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Agilent technologies e coli xl1 blue cells
E Coli Xl1 Blue Cells, supplied by Agilent technologies, used in various techniques. Bioz Stars score: 93/100, based on 10 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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Average 93 stars, based on 10 article reviews
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e coli xl1 blue cells - by Bioz Stars, 2020-04
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Clone Assay:

Article Title: Defining the complementarities between antibodies and haptens to refine our understanding and aid the prediction of a successful binding interaction
Article Snippet: Expression and purification of scAb proteins The plasmid DNA of the anti-hapten scFv were digested with Nco I and Not I restriction enzymes, and cloned into a similarly digested soluble expression vector pIMS147 [ ]. .. The ligated product was transformed into E. coli (XL1-Blue) cells (Agilent Technologies, 200228) by electroporation.

Article Title: TDP-43 Inclusion Bodies Formed in Bacteria Are Structurally Amorphous, Non-Amyloid and Inherently Toxic to Neuroblastoma Cells
Article Snippet: Paragraph title: FL and Ct TDP-43 Gene Cloning and Protein Expression ... Cultures of E. coli XL1 Blue cells (Agilent Technologies, Santa Clara, CA, USA) were transformed with the resulting plasmids containing FL TDP-43 or Ct TDP-43 and were grown overnight at 37°C in LB medium with 100 µg/mL ampicillin under vigorous shaking.

Article Title: Ubiquitin is a versatile scaffold protein for the generation of molecules with de novo binding and advantageous drug-like properties
Article Snippet: Paragraph title: Cloning ... Ligation reactions were purified with the MinElute Reaction Cleanup kit (Qiagen) and transformed into E. coli XL1 blue cells (Agilent).

Article Title: A fully synthetic human Fab antibody library based on fixed VH/VL framework pairings with favorable biophysical properties
Article Snippet: Electrocompetent E. coli MC1061F′ cells with a transformation efficiency of at least 1E+10/µg DNA were used for library cloning. .. For all other molecular cloning procedures, E.coli XL1-Blue cells (Agilent Technologies) were used and during antibody selections phage were propagated in E.coli TG1F+ cells (Agilent Technologies).

Article Title: Genotype of a historic strain of Mycobacterium tuberculosis
Article Snippet: .. DNA was cloned using the CloneJet PCR cloning kit (Fermentas) in Escherichia coli XL1-Blue cells (Agilent). .. Recombinant plasmid DNA was purified using Qiaquick columns and sequenced (GATC Biotech).

Article Title: Folding and activity of mutant cystathionine ?-synthase depends on the position and nature of the purification tag: Characterization of the R266K CBS mutant
Article Snippet: For the preparation of the pET47-NP-hCBS construct, first the original vector was modified in order to allow for cloning into the Apa I and Xho I sites. .. All constructs were transformed into E. coli XL1-Blue cells (Agilent) and their authenticity was confirmed by DNA sequencing.

Article Title: Domain Organization, Catalysis and Regulation of Eukaryotic Cystathionine Beta-Synthases
Article Snippet: Preparation of CBS constructs We used our established constructs for hCBS as models for cloning of yCBS and dCBS into pGEX-6P1 (GE Healthcare) and/or pET-28a (Novagen) vectors and for preparation of their C-terminally truncated forms , . lists all the oligonucleotides used for subcloning and mutagenesis. .. Both yCBS and dCBS constructs, designated as pGEX-6P1-yCBS and pET28-C-dCBS, respectively, were transformed into E. coli XL1-Blue cells (Agilent) and their authenticity was confirmed by DNA sequencing.

Article Title: Regulation of the thermoalkaliphilic F1-ATPase from Caldalkalibacillus thermarum
Article Snippet: .. All cloning was conducted in Escherichia coli XL1-Blue cells (Agilent), and the enzyme used for all PCR was the DNA polymerase from Thermococcus kodakaraensis (Merck Millipore). ..

Centrifugation:

Article Title: TDP-43 Inclusion Bodies Formed in Bacteria Are Structurally Amorphous, Non-Amyloid and Inherently Toxic to Neuroblastoma Cells
Article Snippet: Cultures of E. coli XL1 Blue cells (Agilent Technologies, Santa Clara, CA, USA) were transformed with the resulting plasmids containing FL TDP-43 or Ct TDP-43 and were grown overnight at 37°C in LB medium with 100 µg/mL ampicillin under vigorous shaking. .. Cells were harvested by centrifugation, resuspended in PBS buffer (137 mM NaCl, 2.7 mM KCl, 4.3 mM Na2 HPO4 , 1.4 mM KH2 PO4 , 1 mM EDTA, 1 mM β-mercaptoethanol, 0.1 mM PMSF, at pH 7.3) and then lysed by 30 min incubation with 1 mg/mL HEWL in ice, followed by sonication at 40 kHz (five cycles of 30 s each).

