bl21  (New England Biolabs)


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    BL21 DE3 Competent E coli
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    BL21 DE3 Competent E coli 20x0 05 ml
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    c2527h
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    New England Biolabs bl21
    BL21 DE3 Competent E coli
    BL21 DE3 Competent E coli 20x0 05 ml
    https://www.bioz.com/result/bl21/product/New England Biolabs
    Average 99 stars, based on 551 article reviews
    Price from $9.99 to $1999.99
    bl21 - by Bioz Stars, 2020-09
    99/100 stars

    Images

    1) Product Images from "The traffic ATPase PilF interacts with the inner membrane platform of the DNA translocator and type IV pili from Thermus thermophilus"

    Article Title: The traffic ATPase PilF interacts with the inner membrane platform of the DNA translocator and type IV pili from Thermus thermophilus

    Journal: FEBS Open Bio

    doi: 10.1002/2211-5463.12548

    Purification of PilMNO, PilMN, PilM, and PilN. His 6 ‐PilMNO‐Strep (lane 1), His 6 ‐PilMN‐Strep (lane 2), His 6 ‐PilM (lane 3), and PilN‐Strep (lane 4) were purified from Escherichia coli BL21 (DE3) cells as described in Experimental procedures . PilMNO, PilMN, and PilN were isolated from membranes, whereas PilM was isolated from soluble fractions. To separate His 6 ‐PilM (44 kDa), PilN (23 kDa), and PilO‐Strep (22.5 kDa), 4–20% SDS/PAGE was performed (lane 1). All other protein preparations were separated by 14% SDS/PAGE (A). The identity of the proteins was confirmed by western blot using polyclonal antisera against PilM (B) and PilN (C) or using Strep‐Tactin HRP conjugate to detect PilO‐Strep (D). Dotted lines indicate excised lanes from different but comparable gels or blots.
    Figure Legend Snippet: Purification of PilMNO, PilMN, PilM, and PilN. His 6 ‐PilMNO‐Strep (lane 1), His 6 ‐PilMN‐Strep (lane 2), His 6 ‐PilM (lane 3), and PilN‐Strep (lane 4) were purified from Escherichia coli BL21 (DE3) cells as described in Experimental procedures . PilMNO, PilMN, and PilN were isolated from membranes, whereas PilM was isolated from soluble fractions. To separate His 6 ‐PilM (44 kDa), PilN (23 kDa), and PilO‐Strep (22.5 kDa), 4–20% SDS/PAGE was performed (lane 1). All other protein preparations were separated by 14% SDS/PAGE (A). The identity of the proteins was confirmed by western blot using polyclonal antisera against PilM (B) and PilN (C) or using Strep‐Tactin HRP conjugate to detect PilO‐Strep (D). Dotted lines indicate excised lanes from different but comparable gels or blots.

    Techniques Used: Purification, Isolation, SDS Page, Western Blot

    2) Product Images from "FabV/Triclosan Is an Antibiotic-Free and Cost-Effective Selection System for Efficient Maintenance of High and Medium -Copy Number Plasmids in Escherichia coli"

    Article Title: FabV/Triclosan Is an Antibiotic-Free and Cost-Effective Selection System for Efficient Maintenance of High and Medium -Copy Number Plasmids in Escherichia coli

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0129547

    Plasmid DNA yield. E . coli DH5α and JM109 were transformed with high-copy number pUC19-Bla, pUC19-FabV, pUC19-FabI, and medium-copy number pSA-Hp24-Bla, pSA-Hp24-FabV, pBR322-FabI plasmids and selected on LB agar plates containing 1μM Triclosan (for FabV/FabI plasmids) and or 100μg mL -1 ampicillin (for Bla plasmids). Seed cultures were used to inoculated 5mL LB broth in 50mL flasks and cultured for another 18 hours at 37 or 30°C while shaking at 250rpm. Cell density was then measured by absorbing the diluted samples at 600nm and normalized to 2 OD600. One mL of the normalized culture was then used to extract the plasmid following supplied protocol. The quantity of the extracted plasmid DNA was measured by fluorometry using Qubit Fluorometer and Qubit dsDNA BR Assay Kit (Invitrogen, life technology). Fig 4A shows plasmid DNA yield when bacteria were grown at 37°C, whereas Fig 4B shows the plasmid DNA yield from cultures incubated at 30°C. Error bars show standard deviations calculated from at-least six (6) independent experiments performed in triplicate.
    Figure Legend Snippet: Plasmid DNA yield. E . coli DH5α and JM109 were transformed with high-copy number pUC19-Bla, pUC19-FabV, pUC19-FabI, and medium-copy number pSA-Hp24-Bla, pSA-Hp24-FabV, pBR322-FabI plasmids and selected on LB agar plates containing 1μM Triclosan (for FabV/FabI plasmids) and or 100μg mL -1 ampicillin (for Bla plasmids). Seed cultures were used to inoculated 5mL LB broth in 50mL flasks and cultured for another 18 hours at 37 or 30°C while shaking at 250rpm. Cell density was then measured by absorbing the diluted samples at 600nm and normalized to 2 OD600. One mL of the normalized culture was then used to extract the plasmid following supplied protocol. The quantity of the extracted plasmid DNA was measured by fluorometry using Qubit Fluorometer and Qubit dsDNA BR Assay Kit (Invitrogen, life technology). Fig 4A shows plasmid DNA yield when bacteria were grown at 37°C, whereas Fig 4B shows the plasmid DNA yield from cultures incubated at 30°C. Error bars show standard deviations calculated from at-least six (6) independent experiments performed in triplicate.

