bl21 de3 strain  (Millipore)


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    Structured Review

    Millipore bl21 de3 strain
    Bl21 De3 Strain, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 2 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/bl21 de3 strain/product/Millipore
    Average 99 stars, based on 2 article reviews
    Price from $9.99 to $1999.99
    bl21 de3 strain - by Bioz Stars, 2020-04
    99/100 stars

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    Related Articles

    Clone Assay:

    Article Title: A simple vector system to improve performance and utilisation of recombinant antibodies
    Article Snippet: Paragraph title: Reagents, scFv clones & bacterial strains ... E. coli BL21 (DE3) and Origami 2™ (DE3) cells and the pET26b(+) plasmid were from Novagen (San Diego, CA).

    Article Title: Genomic and molecular characterization of a novel quorum sensing molecule in Bacillus licheniformis
    Article Snippet: The expression strain [E. coli BL21 (DE3)] and E. coli TOP10 were used for cloning/transformations and were selected on LBA supplemented with ampicilin (100 µgml−1 ). .. E. coli BL21 ComX producer strain was cultivated in M9 minimal salts solution (sigma).

    Article Title: The gaf Fimbrial Gene Cluster of Escherichia coli Expresses a Full-Size and a Truncated Soluble Adhesin Protein
    Article Snippet: Plasmid pRR-5 contains the complete gaf gene cluster from E. coli strain IHE11165 on a 7-kb DNA fragment in pACYC184 , and pHUB110 contains a 6-bp in-frame deletion within the coding region of gafD , resulting in G fimbriae lacking GlcNAc-binding capacity ( ). pHUB113 ( ) contains the gafD reading frame cloned into pUC19. .. E. coli strain BL21 λDE3 ( ompT lon ) and expression vector pET-22b(+) were from Novagen Inc. (Madison, Wis.).

    Article Title: Molecular cloning and in-silico characterization of high temperature stress responsive pAPX gene isolated from heat tolerant Indian wheat cv. Raj 3765
    Article Snippet: The positive clones were further screened by colony PCR using gene specific primers of TapAPX . .. The expression study of TapAPX gene in prokaryotic system was done by transforming the pET-28a-TapAPX recombinant plasmid in E. coli BL21 cells (Novagen, USA) using heat shock method [ ].

    Centrifugation:

    Article Title: A new hybrid bacteriocin, Ent35-MccV, displays antimicrobial activity against pathogenic Gram-positive and Gram-negative bacteria
    Article Snippet: 2.3 Gene expression and bacteriocins purification for activity determinations Ent35–MccV was expressed from pMA24 in E. coli BL21 [DE3] (pLysS) grown in LB (Sigma) at 37 °C. .. After 3 h, cells were collected by centrifugation in 10 mM HEPES, pH 7, PMSF 0.5 mM (buffer A), lysed using a French press and centrifuged at 47,000xg , 4 °C for 1 h. The supernatant was heated (10 min, 100 °C) and centrifuged (10 min, 10,000xg ).

    Article Title: Hypoallergenic Der p 1/Der p 2 combination vaccines for immunotherapy of house dust mite allergy
    Article Snippet: Expression plasmids containing the Der p 2/1 constructs were transformed into E coli strain BL21 (DE3; Novagen, Merck). .. Protein expression was performed in 250 mL of liquid culture by means of induction with 0.5 mmol/L isopropyl-β-D-1-thiogalactopyranoside at an OD600 of 0.8 for 4 hours at 37°C, and cells were harvested by means of centrifugation at 4000 g for 15 minutes at 4°C.

    Article Title: A Second Mitochondrial DNA Primase Is Essential for Cell Growth and Kinetoplast Minicircle DNA Replication in Trypanosoma brucei ▿ ▿ †
    Article Snippet: Both plasmids pJH604 and pJH618 were transformed into E. coli BL21 cells and grown in Terrific broth (Sigma-Aldrich) supplemented with 0.5 M diglycine and ampicillin (100 μg/ml) at 37°C to an optical density of 1.4. .. Cells were harvested by centrifugation, lysed in metal chelate affinity chromatography buffer (Novagen) containing 0.1% Triton X-100, and then sonicated on ice.

    Article Title: Purification and Reconstitution of the Connexin43 Carboxyl Terminus attached to the 4th Transmembrane Domain in Detergent Micelles *
    Article Snippet: The E. coli strains BL21(DE3) (Novagen), Rosetta-2(DE3) (Novagen), C41(DE3) (Lucigen, ( )), and C43(DE3) (Lucigen, ( )) were transformed with the TM4-Cx43CT expression plasmid (see above) and then inoculated into 1 L of Luria Bertani medium (LB), enriched minimal media , or ISOGRO media (Sigma-Aldrich). .. The cells were harvested by centrifugation (1,000 × g for 45 min), washed with PBS buffer, and stored at −20°C.

    Synthesized:

    Article Title: Fluorescence Resonance Energy Transfer Assay for High-Throughput Screening of ADAMTS1 Inhibitors
    Article Snippet: Materials E . coli BL21 (DE3), E. coli DH5a, pET32a(+) vector and his-bind purification kit were purchased from Novagen. .. The peptide, Abz-YPLPRNITEGEARGNVILTAK(Dnp)P-OH, was synthesized by GL Biochem Ltd. (Shanghai, China).

    Article Title: Preparation and Antitumor Activity of CS5931, A Novel Polypeptide from Sea Squirt Ciona Savignyi
    Article Snippet: .. Recombinant Expression of CS5931 from E. coli BL21 (DE3) The expression vector, pET28a/CS5931 synthesized by our lab previously [ ], was transfected into E. coli BL21 (DE3) cells (Novagen, Darmstadt, Germany). .. Cells were incubated in low-salt LB medium (containing 100 µg/mL Kanamycin) at 37 °C with shaking at 200 rpm.

    Mouse Assay:

    Article Title: A novel HPV16 E7-affitoxin for targeted therapy of HPV16-induced human cervical cancer
    Article Snippet: ICR mice, weighing 23-27 g, were purchased from the animal experimental center of Wenzhou Medical University, China. .. The pET21a(+) vector and E.coli BL21 (DE3) were purchased from Novagen and ATCC, respectively.

    Construct:

    Article Title: Hypoallergenic Der p 1/Der p 2 combination vaccines for immunotherapy of house dust mite allergy
    Article Snippet: .. Expression plasmids containing the Der p 2/1 constructs were transformed into E coli strain BL21 (DE3; Novagen, Merck). .. Protein expression was performed in 250 mL of liquid culture by means of induction with 0.5 mmol/L isopropyl-β-D-1-thiogalactopyranoside at an OD600 of 0.8 for 4 hours at 37°C, and cells were harvested by means of centrifugation at 4000 g for 15 minutes at 4°C.