Article Title: Engineered Dengue Virus Domain III Proteins Elicit Cross-Neutralizing Antibody Responses in Mice
Article Snippet: EDIII-expressing phage particles were produced by electroporation into Escherichia coli XL1-Blue cells (Agilent Technologies, Santa Clara, CA) and recovered in 2× yeast extract tryptone (YT) medium containing 50 μg/ml carbenicillin and 10 μg/ml tetracycline at 37°C and 220 rpm for 5 h. Phage were coinfected with M13KO7 helper phage (New England BioLabs, Ipswich, MA) at a final concentration of 1010 PFU per ml and incubated for 1 h at 37°C and 220 rpm. .. Phage were harvested by precipitation with 4% (wt/vol) polyethylene glycol (PEG) 8000 and 3% (wt/vol) NaCl after centrifugation to remove cells.

Amplification:

Article Title: Domain Organization, Catalysis and Regulation of Eukaryotic Cystathionine Beta-Synthases
Article Snippet: The dCBS coding sequence was PCR amplified from a recently reported pGEX-6P1-DMCBS plasmid using the 824 and 825 oligonucleotides. .. Both pET28-C-yCBS WT and pET28-C-yCBS L345* constructs were transformed into E. coli XL1-Blue cells (Agilent) and their authenticity and the presence of a C-terminal 6xHis tag was confirmed by DNA sequencing.

Article Title: TDP-43 Inclusion Bodies Formed in Bacteria Are Structurally Amorphous, Non-Amyloid and Inherently Toxic to Neuroblastoma Cells
Article Snippet: Each amplified sequence and the pGEX-2T plasmid were digested with BamHI and EcoRI (Thermo Fisher Scientific) and combined with the T4 DNA ligase (Thermo Fisher Scientific) to obtain the constructs coding for the GST/FL TDP-43 and the GST/Ct TDP-43 fusion proteins. .. Cultures of E. coli XL1 Blue cells (Agilent Technologies, Santa Clara, CA, USA) were transformed with the resulting plasmids containing FL TDP-43 or Ct TDP-43 and were grown overnight at 37°C in LB medium with 100 µg/mL ampicillin under vigorous shaking.

Article Title: Ubiquitin is a versatile scaffold protein for the generation of molecules with de novo binding and advantageous drug-like properties
Article Snippet: 2.9 Cloning The CXCR4 gene was purchased from Thermo Scientific and PCR amplified with specific primers: pCDNA3-CX forward 5′-GCA TGA ATT CGC CAC CAT GGA GGG GAT CAG TAT ATA C-3′ and pCDNA3-CX reverse 5′-GCA TCT CGA GTT AGC TGG AGT GAA AAC TTG AAG ACT C-3′. .. Ligation reactions were purified with the MinElute Reaction Cleanup kit (Qiagen) and transformed into E. coli XL1 blue cells (Agilent).

Synthesized:

Article Title: MEHMO syndrome mutation EIF2S3-I259M impairs initiator Met-tRNAiMet binding to eukaryotic translation initiation factor eIF2
Article Snippet: Yeast tRNAi Met was synthesized by in vitro transcription with T7 polymerase as described previously ( ). .. His-tagged MetRS was purified from Escherichia coli XL1 Blue cells (Agilent) carrying the E. coli methionyl-tRNA synthetase (MetRS) expression plasmid pRA101 ( ) as described previously ( ).

Article Title: MEHMO syndrome mutation EIF2S3-I259M impairs initiator Met-tRNAiMet binding to eukaryotic translation initiation factor eIF2
Article Snippet: Synthesis and aminoacylation of tRNAi Met Yeast tRNAi Met was synthesized by in vitro transcription with T7 polymerase as described previously ( ). .. His-tagged MetRS was purified from Escherichia coli XL1 Blue cells (Agilent) carrying the E. coli methionyl-tRNA synthetase (MetRS) expression plasmid pRA101 ( ) as described previously ( ).

Construct:

Article Title: Domain Organization, Catalysis and Regulation of Eukaryotic Cystathionine Beta-Synthases
Article Snippet: .. Both pET28-C-yCBS WT and pET28-C-yCBS L345* constructs were transformed into E. coli XL1-Blue cells (Agilent) and their authenticity and the presence of a C-terminal 6xHis tag was confirmed by DNA sequencing. .. In order to prepare the various truncated forms of yCBS and dCBS lacking the regulatory domain , the translational STOP codons were introduced using a QuikChange II XL site directed mutagenesis kit (Agilent).