    Techniques Used: Plasmid Preparation, Transformation Assay, Cell Culture, Incubation

    Morphology of transformed bacteria. E . coli DH5α and JM109 transformed with (A) high-copy number pUC19-Bla, pUC19-FabV, or pUC19-FabI plasmids or (B) medium–copy number pMXB-p24-Bla, pMXBp24-FabV, or pBR322-FabI plasmids were incubated at 30°C for 18 hours on plates with our without selection agent (Ampicillin or Triclosan) and photographed. No appreciable differences noted when the transformants were plated on Agar plates containing selection agent or not.
    Figure Legend Snippet: Morphology of transformed bacteria. E . coli DH5α and JM109 transformed with (A) high-copy number pUC19-Bla, pUC19-FabV, or pUC19-FabI plasmids or (B) medium–copy number pMXB-p24-Bla, pMXBp24-FabV, or pBR322-FabI plasmids were incubated at 30°C for 18 hours on plates with our without selection agent (Ampicillin or Triclosan) and photographed. No appreciable differences noted when the transformants were plated on Agar plates containing selection agent or not.

    Techniques Used: Transformation Assay, Incubation, Selection

    Bacterial growth characteristics. E . coli DH5α, JM109, and BL21(DE3) were transformed with FabV (pUC19-FabV and pSA-HP24-FabV), FabI (pUC19-FabI, pBR322-FabI), or Bla (pUC19–Bla, pSA-HP24-Bla)-plasmids, and the transformants were selected on LB agar plates. Seed cultures were used to inoculate 25mL LB broth in 250mL baffled flasks and cultures grown at 37, 30, and 22°C for up to 12 hours while shaking at 250rpm. Samples were collected at one hour interval and growth was measured by absorbing the diluted samples at 600nm and graphs plotted. Error bars show standard deviations calculated from at-least six (6) independent experiments performed in triplicate.
    Figure Legend Snippet: Bacterial growth characteristics. E . coli DH5α, JM109, and BL21(DE3) were transformed with FabV (pUC19-FabV and pSA-HP24-FabV), FabI (pUC19-FabI, pBR322-FabI), or Bla (pUC19–Bla, pSA-HP24-Bla)-plasmids, and the transformants were selected on LB agar plates. Seed cultures were used to inoculate 25mL LB broth in 250mL baffled flasks and cultures grown at 37, 30, and 22°C for up to 12 hours while shaking at 250rpm. Samples were collected at one hour interval and growth was measured by absorbing the diluted samples at 600nm and graphs plotted. Error bars show standard deviations calculated from at-least six (6) independent experiments performed in triplicate.

    Techniques Used: Transformation Assay

    Bacterial transformation efficiency. Chemically competent E . coli DH5α and JM109 cells were transformed with 100pg μL -1 of purified high-copy number pUC19-Bla, pUC19-FabV, pUC19-FabI, and medium-copy number pSA-Hp24-Bla, pSA-Hp24-FabV, pBR322-FabI plasmids and selected on LB agar plates containing 1μM Triclosan (for FabV/FabI plasmids) and or 100μg mL -1 ampicillin (for Bla plasmids) after 18 hours of incubation at 30°C and transformation efficiency calculated. Fig 5A and B show transformation efficiency of various plasmids in DH5α and JM109 respectively. Fig 5C and D show the effect of incubation time prior to plating the transformants on selection plates. Error bars show standard deviations calculated from at-least six (6) independent experiments performed in triplicate.
    Figure Legend Snippet: Bacterial transformation efficiency. Chemically competent E . coli DH5α and JM109 cells were transformed with 100pg μL -1 of purified high-copy number pUC19-Bla, pUC19-FabV, pUC19-FabI, and medium-copy number pSA-Hp24-Bla, pSA-Hp24-FabV, pBR322-FabI plasmids and selected on LB agar plates containing 1μM Triclosan (for FabV/FabI plasmids) and or 100μg mL -1 ampicillin (for Bla plasmids) after 18 hours of incubation at 30°C and transformation efficiency calculated. Fig 5A and B show transformation efficiency of various plasmids in DH5α and JM109 respectively. Fig 5C and D show the effect of incubation time prior to plating the transformants on selection plates. Error bars show standard deviations calculated from at-least six (6) independent experiments performed in triplicate.