    Article Title: Function and Protective Capacity of Treponema pallidum subsp. pallidum Glycerophosphodiester Phosphodiesterase
    Article Snippet: .. Anti-Gpd polyclonal antiserum was raised in a New Zealand White rabbit by immunizing three times with 100 μg of each of the inclusion bodies purified from E. coli BL21 (DE3) pLysS transformed with the Gpd-pET-3a construct, emulsified in the Ribi adjuvant MPL + TDM + CWS (monophosphoryl lipid A + trehalose dicorynomycolate + cell wall skeleton) (Sigma). ..

    Incubation:

    Article Title: Hypoallergenic Der p 1/Der p 2 combination vaccines for immunotherapy of house dust mite allergy
    Article Snippet: Expression plasmids containing the Der p 2/1 constructs were transformed into E coli strain BL21 (DE3; Novagen, Merck). .. Cells were lysed by using 3 freeze/thaw cycles (−70°C/+50°C), DNA was degraded by means of incubation with 1 μg of DNase I for 10 minutes at room temperature, and cell debris was removed by means of centrifugation (10,000 g for 30 minutes at 4°C).

    Article Title: Preparation and Antitumor Activity of CS5931, A Novel Polypeptide from Sea Squirt Ciona Savignyi
    Article Snippet: Recombinant Expression of CS5931 from E. coli BL21 (DE3) The expression vector, pET28a/CS5931 synthesized by our lab previously [ ], was transfected into E. coli BL21 (DE3) cells (Novagen, Darmstadt, Germany). .. Cells were incubated in low-salt LB medium (containing 100 µg/mL Kanamycin) at 37 °C with shaking at 200 rpm.

    Article Title: Purification and Reconstitution of the Connexin43 Carboxyl Terminus attached to the 4th Transmembrane Domain in Detergent Micelles *
    Article Snippet: The E. coli strains BL21(DE3) (Novagen), Rosetta-2(DE3) (Novagen), C41(DE3) (Lucigen, ( )), and C43(DE3) (Lucigen, ( )) were transformed with the TM4-Cx43CT expression plasmid (see above) and then inoculated into 1 L of Luria Bertani medium (LB), enriched minimal media , or ISOGRO media (Sigma-Aldrich). .. Cultures were incubated at 37°C with continuous agitation.

    Activity Assay:

    Article Title: A new hybrid bacteriocin, Ent35-MccV, displays antimicrobial activity against pathogenic Gram-positive and Gram-negative bacteria
    Article Snippet: .. 2.3 Gene expression and bacteriocins purification for activity determinations Ent35–MccV was expressed from pMA24 in E. coli BL21 [DE3] (pLysS) grown in LB (Sigma) at 37 °C. ..

    Cell Culture:

    Article Title: A novel HPV16 E7-affitoxin for targeted therapy of HPV16-induced human cervical cancer
    Article Snippet: SiHa (ATCC: HTB-35, HPV16 positive, contains about one to two copies of integrated HPV16 genome), CaSki (ATCC: CRL-1550, HPV16 positive, contains about 600 copies of integrated HPV16 genome), HeLa 229 (ATCC: CCL-2.1, HPV18 positive, used as HPV16 negative control cell line), and melanoma tumor A375 (ATCC: CRL-1619, used as HPV negative control cell line) were obtained from the American Type Culture Collection (ATCC, USA) and cultured as previously described . .. The pET21a(+) vector and E.coli BL21 (DE3) were purchased from Novagen and ATCC, respectively.

    Expressing:

    Article Title: A new hybrid bacteriocin, Ent35-MccV, displays antimicrobial activity against pathogenic Gram-positive and Gram-negative bacteria
    Article Snippet: .. 2.3 Gene expression and bacteriocins purification for activity determinations Ent35–MccV was expressed from pMA24 in E. coli BL21 [DE3] (pLysS) grown in LB (Sigma) at 37 °C. ..

    Article Title: Hypoallergenic Der p 1/Der p 2 combination vaccines for immunotherapy of house dust mite allergy
    Article Snippet: .. Expression plasmids containing the Der p 2/1 constructs were transformed into E coli strain BL21 (DE3; Novagen, Merck). .. Protein expression was performed in 250 mL of liquid culture by means of induction with 0.5 mmol/L isopropyl-β-D-1-thiogalactopyranoside at an OD600 of 0.8 for 4 hours at 37°C, and cells were harvested by means of centrifugation at 4000 g for 15 minutes at 4°C.

    Article Title: Chemical complexity of protein determines optimal E. coli expression host; A comparative study using Erythropoietin, Streptokinase and Tumor Necrosis Factor Receptor
    Article Snippet: The bacterial strains used to study expression were E. coli GJ1158, E. coli BL21 (DE3), E. coli BL21 (DE3) pLys S, E. coli BL21 Rosetta (DE3) pLys S. E. coli GJ1158 , derived from E. coli B strain BL21, is a salt inducible strain with a pro U promoter ( ). .. E. coli BL21 (DE3), E. coli BL21 (DE3) pLys S, E. coli BL21 Rosetta (DE3) pLys S (Novagen,) are IPTG inducible strains in which the T7 RNA Polymerase gene is under the control of a Lac UV5 promoter.

    Article Title: Genomic and molecular characterization of a novel quorum sensing molecule in Bacillus licheniformis
    Article Snippet: The expression strain [E. coli BL21 (DE3)] and E. coli TOP10 were used for cloning/transformations and were selected on LBA supplemented with ampicilin (100 µgml−1 ). .. E. coli BL21 ComX producer strain was cultivated in M9 minimal salts solution (sigma).

    Article Title: The gaf Fimbrial Gene Cluster of Escherichia coli Expresses a Full-Size and a Truncated Soluble Adhesin Protein
    Article Snippet: .. E. coli strain BL21 λDE3 ( ompT lon ) and expression vector pET-22b(+) were from Novagen Inc. (Madison, Wis.). .. The bacteria were cultivated at 37°C in Luria broth ( ) containing the appropriate antibiotics.

    Article Title: Molecular cloning and in-silico characterization of high temperature stress responsive pAPX gene isolated from heat tolerant Indian wheat cv. Raj 3765
    Article Snippet: .. The expression study of TapAPX gene in prokaryotic system was done by transforming the pET-28a-TapAPX recombinant plasmid in E. coli BL21 cells (Novagen, USA) using heat shock method [ ]. ..

    Article Title: Preparation and Antitumor Activity of CS5931, A Novel Polypeptide from Sea Squirt Ciona Savignyi
    Article Snippet: .. Recombinant Expression of CS5931 from E. coli BL21 (DE3) The expression vector, pET28a/CS5931 synthesized by our lab previously [ ], was transfected into E. coli BL21 (DE3) cells (Novagen, Darmstadt, Germany). .. Cells were incubated in low-salt LB medium (containing 100 µg/mL Kanamycin) at 37 °C with shaking at 200 rpm.