Article Title: TDP-43 Inclusion Bodies Formed in Bacteria Are Structurally Amorphous, Non-Amyloid and Inherently Toxic to Neuroblastoma Cells
Article Snippet: Each amplified sequence and the pGEX-2T plasmid were digested with BamHI and EcoRI (Thermo Fisher Scientific) and combined with the T4 DNA ligase (Thermo Fisher Scientific) to obtain the constructs coding for the GST/FL TDP-43 and the GST/Ct TDP-43 fusion proteins. .. Cultures of E. coli XL1 Blue cells (Agilent Technologies, Santa Clara, CA, USA) were transformed with the resulting plasmids containing FL TDP-43 or Ct TDP-43 and were grown overnight at 37°C in LB medium with 100 µg/mL ampicillin under vigorous shaking.

Article Title: Folding and activity of mutant cystathionine ?-synthase depends on the position and nature of the purification tag: Characterization of the R266K CBS mutant
Article Snippet: .. All constructs were transformed into E. coli XL1-Blue cells (Agilent) and their authenticity was confirmed by DNA sequencing. .. The R266K missense mutation was subsequently introduced into the wild type CBS constructs by the QuikChange site directed mutagenesis kit (Agilent) using forward (5′-cacgggcattgccAGGaagctgaaggag) and reverse (5′-ctccttcagcttCCTggcaatgcccgtg) oligonucleotides.

Article Title: Domain Organization, Catalysis and Regulation of Eukaryotic Cystathionine Beta-Synthases
Article Snippet: .. Both yCBS and dCBS constructs, designated as pGEX-6P1-yCBS and pET28-C-dCBS, respectively, were transformed into E. coli XL1-Blue cells (Agilent) and their authenticity was confirmed by DNA sequencing. .. The pGEX-6P1-yCBS construct was later used as a template for PCR for recloning of full-length yCBS WT and truncated yCBS L345* into a pET-28a vector using the 794 and 795 and 794 and 796 oligonucleotides, respectively.

Enzyme-linked Immunosorbent Assay:

Article Title: Defining the complementarities between antibodies and haptens to refine our understanding and aid the prediction of a successful binding interaction
Article Snippet: The ligated product was transformed into E. coli (XL1-Blue) cells (Agilent Technologies, 200228) by electroporation. .. The sensitivity of expressed scAb proteins for their target antigen was examined using a scAb binding ELISA [ ] and/or an indirect competition ELISA [ ].

Article Title: Engineered Dengue Virus Domain III Proteins Elicit Cross-Neutralizing Antibody Responses in Mice
Article Snippet: EDIII-expressing phage particles were produced by electroporation into Escherichia coli XL1-Blue cells (Agilent Technologies, Santa Clara, CA) and recovered in 2× yeast extract tryptone (YT) medium containing 50 μg/ml carbenicillin and 10 μg/ml tetracycline at 37°C and 220 rpm for 5 h. Phage were coinfected with M13KO7 helper phage (New England BioLabs, Ipswich, MA) at a final concentration of 1010 PFU per ml and incubated for 1 h at 37°C and 220 rpm. .. Surface expression of EDIII was confirmed by phage ELISA against the anti-FLAG monoclonal antibody M2.

Incubation:

Article Title: MEHMO syndrome mutation EIF2S3-I259M impairs initiator Met-tRNAiMet binding to eukaryotic translation initiation factor eIF2
Article Snippet: His-tagged MetRS was purified from Escherichia coli XL1 Blue cells (Agilent) carrying the E. coli methionyl-tRNA synthetase (MetRS) expression plasmid pRA101 ( ) as described previously ( ). .. Aminoacylation reactions with 5 μM synthesized tRNAi Met , 2 mM adenosine triphosphate-Mg2+ , 0.3 μM [35 S]methionine (Perkin Elmer), 10 mM MgCl2 and 1 μM MetRS were incubated in 1× Reaction Buffer (100 mM HEPES-KOH (pH 7.5), 1 mM DTT and 10 mM KCl) for 30 min at 30°C, and aminoacylated Met-tRNAi Met was purified by phenol/chloroform extraction ( ).