    Techniques Used: Electroporation Bacterial Transformation, Transformation Assay, Purification, Incubation, Selection

    3) Product Images from "FabV/Triclosan Is an Antibiotic-Free and Cost-Effective Selection System for Efficient Maintenance of High and Medium -Copy Number Plasmids in Escherichia coli"

    Article Title: FabV/Triclosan Is an Antibiotic-Free and Cost-Effective Selection System for Efficient Maintenance of High and Medium -Copy Number Plasmids in Escherichia coli

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0129547

    Plasmid DNA yield. E . coli DH5α and JM109 were transformed with high-copy number pUC19-Bla, pUC19-FabV, pUC19-FabI, and medium-copy number pSA-Hp24-Bla, pSA-Hp24-FabV, pBR322-FabI plasmids and selected on LB agar plates containing 1μM Triclosan (for FabV/FabI plasmids) and or 100μg mL -1 ampicillin (for Bla plasmids). Seed cultures were used to inoculated 5mL LB broth in 50mL flasks and cultured for another 18 hours at 37 or 30°C while shaking at 250rpm. Cell density was then measured by absorbing the diluted samples at 600nm and normalized to 2 OD600. One mL of the normalized culture was then used to extract the plasmid following supplied protocol. The quantity of the extracted plasmid DNA was measured by fluorometry using Qubit Fluorometer and Qubit dsDNA BR Assay Kit (Invitrogen, life technology). Fig 4A shows plasmid DNA yield when bacteria were grown at 37°C, whereas Fig 4B shows the plasmid DNA yield from cultures incubated at 30°C. Error bars show standard deviations calculated from at-least six (6) independent experiments performed in triplicate.
    Figure Legend Snippet: Plasmid DNA yield. E . coli DH5α and JM109 were transformed with high-copy number pUC19-Bla, pUC19-FabV, pUC19-FabI, and medium-copy number pSA-Hp24-Bla, pSA-Hp24-FabV, pBR322-FabI plasmids and selected on LB agar plates containing 1μM Triclosan (for FabV/FabI plasmids) and or 100μg mL -1 ampicillin (for Bla plasmids). Seed cultures were used to inoculated 5mL LB broth in 50mL flasks and cultured for another 18 hours at 37 or 30°C while shaking at 250rpm. Cell density was then measured by absorbing the diluted samples at 600nm and normalized to 2 OD600. One mL of the normalized culture was then used to extract the plasmid following supplied protocol. The quantity of the extracted plasmid DNA was measured by fluorometry using Qubit Fluorometer and Qubit dsDNA BR Assay Kit (Invitrogen, life technology). Fig 4A shows plasmid DNA yield when bacteria were grown at 37°C, whereas Fig 4B shows the plasmid DNA yield from cultures incubated at 30°C. Error bars show standard deviations calculated from at-least six (6) independent experiments performed in triplicate.

    Techniques Used: Plasmid Preparation, Transformation Assay, Cell Culture, Incubation

    Morphology of transformed bacteria. E . coli DH5α and JM109 transformed with (A) high-copy number pUC19-Bla, pUC19-FabV, or pUC19-FabI plasmids or (B) medium–copy number pMXB-p24-Bla, pMXBp24-FabV, or pBR322-FabI plasmids were incubated at 30°C for 18 hours on plates with our without selection agent (Ampicillin or Triclosan) and photographed. No appreciable differences noted when the transformants were plated on Agar plates containing selection agent or not.
    Figure Legend Snippet: Morphology of transformed bacteria. E . coli DH5α and JM109 transformed with (A) high-copy number pUC19-Bla, pUC19-FabV, or pUC19-FabI plasmids or (B) medium–copy number pMXB-p24-Bla, pMXBp24-FabV, or pBR322-FabI plasmids were incubated at 30°C for 18 hours on plates with our without selection agent (Ampicillin or Triclosan) and photographed. No appreciable differences noted when the transformants were plated on Agar plates containing selection agent or not.

    Techniques Used: Transformation Assay, Incubation, Selection

    Bacterial growth characteristics. E . coli DH5α, JM109, and BL21(DE3) were transformed with FabV (pUC19-FabV and pSA-HP24-FabV), FabI (pUC19-FabI, pBR322-FabI), or Bla (pUC19–Bla, pSA-HP24-Bla)-plasmids, and the transformants were selected on LB agar plates. Seed cultures were used to inoculate 25mL LB broth in 250mL baffled flasks and cultures grown at 37, 30, and 22°C for up to 12 hours while shaking at 250rpm. Samples were collected at one hour interval and growth was measured by absorbing the diluted samples at 600nm and graphs plotted. Error bars show standard deviations calculated from at-least six (6) independent experiments performed in triplicate.
    Figure Legend Snippet: Bacterial growth characteristics. E . coli DH5α, JM109, and BL21(DE3) were transformed with FabV (pUC19-FabV and pSA-HP24-FabV), FabI (pUC19-FabI, pBR322-FabI), or Bla (pUC19–Bla, pSA-HP24-Bla)-plasmids, and the transformants were selected on LB agar plates. Seed cultures were used to inoculate 25mL LB broth in 250mL baffled flasks and cultures grown at 37, 30, and 22°C for up to 12 hours while shaking at 250rpm. Samples were collected at one hour interval and growth was measured by absorbing the diluted samples at 600nm and graphs plotted. Error bars show standard deviations calculated from at-least six (6) independent experiments performed in triplicate.