    Article Title: Purification and Reconstitution of the Connexin43 Carboxyl Terminus attached to the 4th Transmembrane Domain in Detergent Micelles *
    Article Snippet: .. The E. coli strains BL21(DE3) (Novagen), Rosetta-2(DE3) (Novagen), C41(DE3) (Lucigen, ( )), and C43(DE3) (Lucigen, ( )) were transformed with the TM4-Cx43CT expression plasmid (see above) and then inoculated into 1 L of Luria Bertani medium (LB), enriched minimal media , or ISOGRO media (Sigma-Aldrich). .. Cultures were incubated at 37°C with continuous agitation.

    Transformation Assay:

    Article Title: Hypoallergenic Der p 1/Der p 2 combination vaccines for immunotherapy of house dust mite allergy
    Article Snippet: .. Expression plasmids containing the Der p 2/1 constructs were transformed into E coli strain BL21 (DE3; Novagen, Merck). .. Protein expression was performed in 250 mL of liquid culture by means of induction with 0.5 mmol/L isopropyl-β-D-1-thiogalactopyranoside at an OD600 of 0.8 for 4 hours at 37°C, and cells were harvested by means of centrifugation at 4000 g for 15 minutes at 4°C.

    Article Title: Chemical complexity of protein determines optimal E. coli expression host; A comparative study using Erythropoietin, Streptokinase and Tumor Necrosis Factor Receptor
    Article Snippet: 2.1 Bacterial strains and plasmids For initial transformation of the ligation mixtures, the maintenance strain E. coli DH5α was used. .. E. coli BL21 (DE3), E. coli BL21 (DE3) pLys S, E. coli BL21 Rosetta (DE3) pLys S (Novagen,) are IPTG inducible strains in which the T7 RNA Polymerase gene is under the control of a Lac UV5 promoter.

    Article Title: Function and Protective Capacity of Treponema pallidum subsp. pallidum Glycerophosphodiester Phosphodiesterase
    Article Snippet: .. Anti-Gpd polyclonal antiserum was raised in a New Zealand White rabbit by immunizing three times with 100 μg of each of the inclusion bodies purified from E. coli BL21 (DE3) pLysS transformed with the Gpd-pET-3a construct, emulsified in the Ribi adjuvant MPL + TDM + CWS (monophosphoryl lipid A + trehalose dicorynomycolate + cell wall skeleton) (Sigma). ..

    Article Title: Molecular cloning and in-silico characterization of high temperature stress responsive pAPX gene isolated from heat tolerant Indian wheat cv. Raj 3765
    Article Snippet: The digested PCR product was cloned in pET-28a vector using T4 DNA ligase, the ligated product was transformed in E. coli DH5α and recombinant clones were selected on LA plates supplemented with antibiotic Kanamycin 30 μg/ml. .. The expression study of TapAPX gene in prokaryotic system was done by transforming the pET-28a-TapAPX recombinant plasmid in E. coli BL21 cells (Novagen, USA) using heat shock method [ ].

    Article Title: A Second Mitochondrial DNA Primase Is Essential for Cell Growth and Kinetoplast Minicircle DNA Replication in Trypanosoma brucei ▿ ▿ †
    Article Snippet: .. Both plasmids pJH604 and pJH618 were transformed into E. coli BL21 cells and grown in Terrific broth (Sigma-Aldrich) supplemented with 0.5 M diglycine and ampicillin (100 μg/ml) at 37°C to an optical density of 1.4. ..

    Article Title: Purification and Reconstitution of the Connexin43 Carboxyl Terminus attached to the 4th Transmembrane Domain in Detergent Micelles *
    Article Snippet: .. The E. coli strains BL21(DE3) (Novagen), Rosetta-2(DE3) (Novagen), C41(DE3) (Lucigen, ( )), and C43(DE3) (Lucigen, ( )) were transformed with the TM4-Cx43CT expression plasmid (see above) and then inoculated into 1 L of Luria Bertani medium (LB), enriched minimal media , or ISOGRO media (Sigma-Aldrich). .. Cultures were incubated at 37°C with continuous agitation.

    Derivative Assay:

    Article Title: Chemical complexity of protein determines optimal E. coli expression host; A comparative study using Erythropoietin, Streptokinase and Tumor Necrosis Factor Receptor
    Article Snippet: The bacterial strains used to study expression were E. coli GJ1158, E. coli BL21 (DE3), E. coli BL21 (DE3) pLys S, E. coli BL21 Rosetta (DE3) pLys S. E. coli GJ1158 , derived from E. coli B strain BL21, is a salt inducible strain with a pro U promoter ( ). .. E. coli BL21 (DE3), E. coli BL21 (DE3) pLys S, E. coli BL21 Rosetta (DE3) pLys S (Novagen,) are IPTG inducible strains in which the T7 RNA Polymerase gene is under the control of a Lac UV5 promoter.

    Transfection:

    Article Title: Preparation and Antitumor Activity of CS5931, A Novel Polypeptide from Sea Squirt Ciona Savignyi
    Article Snippet: .. Recombinant Expression of CS5931 from E. coli BL21 (DE3) The expression vector, pET28a/CS5931 synthesized by our lab previously [ ], was transfected into E. coli BL21 (DE3) cells (Novagen, Darmstadt, Germany). .. Cells were incubated in low-salt LB medium (containing 100 µg/mL Kanamycin) at 37 °C with shaking at 200 rpm.

    Ligation:

    Article Title: Chemical complexity of protein determines optimal E. coli expression host; A comparative study using Erythropoietin, Streptokinase and Tumor Necrosis Factor Receptor
    Article Snippet: 2.1 Bacterial strains and plasmids For initial transformation of the ligation mixtures, the maintenance strain E. coli DH5α was used. .. E. coli BL21 (DE3), E. coli BL21 (DE3) pLys S, E. coli BL21 Rosetta (DE3) pLys S (Novagen,) are IPTG inducible strains in which the T7 RNA Polymerase gene is under the control of a Lac UV5 promoter.

    Infection:

    Article Title: Function and Protective Capacity of Treponema pallidum subsp. pallidum Glycerophosphodiester Phosphodiesterase
    Article Snippet: Immune rabbit serum (IRS) was collected from rabbits that had been chronically infected with T. pallidum for > 90 days. .. Anti-Gpd polyclonal antiserum was raised in a New Zealand White rabbit by immunizing three times with 100 μg of each of the inclusion bodies purified from E. coli BL21 (DE3) pLysS transformed with the Gpd-pET-3a construct, emulsified in the Ribi adjuvant MPL + TDM + CWS (monophosphoryl lipid A + trehalose dicorynomycolate + cell wall skeleton) (Sigma).

    Generated:

    Article Title: A simple vector system to improve performance and utilisation of recombinant antibodies
    Article Snippet: E. coli BL21 (DE3) and Origami 2™ (DE3) cells and the pET26b(+) plasmid were from Novagen (San Diego, CA). .. The other scFvs were generated in house to commercially available antigens purchased from R & D Systems (MN, USA), USBiologicals (MA, USA) and Progen (Heidelberg, Germany).