Article Title: TDP-43 Inclusion Bodies Formed in Bacteria Are Structurally Amorphous, Non-Amyloid and Inherently Toxic to Neuroblastoma Cells
Article Snippet: Cultures of E. coli XL1 Blue cells (Agilent Technologies, Santa Clara, CA, USA) were transformed with the resulting plasmids containing FL TDP-43 or Ct TDP-43 and were grown overnight at 37°C in LB medium with 100 µg/mL ampicillin under vigorous shaking. .. Cells were harvested by centrifugation, resuspended in PBS buffer (137 mM NaCl, 2.7 mM KCl, 4.3 mM Na2 HPO4 , 1.4 mM KH2 PO4 , 1 mM EDTA, 1 mM β-mercaptoethanol, 0.1 mM PMSF, at pH 7.3) and then lysed by 30 min incubation with 1 mg/mL HEWL in ice, followed by sonication at 40 kHz (five cycles of 30 s each).

Article Title: MEHMO syndrome mutation EIF2S3-I259M impairs initiator Met-tRNAiMet binding to eukaryotic translation initiation factor eIF2
Article Snippet: His-tagged MetRS was purified from Escherichia coli XL1 Blue cells (Agilent) carrying the E. coli methionyl-tRNA synthetase (MetRS) expression plasmid pRA101 ( ) as described previously ( ). .. Aminoacylation reactions with 5 μM synthesized tRNAi Met , 2 mM adenosine triphosphate-Mg2+ , 0.3 μM [35 S]methionine (Perkin Elmer), 10 mM MgCl2 and 1 μM MetRS were incubated in 1× Reaction Buffer (100 mM HEPES-KOH (pH 7.5), 1 mM DTT and 10 mM KCl) for 30 min at 30°C, and aminoacylated Met-tRNAi Met was purified by phenol/chloroform extraction ( ).

Article Title: Engineered Dengue Virus Domain III Proteins Elicit Cross-Neutralizing Antibody Responses in Mice
Article Snippet: .. EDIII-expressing phage particles were produced by electroporation into Escherichia coli XL1-Blue cells (Agilent Technologies, Santa Clara, CA) and recovered in 2× yeast extract tryptone (YT) medium containing 50 μg/ml carbenicillin and 10 μg/ml tetracycline at 37°C and 220 rpm for 5 h. Phage were coinfected with M13KO7 helper phage (New England BioLabs, Ipswich, MA) at a final concentration of 1010 PFU per ml and incubated for 1 h at 37°C and 220 rpm. .. Phage were harvested by precipitation with 4% (wt/vol) polyethylene glycol (PEG) 8000 and 3% (wt/vol) NaCl after centrifugation to remove cells.

Expressing:

Article Title: MEHMO syndrome mutation EIF2S3-I259M impairs initiator Met-tRNAiMet binding to eukaryotic translation initiation factor eIF2
Article Snippet: .. His-tagged MetRS was purified from Escherichia coli XL1 Blue cells (Agilent) carrying the E. coli methionyl-tRNA synthetase (MetRS) expression plasmid pRA101 ( ) as described previously ( ). .. Aminoacylation reactions with 5 μM synthesized tRNAi Met , 2 mM adenosine triphosphate-Mg2+ , 0.3 μM [35 S]methionine (Perkin Elmer), 10 mM MgCl2 and 1 μM MetRS were incubated in 1× Reaction Buffer (100 mM HEPES-KOH (pH 7.5), 1 mM DTT and 10 mM KCl) for 30 min at 30°C, and aminoacylated Met-tRNAi Met was purified by phenol/chloroform extraction ( ).

Article Title: Defining the complementarities between antibodies and haptens to refine our understanding and aid the prediction of a successful binding interaction
Article Snippet: Paragraph title: Expression and purification of scAb proteins ... The ligated product was transformed into E. coli (XL1-Blue) cells (Agilent Technologies, 200228) by electroporation.

Article Title: TDP-43 Inclusion Bodies Formed in Bacteria Are Structurally Amorphous, Non-Amyloid and Inherently Toxic to Neuroblastoma Cells
Article Snippet: Paragraph title: FL and Ct TDP-43 Gene Cloning and Protein Expression ... Cultures of E. coli XL1 Blue cells (Agilent Technologies, Santa Clara, CA, USA) were transformed with the resulting plasmids containing FL TDP-43 or Ct TDP-43 and were grown overnight at 37°C in LB medium with 100 µg/mL ampicillin under vigorous shaking.

Article Title: Folding and activity of mutant cystathionine ?-synthase depends on the position and nature of the purification tag: Characterization of the R266K CBS mutant
Article Snippet: All constructs were transformed into E. coli XL1-Blue cells (Agilent) and their authenticity was confirmed by DNA sequencing. .. Verified plasmids for wild type as well as R266K mutant CBS were finally transformed into E. coli Rosetta2 (DE3) expression host cells (Novagen).