    Techniques Used: Transformation Assay

    Bacterial transformation efficiency. Chemically competent E . coli DH5α and JM109 cells were transformed with 100pg μL -1 of purified high-copy number pUC19-Bla, pUC19-FabV, pUC19-FabI, and medium-copy number pSA-Hp24-Bla, pSA-Hp24-FabV, pBR322-FabI plasmids and selected on LB agar plates containing 1μM Triclosan (for FabV/FabI plasmids) and or 100μg mL -1 ampicillin (for Bla plasmids) after 18 hours of incubation at 30°C and transformation efficiency calculated. Fig 5A and B show transformation efficiency of various plasmids in DH5α and JM109 respectively. Fig 5C and D show the effect of incubation time prior to plating the transformants on selection plates. Error bars show standard deviations calculated from at-least six (6) independent experiments performed in triplicate.
    Figure Legend Snippet: Bacterial transformation efficiency. Chemically competent E . coli DH5α and JM109 cells were transformed with 100pg μL -1 of purified high-copy number pUC19-Bla, pUC19-FabV, pUC19-FabI, and medium-copy number pSA-Hp24-Bla, pSA-Hp24-FabV, pBR322-FabI plasmids and selected on LB agar plates containing 1μM Triclosan (for FabV/FabI plasmids) and or 100μg mL -1 ampicillin (for Bla plasmids) after 18 hours of incubation at 30°C and transformation efficiency calculated. Fig 5A and B show transformation efficiency of various plasmids in DH5α and JM109 respectively. Fig 5C and D show the effect of incubation time prior to plating the transformants on selection plates. Error bars show standard deviations calculated from at-least six (6) independent experiments performed in triplicate.

    Techniques Used: Electroporation Bacterial Transformation, Transformation Assay, Purification, Incubation, Selection

    4) Product Images from "Identification of the agr Peptide of Listeria monocytogenes"

    Article Title: Identification of the agr Peptide of Listeria monocytogenes

    Journal: Frontiers in Microbiology

    doi: 10.3389/fmicb.2016.00989

    (A) Extracted ion chromatograms performed on culture supernatant of Escherichia coli BL21 DE3 pET29a_ agrBD after induction with IPTG (BD+; upper panel) and the synthetic peptide R5T0 (lower panel; R5T0 molecular formula: C 31 H 41 N 5 O 5 S 2 ). (B) Mass spectrometry fragmentation spectra for chromatographic peaks with retention times of 61.2 and 63.9 min (marked with a box in A ). (C,D) Structure and assignment of fragments detected in MS/MS spectra to fragments of R5T0.
    Figure Legend Snippet: (A) Extracted ion chromatograms performed on culture supernatant of Escherichia coli BL21 DE3 pET29a_ agrBD after induction with IPTG (BD+; upper panel) and the synthetic peptide R5T0 (lower panel; R5T0 molecular formula: C 31 H 41 N 5 O 5 S 2 ). (B) Mass spectrometry fragmentation spectra for chromatographic peaks with retention times of 61.2 and 63.9 min (marked with a box in A ). (C,D) Structure and assignment of fragments detected in MS/MS spectra to fragments of R5T0.

    Techniques Used: Mass Spectrometry

    5) Product Images from "Investigation of the methylerythritol 4-phosphate pathway for microbial terpenoid production through metabolic control analysis"

    Article Title: Investigation of the methylerythritol 4-phosphate pathway for microbial terpenoid production through metabolic control analysis

    Journal: Microbial Cell Factories

    doi: 10.1186/s12934-019-1235-5

    Specific efflux rates of MEP pathway intermediates calculated from their measured concentrations in the supernatant of cultures of the E. coli Bl21 dxs RBS library expressing isp S and idi from pCOLA::IspS-idi ( \documentclass[12pt]{minimal} \usepackage{amsmath} \usepackage{wasysym} \usepackage{amsfonts} \usepackage{amssymb} \usepackage{amsbsy} \usepackage{mathrsfs} \usepackage{upgreek} \setlength{\oddsidemargin}{-69pt} \begin{document}$$\overline{\varvec{x}}$$\end{document} x ¯ ± SD, n = 3). The cultures were induced at OD 600 = 0.1 and the supernatant was sampled at OD 600 ≈ 0.5. The export of MEP pathway intermediates occurred after the induction of isp S expression in the dxs expression library. a Metabolite concentration in the supernatant of the cultures. b DXP and MEcPP efflux rates as a function of dxs expression relative to wild-type levels. Linear fits through zero and second-degree polynomial fits through zero are shown for DXP and MEcPP, respectively in ( a ) and ( b ). c Logarithmic plot of MEcPP efflux rate as a function of dxs expression. The data were fitted to a linear function with a slope of 2.0. The red marked data point was excluded from the fit. d The correlation of MEcPP efflux rate with its intracellular concentration, linear fitted through zero. e Combined efflux of DXP, MEcPP and isoprene in dependents of dxs expression, linear fitted
    Figure Legend Snippet: Specific efflux rates of MEP pathway intermediates calculated from their measured concentrations in the supernatant of cultures of the E. coli Bl21 dxs RBS library expressing isp S and idi from pCOLA::IspS-idi ( \documentclass[12pt]{minimal} \usepackage{amsmath} \usepackage{wasysym} \usepackage{amsfonts} \usepackage{amssymb} \usepackage{amsbsy} \usepackage{mathrsfs} \usepackage{upgreek} \setlength{\oddsidemargin}{-69pt} \begin{document}$$\overline{\varvec{x}}$$\end{document} x ¯ ± SD, n = 3). The cultures were induced at OD 600 = 0.1 and the supernatant was sampled at OD 600 ≈ 0.5. The export of MEP pathway intermediates occurred after the induction of isp S expression in the dxs expression library. a Metabolite concentration in the supernatant of the cultures. b DXP and MEcPP efflux rates as a function of dxs expression relative to wild-type levels. Linear fits through zero and second-degree polynomial fits through zero are shown for DXP and MEcPP, respectively in ( a ) and ( b ). c Logarithmic plot of MEcPP efflux rate as a function of dxs expression. The data were fitted to a linear function with a slope of 2.0. The red marked data point was excluded from the fit. d The correlation of MEcPP efflux rate with its intracellular concentration, linear fitted through zero. e Combined efflux of DXP, MEcPP and isoprene in dependents of dxs expression, linear fitted