    Polymerase Chain Reaction:

    Article Title: Molecular cloning and in-silico characterization of high temperature stress responsive pAPX gene isolated from heat tolerant Indian wheat cv. Raj 3765
    Article Snippet: Paragraph title: RACE (rapid amplification of cDNA ends) PCR of TapAPX gene and heterologous protein expression in E. coli ... The expression study of TapAPX gene in prokaryotic system was done by transforming the pET-28a-TapAPX recombinant plasmid in E. coli BL21 cells (Novagen, USA) using heat shock method [ ].

    Sonication:

    Article Title: A Second Mitochondrial DNA Primase Is Essential for Cell Growth and Kinetoplast Minicircle DNA Replication in Trypanosoma brucei ▿ ▿ †
    Article Snippet: Both plasmids pJH604 and pJH618 were transformed into E. coli BL21 cells and grown in Terrific broth (Sigma-Aldrich) supplemented with 0.5 M diglycine and ampicillin (100 μg/ml) at 37°C to an optical density of 1.4. .. Cells were harvested by centrifugation, lysed in metal chelate affinity chromatography buffer (Novagen) containing 0.1% Triton X-100, and then sonicated on ice.

    Recombinant:

    Article Title: Molecular cloning and in-silico characterization of high temperature stress responsive pAPX gene isolated from heat tolerant Indian wheat cv. Raj 3765
    Article Snippet: .. The expression study of TapAPX gene in prokaryotic system was done by transforming the pET-28a-TapAPX recombinant plasmid in E. coli BL21 cells (Novagen, USA) using heat shock method [ ]. ..

    Article Title: Fluorescence Resonance Energy Transfer Assay for High-Throughput Screening of ADAMTS1 Inhibitors
    Article Snippet: Materials E . coli BL21 (DE3), E. coli DH5a, pET32a(+) vector and his-bind purification kit were purchased from Novagen. .. Recombinant human ADAMTS1 was purchased from R & D Systems.

    Article Title: Preparation and Antitumor Activity of CS5931, A Novel Polypeptide from Sea Squirt Ciona Savignyi
    Article Snippet: .. Recombinant Expression of CS5931 from E. coli BL21 (DE3) The expression vector, pET28a/CS5931 synthesized by our lab previously [ ], was transfected into E. coli BL21 (DE3) cells (Novagen, Darmstadt, Germany). .. Cells were incubated in low-salt LB medium (containing 100 µg/mL Kanamycin) at 37 °C with shaking at 200 rpm.

    Purification:

    Article Title: A new hybrid bacteriocin, Ent35-MccV, displays antimicrobial activity against pathogenic Gram-positive and Gram-negative bacteria
    Article Snippet: .. 2.3 Gene expression and bacteriocins purification for activity determinations Ent35–MccV was expressed from pMA24 in E. coli BL21 [DE3] (pLysS) grown in LB (Sigma) at 37 °C. ..

    Article Title: Hypoallergenic Der p 1/Der p 2 combination vaccines for immunotherapy of house dust mite allergy
    Article Snippet: Paragraph title: Expression and purification of Der p 2/1 mosaic proteins ... Expression plasmids containing the Der p 2/1 constructs were transformed into E coli strain BL21 (DE3; Novagen, Merck).

    Article Title: Function and Protective Capacity of Treponema pallidum subsp. pallidum Glycerophosphodiester Phosphodiesterase
    Article Snippet: .. Anti-Gpd polyclonal antiserum was raised in a New Zealand White rabbit by immunizing three times with 100 μg of each of the inclusion bodies purified from E. coli BL21 (DE3) pLysS transformed with the Gpd-pET-3a construct, emulsified in the Ribi adjuvant MPL + TDM + CWS (monophosphoryl lipid A + trehalose dicorynomycolate + cell wall skeleton) (Sigma). ..

    Article Title: Molecular cloning and in-silico characterization of high temperature stress responsive pAPX gene isolated from heat tolerant Indian wheat cv. Raj 3765
    Article Snippet: 1 μg of PCR purified product of (TapAPX ) was digested with 1 μl each of 20 U/μl of restriction enzymes BamH1 and Sac1 (NEB, USA) in a reaction of 20 μl, the vector pET-28a (500 ng) digested with same set of restriction enzymes. .. The expression study of TapAPX gene in prokaryotic system was done by transforming the pET-28a-TapAPX recombinant plasmid in E. coli BL21 cells (Novagen, USA) using heat shock method [ ].

    Article Title: A Second Mitochondrial DNA Primase Is Essential for Cell Growth and Kinetoplast Minicircle DNA Replication in Trypanosoma brucei ▿ ▿ †
    Article Snippet: Paragraph title: Purification of the PRI2ΔNT and PRI2ΔNTCS proteins. ... Both plasmids pJH604 and pJH618 were transformed into E. coli BL21 cells and grown in Terrific broth (Sigma-Aldrich) supplemented with 0.5 M diglycine and ampicillin (100 μg/ml) at 37°C to an optical density of 1.4.

    Article Title: Fluorescence Resonance Energy Transfer Assay for High-Throughput Screening of ADAMTS1 Inhibitors
    Article Snippet: .. Materials E . coli BL21 (DE3), E. coli DH5a, pET32a(+) vector and his-bind purification kit were purchased from Novagen. .. Heparin sepharose CL-6B column was purchased from Amersham Biosciences.

    Article Title: Purification and Reconstitution of the Connexin43 Carboxyl Terminus attached to the 4th Transmembrane Domain in Detergent Micelles *
    Article Snippet: Paragraph title: Protein expression and purification ... The E. coli strains BL21(DE3) (Novagen), Rosetta-2(DE3) (Novagen), C41(DE3) (Lucigen, ( )), and C43(DE3) (Lucigen, ( )) were transformed with the TM4-Cx43CT expression plasmid (see above) and then inoculated into 1 L of Luria Bertani medium (LB), enriched minimal media , or ISOGRO media (Sigma-Aldrich).

    Sequencing:

    Article Title: Molecular cloning and in-silico characterization of high temperature stress responsive pAPX gene isolated from heat tolerant Indian wheat cv. Raj 3765
    Article Snippet: Sequencing of the clone having TapAPX gene was carried out using T7 promoter primer to reconfirm the presence of TapAPX gene along with the presence of 6X His-tag at the 5 ′ upstream of the expression vector pET-28a. .. The expression study of TapAPX gene in prokaryotic system was done by transforming the pET-28a-TapAPX recombinant plasmid in E. coli BL21 cells (Novagen, USA) using heat shock method [ ].

    Affinity Chromatography:

    Article Title: A Second Mitochondrial DNA Primase Is Essential for Cell Growth and Kinetoplast Minicircle DNA Replication in Trypanosoma brucei ▿ ▿ †
    Article Snippet: Both plasmids pJH604 and pJH618 were transformed into E. coli BL21 cells and grown in Terrific broth (Sigma-Aldrich) supplemented with 0.5 M diglycine and ampicillin (100 μg/ml) at 37°C to an optical density of 1.4. .. Cells were harvested by centrifugation, lysed in metal chelate affinity chromatography buffer (Novagen) containing 0.1% Triton X-100, and then sonicated on ice.

