OD600 data from NanoDrop Spectrophotometer were corrected by factor (x10) (see ).

    Techniques: Software, Concentration Assay

    a The OD600 values of strains Po1f, Po1f-LN-000, Po1f-LN-004, and Po1f-LN-051 cultured in YPD medium, measured at 0, 8, 14, 24, 32, 48, 72, and 96 h. b Squalene production in strains Po1f, Po1f-LN-000, Po1f-LN-004, Po1f-LN-051 cultured in YPD medium for 5 days

    Journal: Biotechnology for Biofuels

    Article Title: Metabolic engineering of oleaginous yeast Yarrowia lipolytica for limonene overproduction

    doi: 10.1186/s13068-016-0626-7

    Figure Lengend Snippet: a The OD600 values of strains Po1f, Po1f-LN-000, Po1f-LN-004, and Po1f-LN-051 cultured in YPD medium, measured at 0, 8, 14, 24, 32, 48, 72, and 96 h. b Squalene production in strains Po1f, Po1f-LN-000, Po1f-LN-004, Po1f-LN-051 cultured in YPD medium for 5 days

    Article Snippet: Analysis Optical densities at 600 nm (OD600 ) were measured using a Shimadzu UV-1800 spectrophotometer (Shimadzu Co., Kyoto, Japan).

    Techniques: Cell Culture

    Effect of nicotine on initial attachment of S. epidermidis . Primary attachment of the SE1457 to polystyrene surfaces. Overnight cultures grown to an OD600 of 0.6 were diluted with TSB (Control) and TSB supplemented with 4000 μg/ml nicotine (Nicotine) to an OD 600 of 0.1 in PBS and inoculated into a six-well polyethylene plate for 1 h at 37°C. The attached cells were photographed under a light microscope (A) and the cell numbers were counted by Image J software (B) .

    Journal: Frontiers in Microbiology

    Article Title: Nicotine Enhances Staphylococcus epidermidis Biofilm Formation by Altering the Bacterial Autolysis, Extracellular DNA Releasing, and Polysaccharide Intercellular Adhesin Production

    doi: 10.3389/fmicb.2018.02575

    Figure Lengend Snippet: Effect of nicotine on initial attachment of S. epidermidis . Primary attachment of the SE1457 to polystyrene surfaces. Overnight cultures grown to an OD600 of 0.6 were diluted with TSB (Control) and TSB supplemented with 4000 μg/ml nicotine (Nicotine) to an OD 600 of 0.1 in PBS and inoculated into a six-well polyethylene plate for 1 h at 37°C. The attached cells were photographed under a light microscope (A) and the cell numbers were counted by Image J software (B) .

    Article Snippet: The diluted cultures were transferred to the wells of a polystyrene microtiter plate (200 μL/well) and incubated at 37°C for 24 h. The cell density was measured at OD600 using a microtiter plate reader (BioRAD, United States).

    Techniques: Light Microscopy, Software

    Effect of nicotine on extracellular DNA release and bacterial autolysis. (A) The amount of eDNA in SE1457 WT and Δ arlRS strains biofilms with and without 4000 μg/ml nicotine treatment was determined by measuring the fluorescence of PI-bound eDNA with the excitation/emission wavelength at 535/610 nm. Relative amounts of eDNA per OD600 unit were calculated. The experiment was performed in triplicates. ( ∗ P

    Journal: Frontiers in Microbiology

    Article Title: Nicotine Enhances Staphylococcus epidermidis Biofilm Formation by Altering the Bacterial Autolysis, Extracellular DNA Releasing, and Polysaccharide Intercellular Adhesin Production

    doi: 10.3389/fmicb.2018.02575

    Figure Lengend Snippet: Effect of nicotine on extracellular DNA release and bacterial autolysis. (A) The amount of eDNA in SE1457 WT and Δ arlRS strains biofilms with and without 4000 μg/ml nicotine treatment was determined by measuring the fluorescence of PI-bound eDNA with the excitation/emission wavelength at 535/610 nm. Relative amounts of eDNA per OD600 unit were calculated. The experiment was performed in triplicates. ( ∗ P

    Article Snippet: The diluted cultures were transferred to the wells of a polystyrene microtiter plate (200 μL/well) and incubated at 37°C for 24 h. The cell density was measured at OD600 using a microtiter plate reader (BioRAD, United States).

    Techniques: Fluorescence

    SPI-1 induction varies in M9 minimal medium. A, C, E. P prgH activity of the SPI-1 reporter strain (P prgH -gfp [LVA]) from one representative experiment. B, D, F. Comparing LB-M to glucose M9, glucose M9 containing different brands of Casamino acids, and 3 alternative carbon sources in M9, respectively. % promoter activity was calculated from n = 3 at maximum promoter activity of GFP/OD600; normalized to LB-Miller grown bacteria (B and F) or those grown with CAA3 (D). The Casamino acids used are ‘existing’ Bacto (CAA1), Bacto (CAA2), Bacto Technical (CAA3), Sigma (CAA4), Remel (CAA5). Statistical analyses were unpaired t-test with Welch’s correction (B) or a one-way ANOVA with Dunnett’s multiple comparisons test (D, F). Data shown are mean ±SD (* p