    Techniques Used: Expressing, Concentration Assay

    6) Product Images from "Pyocin S5 import into Pseudomonas aeruginosa reveals a generic mode of bacteriocin transport"

    Article Title: Pyocin S5 import into Pseudomonas aeruginosa reveals a generic mode of bacteriocin transport

    Journal: bioRxiv

    doi: 10.1101/856047

    FptA and TonB1 constitute the minimal system for PyoS5-susceptibility in E. coli . (A) Susceptibility to PyoS5 (3 and 60 μM) was assessed for P. aeruginosa YHP17, (B) for E. coli BL21 (DE3) expressing FptA, (C) for E. coli BL21 (DE3) expressing TonB-B1, and (D) for E. coli BL21 (DE3) expressing FptA and TonB-B1. Zones of clearance are observed in all E. coli samples for the ColBPyoS5 (13 μM) control (B-D) and for both concentrations of PyoS5 in E. coli expressing FptA and TonB-B1 (D). In the P. aeruginosa control clearance zones were observed for PyoS5 (A).
    Figure Legend Snippet: FptA and TonB1 constitute the minimal system for PyoS5-susceptibility in E. coli . (A) Susceptibility to PyoS5 (3 and 60 μM) was assessed for P. aeruginosa YHP17, (B) for E. coli BL21 (DE3) expressing FptA, (C) for E. coli BL21 (DE3) expressing TonB-B1, and (D) for E. coli BL21 (DE3) expressing FptA and TonB-B1. Zones of clearance are observed in all E. coli samples for the ColBPyoS5 (13 μM) control (B-D) and for both concentrations of PyoS5 in E. coli expressing FptA and TonB-B1 (D). In the P. aeruginosa control clearance zones were observed for PyoS5 (A).

    Techniques Used: Expressing

    FpvAI and TonB1 constitute the minimal system for PyoS2- and PyoS4-susceptibility. Susceptibility to 1 μM PyoS2, PyoS4, PyoS5 and ColBPyoS5 was assessed for (A) P. aeruginosa YHP17, (B) E. coli BL21 (DE3) expressing FpvAI, (C) E. coli BL21(DE3) expressing TonB-B1, and (D) E. coli BL21 (DE3) expressing FpvAI and TonB-B1. Zones of clearance were observed for all E. coli conditions tested when ColBPyoS5 was applied (B-D). Similarly, E. coli expressing FpvAI and TonB-B1 was susceptible to both PyoS2 and PyoS4, as was the P. aeruginosa control (A+D). Zones of clearance for PyoS5 were only observed in P. aeruginosa .
    Figure Legend Snippet: FpvAI and TonB1 constitute the minimal system for PyoS2- and PyoS4-susceptibility. Susceptibility to 1 μM PyoS2, PyoS4, PyoS5 and ColBPyoS5 was assessed for (A) P. aeruginosa YHP17, (B) E. coli BL21 (DE3) expressing FpvAI, (C) E. coli BL21(DE3) expressing TonB-B1, and (D) E. coli BL21 (DE3) expressing FpvAI and TonB-B1. Zones of clearance were observed for all E. coli conditions tested when ColBPyoS5 was applied (B-D). Similarly, E. coli expressing FpvAI and TonB-B1 was susceptible to both PyoS2 and PyoS4, as was the P. aeruginosa control (A+D). Zones of clearance for PyoS5 were only observed in P. aeruginosa .

    Techniques Used: Expressing

    Related Articles

    Clone Assay:

    Article Title: Investigation of the methylerythritol 4-phosphate pathway for microbial terpenoid production through metabolic control analysis
    Article Snippet: .. Bacterial strains and culture conditions Standard cloning and metabolic engineering were carried out using E. coli strains 10β and BL21 (DE3), respectively (both supplied by New England Biolabs, Ipswich, MA, USA). .. For general cloning, the bacteria were cultivated in lysogenic broth (LB) medium, whereas for all other experiments the bacteria were cultivated in M9 medium supplemented with 0.5% (w/v) glucose and appropriate antibiotics (50 μg mL−1 kanamycin, 100 μg mL−1 ampicillin and/or 25 μg mL−1 chloramphenicol) at 37 °C [ ].

    Cell Culture:

    Article Title: Cloning and Characterization of a Novel Drosophila Stress Induced DNase
    Article Snippet: .. Expression of the SID protein in bacteria After verifying the sequence of the pCold-sid clone, the vector was transformed into BL21 (DE3) pLysS competent cells (NEB, Ipswich, MA) and a single colony was cultured in LB containing 100 µg/ml ampicillin at 37°C overnight. ..