    Rapid Amplification of cDNA Ends:

    Article Title: Molecular cloning and in-silico characterization of high temperature stress responsive pAPX gene isolated from heat tolerant Indian wheat cv. Raj 3765
    Article Snippet: Paragraph title: RACE (rapid amplification of cDNA ends) PCR of TapAPX gene and heterologous protein expression in E. coli ... The expression study of TapAPX gene in prokaryotic system was done by transforming the pET-28a-TapAPX recombinant plasmid in E. coli BL21 cells (Novagen, USA) using heat shock method [ ].

    Plasmid Preparation:

    Article Title: A simple vector system to improve performance and utilisation of recombinant antibodies
    Article Snippet: .. E. coli BL21 (DE3) and Origami 2™ (DE3) cells and the pET26b(+) plasmid were from Novagen (San Diego, CA). ..

    Article Title: Chemical complexity of protein determines optimal E. coli expression host; A comparative study using Erythropoietin, Streptokinase and Tumor Necrosis Factor Receptor
    Article Snippet: E. coli BL21 (DE3), E. coli BL21 (DE3) pLys S, E. coli BL21 Rosetta (DE3) pLys S (Novagen,) are IPTG inducible strains in which the T7 RNA Polymerase gene is under the control of a Lac UV5 promoter. .. The expression vector used in this study is pRSET A obtained from Invitrogen Life Technologies, USA. pRSET is a high copy number plasmid with a pUC origin of replication.

    Article Title: A Single Codon Optimization Enhances Recombinant Human TNF-α Vaccine Expression in Escherichia coli
    Article Snippet: .. The vector pET-22b and E. coli strain BL21 (DE3) were purchased from Novagen (San Diego, CA). .. Isopropyl β -D-1-thiogalactopyranoside (IPTG) was from Sigma-Aldrich (St. Louis, MO).

    Article Title: The gaf Fimbrial Gene Cluster of Escherichia coli Expresses a Full-Size and a Truncated Soluble Adhesin Protein
    Article Snippet: .. E. coli strain BL21 λDE3 ( ompT lon ) and expression vector pET-22b(+) were from Novagen Inc. (Madison, Wis.). .. The bacteria were cultivated at 37°C in Luria broth ( ) containing the appropriate antibiotics.

    Article Title: Molecular cloning and in-silico characterization of high temperature stress responsive pAPX gene isolated from heat tolerant Indian wheat cv. Raj 3765
    Article Snippet: .. The expression study of TapAPX gene in prokaryotic system was done by transforming the pET-28a-TapAPX recombinant plasmid in E. coli BL21 cells (Novagen, USA) using heat shock method [ ]. ..

    Article Title: A novel HPV16 E7-affitoxin for targeted therapy of HPV16-induced human cervical cancer
    Article Snippet: .. The pET21a(+) vector and E.coli BL21 (DE3) were purchased from Novagen and ATCC, respectively. .. Reagents The reagents used, including Cell Counting Kit-8 (CCK-8) (Dojindo, Japan), RPMI-1640 (Gibco, USA), fetal bovine serum (FBS) (Gibco, USA), penicillin (Gibco, USA), trypsin-EDTA (Gibco, USA), streptomycin (Sigma Aldrich, Saint Louis, USA), Isopropyl-D-thiogalactopyranoside (IPTG) (Sigma Aldrich, Saint Louis, USA), Ni-NTA agarose (Qiagen Inc., Valencia, CA), and DyLight-755 (Thermo Fisher Scientific, USA), were purchased from commercial sources.

    Article Title: Fluorescence Resonance Energy Transfer Assay for High-Throughput Screening of ADAMTS1 Inhibitors
    Article Snippet: .. Materials E . coli BL21 (DE3), E. coli DH5a, pET32a(+) vector and his-bind purification kit were purchased from Novagen. .. Heparin sepharose CL-6B column was purchased from Amersham Biosciences.

    Article Title: Preparation and Antitumor Activity of CS5931, A Novel Polypeptide from Sea Squirt Ciona Savignyi
    Article Snippet: .. Recombinant Expression of CS5931 from E. coli BL21 (DE3) The expression vector, pET28a/CS5931 synthesized by our lab previously [ ], was transfected into E. coli BL21 (DE3) cells (Novagen, Darmstadt, Germany). .. Cells were incubated in low-salt LB medium (containing 100 µg/mL Kanamycin) at 37 °C with shaking at 200 rpm.

    Article Title: Identification and molecular characterization of a metagenome-derived L-lysine decarboxylase gene from subtropical soil microorganisms
    Article Snippet: .. Plasmid pET-30a(+) (Novagen) and bacterial strain E . coli BL21 (DE3) pLysS (Novagen) were used as the expression vector and host, respectively. ..

    Article Title: Purification and Reconstitution of the Connexin43 Carboxyl Terminus attached to the 4th Transmembrane Domain in Detergent Micelles *
    Article Snippet: .. The E. coli strains BL21(DE3) (Novagen), Rosetta-2(DE3) (Novagen), C41(DE3) (Lucigen, ( )), and C43(DE3) (Lucigen, ( )) were transformed with the TM4-Cx43CT expression plasmid (see above) and then inoculated into 1 L of Luria Bertani medium (LB), enriched minimal media , or ISOGRO media (Sigma-Aldrich). .. Cultures were incubated at 37°C with continuous agitation.

    Negative Control:

    Article Title: A novel HPV16 E7-affitoxin for targeted therapy of HPV16-induced human cervical cancer
    Article Snippet: SiHa (ATCC: HTB-35, HPV16 positive, contains about one to two copies of integrated HPV16 genome), CaSki (ATCC: CRL-1550, HPV16 positive, contains about 600 copies of integrated HPV16 genome), HeLa 229 (ATCC: CCL-2.1, HPV18 positive, used as HPV16 negative control cell line), and melanoma tumor A375 (ATCC: CRL-1619, used as HPV negative control cell line) were obtained from the American Type Culture Collection (ATCC, USA) and cultured as previously described . .. The pET21a(+) vector and E.coli BL21 (DE3) were purchased from Novagen and ATCC, respectively.

    Positron Emission Tomography:

    Article Title: A Single Codon Optimization Enhances Recombinant Human TNF-α Vaccine Expression in Escherichia coli
    Article Snippet: .. The vector pET-22b and E. coli strain BL21 (DE3) were purchased from Novagen (San Diego, CA). .. Isopropyl β -D-1-thiogalactopyranoside (IPTG) was from Sigma-Aldrich (St. Louis, MO).

    Article Title: The gaf Fimbrial Gene Cluster of Escherichia coli Expresses a Full-Size and a Truncated Soluble Adhesin Protein
    Article Snippet: .. E. coli strain BL21 λDE3 ( ompT lon ) and expression vector pET-22b(+) were from Novagen Inc. (Madison, Wis.). .. The bacteria were cultivated at 37°C in Luria broth ( ) containing the appropriate antibiotics.