    Journal: PLoS ONE

    Article Title: Inherent Variability of Growth Media Impacts the Ability of Salmonella Typhimurium to Interact with Host Cells

    doi: 10.1371/journal.pone.0157043

    Figure Lengend Snippet: SPI-1 induction varies in M9 minimal medium. A, C, E. P prgH activity of the SPI-1 reporter strain (P prgH -gfp [LVA]) from one representative experiment. B, D, F. Comparing LB-M to glucose M9, glucose M9 containing different brands of Casamino acids, and 3 alternative carbon sources in M9, respectively. % promoter activity was calculated from n = 3 at maximum promoter activity of GFP/OD600; normalized to LB-Miller grown bacteria (B and F) or those grown with CAA3 (D). The Casamino acids used are ‘existing’ Bacto (CAA1), Bacto (CAA2), Bacto Technical (CAA3), Sigma (CAA4), Remel (CAA5). Statistical analyses were unpaired t-test with Welch’s correction (B) or a one-way ANOVA with Dunnett’s multiple comparisons test (D, F). Data shown are mean ±SD (* p

    Article Snippet: The OD600 after the 3.5 h subculture was 4.5, 3.1, and 4.2 for US Biological, Difco, and Sigma, respectively.

    Techniques: Activity Assay

    Comparison of bacterial invasiveness after grown in different brands of LB-Miller. A. Invasion assays were conducted on HeLa cells using SL1344 bacteria grown in US Biological (US-B), Difco, or Sigma LB-Miller. Data are % of the inoculum found to be intracellular at 1.5 h post-infection, normalized to US-B invasion from n = 3, showing mean ±SD. Plate reader growth curves (B) and P prgH activity (C) of the SPI-1 reporter strain (P prgH -gfp [LVA]) are shown for one representative experiment (n = 3). D. % promoter activity was calculated from n = 3 of GFP/OD600 at maximum promoter activity; normalized to US-B LB-M grown bacteria. Data are mean ±SD; statistical analyses were done using a one-way ANOVA with Dunnett’s multiple comparisons test (* p

    Journal: PLoS ONE

    Article Title: Inherent Variability of Growth Media Impacts the Ability of Salmonella Typhimurium to Interact with Host Cells

    doi: 10.1371/journal.pone.0157043

    Figure Lengend Snippet: Comparison of bacterial invasiveness after grown in different brands of LB-Miller. A. Invasion assays were conducted on HeLa cells using SL1344 bacteria grown in US Biological (US-B), Difco, or Sigma LB-Miller. Data are % of the inoculum found to be intracellular at 1.5 h post-infection, normalized to US-B invasion from n = 3, showing mean ±SD. Plate reader growth curves (B) and P prgH activity (C) of the SPI-1 reporter strain (P prgH -gfp [LVA]) are shown for one representative experiment (n = 3). D. % promoter activity was calculated from n = 3 of GFP/OD600 at maximum promoter activity; normalized to US-B LB-M grown bacteria. Data are mean ±SD; statistical analyses were done using a one-way ANOVA with Dunnett’s multiple comparisons test (* p

    Article Snippet: The OD600 after the 3.5 h subculture was 4.5, 3.1, and 4.2 for US Biological, Difco, and Sigma, respectively.

    Techniques: Infection, Activity Assay

    Actin mutant alleles affecting the activation of ExoY. ( a ) Drop tests of serial dilutions of S. cerevisiae strains expressing wild-type (wt) actin (SC489), D25Y/D222G (SC690) or D25N (SC691) and p1593 for galactose-induced expression of ExoY or the corresponding vector control YEpGal555. Cell suspensions were normalized to an OD600 of 1.0 and 5-fold serial dilutions were applied as 3 μl drops on SD or SG-agar plates. ( b ) Western blot analysis to verify expression of Myc-ExoY in SC690 and SC691 with anti-Myc or anti-RPS9 (loading control). ( c ) Western blot analysis to verify expression of actin in SC489 (1), SC690 (2) and SC691 (3) with anti-actin or anti-RPS9 (loading control). Uncropped images of ( b , c ) are shown in Supplementary Fig. 10 .

    Journal: Nature Communications

    Article Title: Actin activates Pseudomonas aeruginosa ExoY nucleotidyl cyclase toxin and ExoY-like effector domains from MARTX toxins

    doi: 10.1038/ncomms13582

    Figure Lengend Snippet: Actin mutant alleles affecting the activation of ExoY. ( a ) Drop tests of serial dilutions of S. cerevisiae strains expressing wild-type (wt) actin (SC489), D25Y/D222G (SC690) or D25N (SC691) and p1593 for galactose-induced expression of ExoY or the corresponding vector control YEpGal555. Cell suspensions were normalized to an OD600 of 1.0 and 5-fold serial dilutions were applied as 3 μl drops on SD or SG-agar plates. ( b ) Western blot analysis to verify expression of Myc-ExoY in SC690 and SC691 with anti-Myc or anti-RPS9 (loading control). ( c ) Western blot analysis to verify expression of actin in SC489 (1), SC690 (2) and SC691 (3) with anti-actin or anti-RPS9 (loading control). Uncropped images of ( b , c ) are shown in Supplementary Fig. 10 .

    Article Snippet: Extracts from S. cerevisiae were prepared from 100 ml cultures grown in yeast extract peptone dextrose to an OD600 between 0.5 and 2, at which cells were harvested by centrifugation, washed once with water, and resuspended in 300 μl yeast lysis buffer (50 mM Tris-HCl pH 7.4, 50 mM KCl, 1 mM DTT, complete EDTA-free protease inhibitor cocktail (Roche)).

    Techniques: Mutagenesis, Activation Assay, Expressing, Plasmid Preparation, Western Blot