    Purification:

    Article Title: Targeted Isolation of Antibodies Directed against Major Sites of SIV Env Vulnerability
    Article Snippet: .. Briefly, CVN was expressed in the BL21-DE3 E . coli strain (New England Biolabs), followed by purification using reversed-phase chromatograpy (Sep-Pak Vac 35cc (10g) tC18 cartridges, Waters) and gel-filtration (Superdex 75, GE Healthcare) to ensure separation of monomeric and domain-swapped dimeric CVN. .. Targeted isolation of SIV-specific B cells by fluorescence activated cell sorting (FACS) Cryopreserved PBMC were thawed and stained with LIVE/DEAD Fixable Violet Dead Cell Stain (Life Technologies) as previously described [ , ].

    Article Title: Nse1, Nse2, and a Novel Subunit of the Smc5-Smc6 Complex, Nse3, Play a Crucial Role in Meiosis
    Article Snippet: .. MBP-Nse3 fusion protein was expressed in BL21 (DE3), purified on amylose resin (NEB) and used to inoculate rabbits. .. The resulting sera were affinity purified against the MBP-Nse3 protein and used at a 1:400 dilution for Western blotting.

    other:

    Article Title: The traffic ATPase PilF interacts with the inner membrane platform of the DNA translocator and type IV pili from Thermus thermophilus
    Article Snippet: Organisms and cultivation Escherichia coli DH5α, BL21 (DE3) and T7 Express cells harboring pET28a plasmids (New England Biolabs, Ipswich, MA, USA) were grown at 37 °C in LB medium or 2 × YT medium containing 20 μg·mL−1 kanamycin.

    Expressing:

    Article Title: Cloning and Characterization of a Novel Drosophila Stress Induced DNase
    Article Snippet: .. Expression of the SID protein in bacteria After verifying the sequence of the pCold-sid clone, the vector was transformed into BL21 (DE3) pLysS competent cells (NEB, Ipswich, MA) and a single colony was cultured in LB containing 100 µg/ml ampicillin at 37°C overnight. ..

    Sequencing:

    Article Title: Cloning and Characterization of a Novel Drosophila Stress Induced DNase
    Article Snippet: .. Expression of the SID protein in bacteria After verifying the sequence of the pCold-sid clone, the vector was transformed into BL21 (DE3) pLysS competent cells (NEB, Ipswich, MA) and a single colony was cultured in LB containing 100 µg/ml ampicillin at 37°C overnight. ..

    Transformation Assay:

    Article Title: Postnatal Chick Choroids Exhibit Increased Retinaldehyde Dehydrogenase Activity During Recovery From Form Deprivation Induced Myopia
    Article Snippet: .. RALDH1, 2, and 3/pET15b plasmids were transformed into BL21 (DE3) Escherichiacoli (New England Biolabs), according to the manufacturer's protocol and plated on LB agarose plates with carbenicillin selectivity (100 μg/mL; Sigma-Aldrich Corp.). ..

    Article Title: Identification of the agr Peptide of Listeria monocytogenes
    Article Snippet: .. AIP Production in E. coli For heterologous AIP production, pET29a_agrB or pET29a_agrBD were transformed into E. coli BL21(DE3) (New England Biolabs) and transformants were selected on LB agar containing kanamycin. ..

    Article Title: Cloning and Characterization of a Novel Drosophila Stress Induced DNase
    Article Snippet: .. Expression of the SID protein in bacteria After verifying the sequence of the pCold-sid clone, the vector was transformed into BL21 (DE3) pLysS competent cells (NEB, Ipswich, MA) and a single colony was cultured in LB containing 100 µg/ml ampicillin at 37°C overnight. ..

    Plasmid Preparation:

    Article Title: Cloning and Characterization of a Novel Drosophila Stress Induced DNase
    Article Snippet: .. Expression of the SID protein in bacteria After verifying the sequence of the pCold-sid clone, the vector was transformed into BL21 (DE3) pLysS competent cells (NEB, Ipswich, MA) and a single colony was cultured in LB containing 100 µg/ml ampicillin at 37°C overnight. ..

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  • 99
    New England Biolabs bl21
    Purification of PilMNO, PilMN, PilM, and PilN. His 6 ‐PilMNO‐Strep (lane 1), His 6 ‐PilMN‐Strep (lane 2), His 6 ‐PilM (lane 3), and PilN‐Strep (lane 4) were purified from Escherichia coli <t>BL21</t> (DE3) cells as described in Experimental procedures . PilMNO, PilMN, and PilN were isolated from membranes, whereas PilM was isolated from soluble fractions. To separate His 6 ‐PilM (44 kDa), PilN (23 kDa), and PilO‐Strep (22.5 kDa), 4–20% SDS/PAGE was performed (lane 1). All other protein preparations were separated by 14% SDS/PAGE (A). The identity of the proteins was confirmed by western blot using polyclonal antisera against PilM (B) and PilN (C) or using Strep‐Tactin HRP conjugate to detect PilO‐Strep (D). Dotted lines indicate excised lanes from different but comparable gels or blots.
    Bl21, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 99/100, based on 235 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Purification of PilMNO, PilMN, PilM, and PilN. His 6 ‐PilMNO‐Strep (lane 1), His 6 ‐PilMN‐Strep (lane 2), His 6 ‐PilM (lane 3), and PilN‐Strep (lane 4) were purified from Escherichia coli BL21 (DE3) cells as described in Experimental procedures . PilMNO, PilMN, and PilN were isolated from membranes, whereas PilM was isolated from soluble fractions. To separate His 6 ‐PilM (44 kDa), PilN (23 kDa), and PilO‐Strep (22.5 kDa), 4–20% SDS/PAGE was performed (lane 1). All other protein preparations were separated by 14% SDS/PAGE (A). The identity of the proteins was confirmed by western blot using polyclonal antisera against PilM (B) and PilN (C) or using Strep‐Tactin HRP conjugate to detect PilO‐Strep (D). Dotted lines indicate excised lanes from different but comparable gels or blots.