    Article Title: Molecular cloning and in-silico characterization of high temperature stress responsive pAPX gene isolated from heat tolerant Indian wheat cv. Raj 3765
    Article Snippet: .. The expression study of TapAPX gene in prokaryotic system was done by transforming the pET-28a-TapAPX recombinant plasmid in E. coli BL21 cells (Novagen, USA) using heat shock method [ ]. ..

    Article Title: Identification and molecular characterization of a metagenome-derived L-lysine decarboxylase gene from subtropical soil microorganisms
    Article Snippet: .. Plasmid pET-30a(+) (Novagen) and bacterial strain E . coli BL21 (DE3) pLysS (Novagen) were used as the expression vector and host, respectively. ..

    Concentration Assay:

    Article Title: Molecular cloning and in-silico characterization of high temperature stress responsive pAPX gene isolated from heat tolerant Indian wheat cv. Raj 3765
    Article Snippet: The expression study of TapAPX gene in prokaryotic system was done by transforming the pET-28a-TapAPX recombinant plasmid in E. coli BL21 cells (Novagen, USA) using heat shock method [ ]. .. Isopropyl β-D thiogalacto pyranoside (IPTG), an inducer of T7 promoter in pET-28a vector, was added at final concentration of 1 mM when O.D of the culture reached an absorbance of 0.5 at 600 nm.

    Article Title: Preparation and Antitumor Activity of CS5931, A Novel Polypeptide from Sea Squirt Ciona Savignyi
    Article Snippet: Recombinant Expression of CS5931 from E. coli BL21 (DE3) The expression vector, pET28a/CS5931 synthesized by our lab previously [ ], was transfected into E. coli BL21 (DE3) cells (Novagen, Darmstadt, Germany). .. After the value of OD600 of the culture medium reached around 0.5–0.7, IPTG was added at a final concentration of 0.75 mM and the cells were incubated overnight at 16 °C.

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    Millipore e coli strain bl21
    Zymographic detection of the autolytic activity in recombinant His-tagged Atl. Protein extracts from E. coli <t>BL21</t> cells grown with IPTG were used in this study. The protein extract was separated by 12.5% SDS-gel electrophoresis. The SDS gel was impregnated with autoclaved S. aureus RN450 cells. Autolytic activity was detected by renaturing the autolysins by incubation of the gel in 50 mM Tri-HCl buffer, pH 7.5, containing 0.05% Triton X-100. Lane 1: full length His-tagged Atl; lane 2: Atl-1; lane 3: Atl-2; lane 4: Atl-3; lane 5: Atl-4; lane 6: Atl-5; lane 7: Atl-6; lane 8: Atl-7; lane 9: Atl-8.
    E Coli Strain Bl21, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 284 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Zymographic detection of the autolytic activity in recombinant His-tagged Atl. Protein extracts from E. coli BL21 cells grown with IPTG were used in this study. The protein extract was separated by 12.5% SDS-gel electrophoresis. The SDS gel was impregnated with autoclaved S. aureus RN450 cells. Autolytic activity was detected by renaturing the autolysins by incubation of the gel in 50 mM Tri-HCl buffer, pH 7.5, containing 0.05% Triton X-100. Lane 1: full length His-tagged Atl; lane 2: Atl-1; lane 3: Atl-2; lane 4: Atl-3; lane 5: Atl-4; lane 6: Atl-5; lane 7: Atl-6; lane 8: Atl-7; lane 9: Atl-8.

    Journal: International Journal of Microbiology

    Article Title: High Level Expression and Purification of Atl, the Major Autolytic Protein of Staphylococcus aureus

    doi: 10.1155/2014/615965

    Figure Lengend Snippet: Zymographic detection of the autolytic activity in recombinant His-tagged Atl. Protein extracts from E. coli BL21 cells grown with IPTG were used in this study. The protein extract was separated by 12.5% SDS-gel electrophoresis. The SDS gel was impregnated with autoclaved S. aureus RN450 cells. Autolytic activity was detected by renaturing the autolysins by incubation of the gel in 50 mM Tri-HCl buffer, pH 7.5, containing 0.05% Triton X-100. Lane 1: full length His-tagged Atl; lane 2: Atl-1; lane 3: Atl-2; lane 4: Atl-3; lane 5: Atl-4; lane 6: Atl-5; lane 7: Atl-6; lane 8: Atl-7; lane 9: Atl-8.

    Article Snippet: In addition, E. coli strain BL21 (EMD Millipore) was used for all protein expression studies.

    Techniques: Activity Assay, Recombinant, SDS-Gel, Electrophoresis, Incubation

    Coomassie stained gels showing purity of recombinant His-tagged Atl. Recombinant Atl and Atl-1 proteins were overproduced in E. coli and purified as described in the Materials and Methods section. Lane M: standard protein markers; lanes 1 and 2: protein extract from E. coli BL21 cells transformed with plasmids pRSETA- atl and grown without and with IPTG, respectively; lane 3: purified His-tagged Atl; lanes 4 and 5: protein extract from E. coli BL21 cells transformed with plasmids pRSETA- atl1 and grown without and with IPTG, respectively; lane 6: purified His-tagged Atl-1.

    Journal: International Journal of Microbiology

    Article Title: High Level Expression and Purification of Atl, the Major Autolytic Protein of Staphylococcus aureus

    doi: 10.1155/2014/615965

    Figure Lengend Snippet: Coomassie stained gels showing purity of recombinant His-tagged Atl. Recombinant Atl and Atl-1 proteins were overproduced in E. coli and purified as described in the Materials and Methods section. Lane M: standard protein markers; lanes 1 and 2: protein extract from E. coli BL21 cells transformed with plasmids pRSETA- atl and grown without and with IPTG, respectively; lane 3: purified His-tagged Atl; lanes 4 and 5: protein extract from E. coli BL21 cells transformed with plasmids pRSETA- atl1 and grown without and with IPTG, respectively; lane 6: purified His-tagged Atl-1.

    Article Snippet: In addition, E. coli strain BL21 (EMD Millipore) was used for all protein expression studies.

    Techniques: Staining, Recombinant, Purification, Transformation Assay

    Zymographic detection of the autolytic activity in purified recombinant His-tagged Atl. Lane 1: total protein extract from S. aureus cells. Lanes 2 and 4: purified His-tagged Atl and Atl-1, respectively; lanes 3 and 5: protein extract from E. coli BL21 cells transformed with plasmids pRSETA- atl or pRSETA- atl1 and grown with IPTG, respectively.

    Journal: International Journal of Microbiology

    Article Title: High Level Expression and Purification of Atl, the Major Autolytic Protein of Staphylococcus aureus

    doi: 10.1155/2014/615965

    Figure Lengend Snippet: Zymographic detection of the autolytic activity in purified recombinant His-tagged Atl. Lane 1: total protein extract from S. aureus cells. Lanes 2 and 4: purified His-tagged Atl and Atl-1, respectively; lanes 3 and 5: protein extract from E. coli BL21 cells transformed with plasmids pRSETA- atl or pRSETA- atl1 and grown with IPTG, respectively.