    Journal: FEBS Open Bio

    Article Title: The traffic ATPase PilF interacts with the inner membrane platform of the DNA translocator and type IV pili from Thermus thermophilus

    doi: 10.1002/2211-5463.12548

    Figure Lengend Snippet: Purification of PilMNO, PilMN, PilM, and PilN. His 6 ‐PilMNO‐Strep (lane 1), His 6 ‐PilMN‐Strep (lane 2), His 6 ‐PilM (lane 3), and PilN‐Strep (lane 4) were purified from Escherichia coli BL21 (DE3) cells as described in Experimental procedures . PilMNO, PilMN, and PilN were isolated from membranes, whereas PilM was isolated from soluble fractions. To separate His 6 ‐PilM (44 kDa), PilN (23 kDa), and PilO‐Strep (22.5 kDa), 4–20% SDS/PAGE was performed (lane 1). All other protein preparations were separated by 14% SDS/PAGE (A). The identity of the proteins was confirmed by western blot using polyclonal antisera against PilM (B) and PilN (C) or using Strep‐Tactin HRP conjugate to detect PilO‐Strep (D). Dotted lines indicate excised lanes from different but comparable gels or blots.

    Article Snippet: Organisms and cultivation Escherichia coli DH5α, BL21 (DE3) and T7 Express cells harboring pET28a plasmids (New England Biolabs, Ipswich, MA, USA) were grown at 37 °C in LB medium or 2 × YT medium containing 20 μg·mL−1 kanamycin.

    Techniques: Purification, Isolation, SDS Page, Western Blot

    (A) Extracted ion chromatograms performed on culture supernatant of Escherichia coli BL21 DE3 pET29a_ agrBD after induction with IPTG (BD+; upper panel) and the synthetic peptide R5T0 (lower panel; R5T0 molecular formula: C 31 H 41 N 5 O 5 S 2 ). (B) Mass spectrometry fragmentation spectra for chromatographic peaks with retention times of 61.2 and 63.9 min (marked with a box in A ). (C,D) Structure and assignment of fragments detected in MS/MS spectra to fragments of R5T0.

    Journal: Frontiers in Microbiology

    Article Title: Identification of the agr Peptide of Listeria monocytogenes

    doi: 10.3389/fmicb.2016.00989

    Figure Lengend Snippet: (A) Extracted ion chromatograms performed on culture supernatant of Escherichia coli BL21 DE3 pET29a_ agrBD after induction with IPTG (BD+; upper panel) and the synthetic peptide R5T0 (lower panel; R5T0 molecular formula: C 31 H 41 N 5 O 5 S 2 ). (B) Mass spectrometry fragmentation spectra for chromatographic peaks with retention times of 61.2 and 63.9 min (marked with a box in A ). (C,D) Structure and assignment of fragments detected in MS/MS spectra to fragments of R5T0.

    Article Snippet: AIP Production in E. coli For heterologous AIP production, pET29a_agrB or pET29a_agrBD were transformed into E. coli BL21(DE3) (New England Biolabs) and transformants were selected on LB agar containing kanamycin.

    Techniques: Mass Spectrometry

    Specific efflux rates of MEP pathway intermediates calculated from their measured concentrations in the supernatant of cultures of the E. coli Bl21 dxs RBS library expressing isp S and idi from pCOLA::IspS-idi ( \documentclass[12pt]{minimal} \usepackage{amsmath} \usepackage{wasysym} \usepackage{amsfonts} \usepackage{amssymb} \usepackage{amsbsy} \usepackage{mathrsfs} \usepackage{upgreek} \setlength{\oddsidemargin}{-69pt} \begin{document}$$\overline{\varvec{x}}$$\end{document} x ¯ ± SD, n = 3). The cultures were induced at OD 600 = 0.1 and the supernatant was sampled at OD 600 ≈ 0.5. The export of MEP pathway intermediates occurred after the induction of isp S expression in the dxs expression library. a Metabolite concentration in the supernatant of the cultures. b DXP and MEcPP efflux rates as a function of dxs expression relative to wild-type levels. Linear fits through zero and second-degree polynomial fits through zero are shown for DXP and MEcPP, respectively in ( a ) and ( b ). c Logarithmic plot of MEcPP efflux rate as a function of dxs expression. The data were fitted to a linear function with a slope of 2.0. The red marked data point was excluded from the fit. d The correlation of MEcPP efflux rate with its intracellular concentration, linear fitted through zero. e Combined efflux of DXP, MEcPP and isoprene in dependents of dxs expression, linear fitted

    Journal: Microbial Cell Factories

    Article Title: Investigation of the methylerythritol 4-phosphate pathway for microbial terpenoid production through metabolic control analysis

    doi: 10.1186/s12934-019-1235-5

    Figure Lengend Snippet: Specific efflux rates of MEP pathway intermediates calculated from their measured concentrations in the supernatant of cultures of the E. coli Bl21 dxs RBS library expressing isp S and idi from pCOLA::IspS-idi ( \documentclass[12pt]{minimal} \usepackage{amsmath} \usepackage{wasysym} \usepackage{amsfonts} \usepackage{amssymb} \usepackage{amsbsy} \usepackage{mathrsfs} \usepackage{upgreek} \setlength{\oddsidemargin}{-69pt} \begin{document}$$\overline{\varvec{x}}$$\end{document} x ¯ ± SD, n = 3). The cultures were induced at OD 600 = 0.1 and the supernatant was sampled at OD 600 ≈ 0.5. The export of MEP pathway intermediates occurred after the induction of isp S expression in the dxs expression library. a Metabolite concentration in the supernatant of the cultures. b DXP and MEcPP efflux rates as a function of dxs expression relative to wild-type levels. Linear fits through zero and second-degree polynomial fits through zero are shown for DXP and MEcPP, respectively in ( a ) and ( b ). c Logarithmic plot of MEcPP efflux rate as a function of dxs expression. The data were fitted to a linear function with a slope of 2.0. The red marked data point was excluded from the fit. d The correlation of MEcPP efflux rate with its intracellular concentration, linear fitted through zero. e Combined efflux of DXP, MEcPP and isoprene in dependents of dxs expression, linear fitted

    Article Snippet: Bacterial strains and culture conditions Standard cloning and metabolic engineering were carried out using E. coli strains 10β and BL21 (DE3), respectively (both supplied by New England Biolabs, Ipswich, MA, USA).

    Techniques: Expressing, Concentration Assay

    FptA and TonB1 constitute the minimal system for PyoS5-susceptibility in E. coli . (A) Susceptibility to PyoS5 (3 and 60 μM) was assessed for P. aeruginosa YHP17, (B) for E. coli BL21 (DE3) expressing FptA, (C) for E. coli BL21 (DE3) expressing TonB-B1, and (D) for E. coli BL21 (DE3) expressing FptA and TonB-B1. Zones of clearance are observed in all E. coli samples for the ColBPyoS5 (13 μM) control (B-D) and for both concentrations of PyoS5 in E. coli expressing FptA and TonB-B1 (D). In the P. aeruginosa control clearance zones were observed for PyoS5 (A).

    Journal: bioRxiv

    Article Title: Pyocin S5 import into Pseudomonas aeruginosa reveals a generic mode of bacteriocin transport

    doi: 10.1101/856047

    Figure Lengend Snippet: FptA and TonB1 constitute the minimal system for PyoS5-susceptibility in E. coli . (A) Susceptibility to PyoS5 (3 and 60 μM) was assessed for P. aeruginosa YHP17, (B) for E. coli BL21 (DE3) expressing FptA, (C) for E. coli BL21 (DE3) expressing TonB-B1, and (D) for E. coli BL21 (DE3) expressing FptA and TonB-B1. Zones of clearance are observed in all E. coli samples for the ColBPyoS5 (13 μM) control (B-D) and for both concentrations of PyoS5 in E. coli expressing FptA and TonB-B1 (D). In the P. aeruginosa control clearance zones were observed for PyoS5 (A).

    Article Snippet: Chemically competent E. coli NEB5α and BL21 (DE3) were purchased from NEB.

    Techniques: Expressing

    FpvAI and TonB1 constitute the minimal system for PyoS2- and PyoS4-susceptibility. Susceptibility to 1 μM PyoS2, PyoS4, PyoS5 and ColBPyoS5 was assessed for (A) P. aeruginosa YHP17, (B) E. coli BL21 (DE3) expressing FpvAI, (C) E. coli BL21(DE3) expressing TonB-B1, and (D) E. coli BL21 (DE3) expressing FpvAI and TonB-B1. Zones of clearance were observed for all E. coli conditions tested when ColBPyoS5 was applied (B-D). Similarly, E. coli expressing FpvAI and TonB-B1 was susceptible to both PyoS2 and PyoS4, as was the P. aeruginosa control (A+D). Zones of clearance for PyoS5 were only observed in P. aeruginosa .

    Journal: bioRxiv

    Article Title: Pyocin S5 import into Pseudomonas aeruginosa reveals a generic mode of bacteriocin transport

    doi: 10.1101/856047

    Figure Lengend Snippet: FpvAI and TonB1 constitute the minimal system for PyoS2- and PyoS4-susceptibility. Susceptibility to 1 μM PyoS2, PyoS4, PyoS5 and ColBPyoS5 was assessed for (A) P. aeruginosa YHP17, (B) E. coli BL21 (DE3) expressing FpvAI, (C) E. coli BL21(DE3) expressing TonB-B1, and (D) E. coli BL21 (DE3) expressing FpvAI and TonB-B1. Zones of clearance were observed for all E. coli conditions tested when ColBPyoS5 was applied (B-D). Similarly, E. coli expressing FpvAI and TonB-B1 was susceptible to both PyoS2 and PyoS4, as was the P. aeruginosa control (A+D). Zones of clearance for PyoS5 were only observed in P. aeruginosa .

    Article Snippet: Chemically competent E. coli NEB5α and BL21 (DE3) were purchased from NEB.

    Techniques: Expressing