    Article Snippet: In addition, E. coli strain BL21 (EMD Millipore) was used for all protein expression studies.

    Techniques: Activity Assay, Purification, Recombinant, Transformation Assay

    Coomassie stained gels demonstrating overexpression of recombinant Atl. The 12.5% SDS-PAGE contains protein extracts of E. coli BL21 cells with pRSETA plasmids expressing either full length or truncated Atl proteins. The odd number labels are cells grown without IPTG and the even number labels are the cells grown with IPTG. Lane M: standard protein markers; lanes 1 and 2 - Atl (full length His-tagged Atl); lanes 3 and 4: Atl-1; lanes 5 and 6: Atl-2; lanes 7 and 8: Atl-3; lanes 9 and 10: Atl-4; lanes 11 and 12: Atl-6; lanes 13 and 14: Atl-5; lanes 15 and 16: Atl-7; lanes 17 and 18: Atl-8. The Atl number suffixes are indicated in Table 1 .

    Journal: International Journal of Microbiology

    Article Title: High Level Expression and Purification of Atl, the Major Autolytic Protein of Staphylococcus aureus

    doi: 10.1155/2014/615965

    Figure Lengend Snippet: Coomassie stained gels demonstrating overexpression of recombinant Atl. The 12.5% SDS-PAGE contains protein extracts of E. coli BL21 cells with pRSETA plasmids expressing either full length or truncated Atl proteins. The odd number labels are cells grown without IPTG and the even number labels are the cells grown with IPTG. Lane M: standard protein markers; lanes 1 and 2 - Atl (full length His-tagged Atl); lanes 3 and 4: Atl-1; lanes 5 and 6: Atl-2; lanes 7 and 8: Atl-3; lanes 9 and 10: Atl-4; lanes 11 and 12: Atl-6; lanes 13 and 14: Atl-5; lanes 15 and 16: Atl-7; lanes 17 and 18: Atl-8. The Atl number suffixes are indicated in Table 1 .

    Article Snippet: In addition, E. coli strain BL21 (EMD Millipore) was used for all protein expression studies.

    Techniques: Staining, Over Expression, Recombinant, SDS Page, Expressing

    Binding of LsrB to endogenous AI-2. Purified LsrB (BL21) and LsrB (BL21∆luxS) proteins (5 mg/ml) were incubated for 10 min at 37, 50 and 60 °C to release endogenous AI-2, respectively. After incubation, the LsrB proteins were removed by ultrafiltration (10,000-Da cut-off; EMD Millipore), and the filtered reaction products were tested for AI-2 activity using a V. harveyi BB170 bioassay. The extent of LsrB binding to endogenous AI-2 was evaluated using an AI-2 assay, which showed that recombinant LsrB (BL21) bound to endogenous AI-2 (produced by wild-type strain BL21) and was released from LsrB (BL21) at 50 or 60 °C, respectively ( a ). However, since the luxS mutant BL21∆luxS did not produce endogenous AI-2, no AI-2 could be released from the recombinant LsrB (BL21∆luxS) ( b ). Moreover, the recombinant LuxS protein, which was expressed in strain BL21 (pColdTF-lsrB) as a negative control, also showed no AI-2 binding activity ( c ). AI-2 (10 μM) was used as a positive control

    Journal: AMB Express

    Article Title: LsrB-based and temperature-dependent identification of bacterial AI-2 receptor

    doi: 10.1186/s13568-017-0486-y

    Figure Lengend Snippet: Binding of LsrB to endogenous AI-2. Purified LsrB (BL21) and LsrB (BL21∆luxS) proteins (5 mg/ml) were incubated for 10 min at 37, 50 and 60 °C to release endogenous AI-2, respectively. After incubation, the LsrB proteins were removed by ultrafiltration (10,000-Da cut-off; EMD Millipore), and the filtered reaction products were tested for AI-2 activity using a V. harveyi BB170 bioassay. The extent of LsrB binding to endogenous AI-2 was evaluated using an AI-2 assay, which showed that recombinant LsrB (BL21) bound to endogenous AI-2 (produced by wild-type strain BL21) and was released from LsrB (BL21) at 50 or 60 °C, respectively ( a ). However, since the luxS mutant BL21∆luxS did not produce endogenous AI-2, no AI-2 could be released from the recombinant LsrB (BL21∆luxS) ( b ). Moreover, the recombinant LuxS protein, which was expressed in strain BL21 (pColdTF-lsrB) as a negative control, also showed no AI-2 binding activity ( c ). AI-2 (10 μM) was used as a positive control

    Article Snippet: Cell-free culture fluid (CF) was prepared as follows: E. coli strains BL21 (DE3), BL21∆luxS and DH5a were grown in LB at 37 °C, and then pelleted by centrifugation at 12,000g at 4 °C for 10 min. Then, the resulting supernatants were filtered through a 0.22-μm filter (EMD Millipore, Bedford, MA, USA) to obtain CF samples.

    Techniques: Binding Assay, Purification, Incubation, Activity Assay, Recombinant, Produced, Mutagenesis, Negative Control, Positive Control

    Schematic chart of strategy for binding of LsrB to endogenous AI-2 or exogenous AI-2. The extent of LsrB binding to endogenous AI-2 was evaluated using an AI-2 assay, which showed that recombinant LsrB (BL21) bound to endogenous AI-2 (produced by wild-type strain BL21) and was released from LsrB (BL21) at 50 or 60 °C. However, since the luxS mutant BL21∆luxS did not produce endogenous AI-2, no AI-2 could be released from the recombinant LsrB (BL21∆luxS). The combination of endogenous AI-2 produced by BL21 (DE3), which is known to interfere with the binding of AI-2 to LsrB (BL21), resulted in the loss in the ability of LsrB (BL21) to bind to exogenous AI-2. Hence, the BL21 (DE3) luxS mutant, which was incapable of producing endogenous AI-2, but could bind to exogenous AI-2

    Journal: AMB Express

    Article Title: LsrB-based and temperature-dependent identification of bacterial AI-2 receptor

    doi: 10.1186/s13568-017-0486-y

    Figure Lengend Snippet: Schematic chart of strategy for binding of LsrB to endogenous AI-2 or exogenous AI-2. The extent of LsrB binding to endogenous AI-2 was evaluated using an AI-2 assay, which showed that recombinant LsrB (BL21) bound to endogenous AI-2 (produced by wild-type strain BL21) and was released from LsrB (BL21) at 50 or 60 °C. However, since the luxS mutant BL21∆luxS did not produce endogenous AI-2, no AI-2 could be released from the recombinant LsrB (BL21∆luxS). The combination of endogenous AI-2 produced by BL21 (DE3), which is known to interfere with the binding of AI-2 to LsrB (BL21), resulted in the loss in the ability of LsrB (BL21) to bind to exogenous AI-2. Hence, the BL21 (DE3) luxS mutant, which was incapable of producing endogenous AI-2, but could bind to exogenous AI-2

    Article Snippet: Cell-free culture fluid (CF) was prepared as follows: E. coli strains BL21 (DE3), BL21∆luxS and DH5a were grown in LB at 37 °C, and then pelleted by centrifugation at 12,000g at 4 °C for 10 min. Then, the resulting supernatants were filtered through a 0.22-μm filter (EMD Millipore, Bedford, MA, USA) to obtain CF samples.

    Techniques: Binding Assay, Recombinant, Produced, Mutagenesis

    Identification of the luxS mutant BL21∆ luxS. a Schematic chart of strategy for producing the BL21 luxS deletion mutant. The luxS was deleted by replacing the partial gene sequence of luxS with kanamycin resistance cassette at Sal I cleavage sites. The primers used for the confirmation of the luxS deletion are also indicated. b Identification of the luxS mutant BL21∆ luxS. Lane M: DL 2000 DNA marker (D501A; Takara); lane 1: the wild-type strain BL21 showed a 2519-bp PCR product using primers LuxS-OutF/LuxS-OutR; lane 2: the mutant BL21∆ luxS with cure of the kanamycin resistance cassette showed a 2209-bp PCR product using primers LuxS-OutF/LuxS-OutR; lane 3: negative control; lane 4: the wild-type strain DE17 showed a 307-bp PCR product using primers LuxS-inF/LuxS-inR; lane 5: the mutant DE17∆ pfs showed no PCR products using primers LuxS-inF/LuxS-inR; lane 6: negative control

    Journal: AMB Express

    Article Title: LsrB-based and temperature-dependent identification of bacterial AI-2 receptor

    doi: 10.1186/s13568-017-0486-y

    Figure Lengend Snippet: Identification of the luxS mutant BL21∆ luxS. a Schematic chart of strategy for producing the BL21 luxS deletion mutant. The luxS was deleted by replacing the partial gene sequence of luxS with kanamycin resistance cassette at Sal I cleavage sites. The primers used for the confirmation of the luxS deletion are also indicated. b Identification of the luxS mutant BL21∆ luxS. Lane M: DL 2000 DNA marker (D501A; Takara); lane 1: the wild-type strain BL21 showed a 2519-bp PCR product using primers LuxS-OutF/LuxS-OutR; lane 2: the mutant BL21∆ luxS with cure of the kanamycin resistance cassette showed a 2209-bp PCR product using primers LuxS-OutF/LuxS-OutR; lane 3: negative control; lane 4: the wild-type strain DE17 showed a 307-bp PCR product using primers LuxS-inF/LuxS-inR; lane 5: the mutant DE17∆ pfs showed no PCR products using primers LuxS-inF/LuxS-inR; lane 6: negative control

    Article Snippet: Cell-free culture fluid (CF) was prepared as follows: E. coli strains BL21 (DE3), BL21∆luxS and DH5a were grown in LB at 37 °C, and then pelleted by centrifugation at 12,000g at 4 °C for 10 min. Then, the resulting supernatants were filtered through a 0.22-μm filter (EMD Millipore, Bedford, MA, USA) to obtain CF samples.

    Techniques: Mutagenesis, Sequencing, Marker, Polymerase Chain Reaction, Negative Control

    The AI-2 activity in BL21∆luxS. The wild strain BL21 secretes AI-2-like molecules, and can induce V . harveyi BB170 bioluminescence, whereas no bioluminescence induction was observed for the mutant BL21∆ luxS . V . harveyi BB152 served as a positive control and E. coli DH5α as a negative control. The figure represents the means of the results from three independent experiments. The error bars indicate standard deviations

    Journal: AMB Express

    Article Title: LsrB-based and temperature-dependent identification of bacterial AI-2 receptor

    doi: 10.1186/s13568-017-0486-y

    Figure Lengend Snippet: The AI-2 activity in BL21∆luxS. The wild strain BL21 secretes AI-2-like molecules, and can induce V . harveyi BB170 bioluminescence, whereas no bioluminescence induction was observed for the mutant BL21∆ luxS . V . harveyi BB152 served as a positive control and E. coli DH5α as a negative control. The figure represents the means of the results from three independent experiments. The error bars indicate standard deviations

    Article Snippet: Cell-free culture fluid (CF) was prepared as follows: E. coli strains BL21 (DE3), BL21∆luxS and DH5a were grown in LB at 37 °C, and then pelleted by centrifugation at 12,000g at 4 °C for 10 min. Then, the resulting supernatants were filtered through a 0.22-μm filter (EMD Millipore, Bedford, MA, USA) to obtain CF samples.

    Techniques: Activity Assay, Mutagenesis, Positive Control, Negative Control

    Identification of the luxS mutant BL21∆ luxS. a Schematic chart of strategy for producing the BL21 luxS deletion mutant. The luxS was deleted by replacing the partial gene sequence of luxS with kanamycin resistance cassette at Sal I cleavage sites.

    Journal: AMB Express

    Article Title: LsrB-based and temperature-dependent identification of bacterial AI-2 receptor

    doi: 10.1186/s13568-017-0486-y

    Figure Lengend Snippet: Identification of the luxS mutant BL21∆ luxS. a Schematic chart of strategy for producing the BL21 luxS deletion mutant. The luxS was deleted by replacing the partial gene sequence of luxS with kanamycin resistance cassette at Sal I cleavage sites.

    Article Snippet: Cell-free culture fluid (CF) was prepared as follows: E. coli strains BL21 (DE3), BL21∆luxS and DH5a were grown in LB at 37 °C, and then pelleted by centrifugation at 12,000 g at 4 °C for 10 min. Then, the resulting supernatants were filtered through a 0.22-μm filter (EMD Millipore, Bedford, MA, USA) to obtain CF samples.

    Techniques: Mutagenesis, Sequencing

    The AI-2 activity in BL21∆luxS. The wild strain BL21 secretes AI-2-like molecules, and can induce V . harveyi BB170 bioluminescence, whereas no bioluminescence induction was observed for the mutant BL21∆ luxS . V . harveyi BB152 served as

    Journal: AMB Express

    Article Title: LsrB-based and temperature-dependent identification of bacterial AI-2 receptor

    doi: 10.1186/s13568-017-0486-y

    Figure Lengend Snippet: The AI-2 activity in BL21∆luxS. The wild strain BL21 secretes AI-2-like molecules, and can induce V . harveyi BB170 bioluminescence, whereas no bioluminescence induction was observed for the mutant BL21∆ luxS . V . harveyi BB152 served as

    Article Snippet: Cell-free culture fluid (CF) was prepared as follows: E. coli strains BL21 (DE3), BL21∆luxS and DH5a were grown in LB at 37 °C, and then pelleted by centrifugation at 12,000 g at 4 °C for 10 min. Then, the resulting supernatants were filtered through a 0.22-μm filter (EMD Millipore, Bedford, MA, USA) to obtain CF samples.

    Techniques: Activity Assay, Mutagenesis