Article Title: MEHMO syndrome mutation EIF2S3-I259M impairs initiator Met-tRNAiMet binding to eukaryotic translation initiation factor eIF2
Article Snippet: .. His-tagged MetRS was purified from Escherichia coli XL1 Blue cells (Agilent) carrying the E. coli methionyl-tRNA synthetase (MetRS) expression plasmid pRA101 ( ) as described previously ( ). .. Aminoacylation reactions with 5 μM synthesized tRNAi Met , 2 mM adenosine triphosphate-Mg2+ , 0.3 μM [35 S]methionine (Perkin Elmer), 10 mM MgCl2 and 1 μM MetRS were incubated in 1× Reaction Buffer (100 mM HEPES-KOH (pH 7.5), 1 mM DTT and 10 mM KCl) for 30 min at 30°C, and aminoacylated Met-tRNAi Met was purified by phenol/chloroform extraction ( ).

Article Title: Engineered Dengue Virus Domain III Proteins Elicit Cross-Neutralizing Antibody Responses in Mice
Article Snippet: EDIII-expressing phage particles were produced by electroporation into Escherichia coli XL1-Blue cells (Agilent Technologies, Santa Clara, CA) and recovered in 2× yeast extract tryptone (YT) medium containing 50 μg/ml carbenicillin and 10 μg/ml tetracycline at 37°C and 220 rpm for 5 h. Phage were coinfected with M13KO7 helper phage (New England BioLabs, Ipswich, MA) at a final concentration of 1010 PFU per ml and incubated for 1 h at 37°C and 220 rpm. .. Surface expression of EDIII was confirmed by phage ELISA against the anti-FLAG monoclonal antibody M2.

Article Title: Regulation of the thermoalkaliphilic F1-ATPase from Caldalkalibacillus thermarum
Article Snippet: Paragraph title: Expression Plasmids. ... All cloning was conducted in Escherichia coli XL1-Blue cells (Agilent), and the enzyme used for all PCR was the DNA polymerase from Thermococcus kodakaraensis (Merck Millipore).

Modification:

Article Title: Folding and activity of mutant cystathionine ?-synthase depends on the position and nature of the purification tag: Characterization of the R266K CBS mutant
Article Snippet: Subsequently, the CBS fragment was ligated into an Apa I- Xho I linearized modified pET-47b(+) vector. .. All constructs were transformed into E. coli XL1-Blue cells (Agilent) and their authenticity was confirmed by DNA sequencing.

Transformation Assay:

Article Title: Domain Organization, Catalysis and Regulation of Eukaryotic Cystathionine Beta-Synthases
Article Snippet: .. Both pET28-C-yCBS WT and pET28-C-yCBS L345* constructs were transformed into E. coli XL1-Blue cells (Agilent) and their authenticity and the presence of a C-terminal 6xHis tag was confirmed by DNA sequencing. .. In order to prepare the various truncated forms of yCBS and dCBS lacking the regulatory domain , the translational STOP codons were introduced using a QuikChange II XL site directed mutagenesis kit (Agilent).

Article Title: His224 Alters the R2 Drug Binding Site and Phe218 Influences the Catalytic Efficiency of the Metallo-β-Lactamase VIM-7
Article Snippet: .. Parental DNA was digested with DpnI, and E. coli XL1-Blue cells (Agilent Biosciences) were transformed with the PCR products for clone selection. .. Mutations were verified by DNA sequencing as described above, using insert flanking T7 primers ( ).

Article Title: Defining the complementarities between antibodies and haptens to refine our understanding and aid the prediction of a successful binding interaction
Article Snippet: .. The ligated product was transformed into E. coli (XL1-Blue) cells (Agilent Technologies, 200228) by electroporation. .. Expressions of scAbs in transformed E. coli XL-1 Blue bacterial cells were carried out in Terrific Broth according to standard protocols [ ].

Article Title: TDP-43 Inclusion Bodies Formed in Bacteria Are Structurally Amorphous, Non-Amyloid and Inherently Toxic to Neuroblastoma Cells
Article Snippet: .. Cultures of E. coli XL1 Blue cells (Agilent Technologies, Santa Clara, CA, USA) were transformed with the resulting plasmids containing FL TDP-43 or Ct TDP-43 and were grown overnight at 37°C in LB medium with 100 µg/mL ampicillin under vigorous shaking. ..

Article Title: Ubiquitin is a versatile scaffold protein for the generation of molecules with de novo binding and advantageous drug-like properties
Article Snippet: .. Ligation reactions were purified with the MinElute Reaction Cleanup kit (Qiagen) and transformed into E. coli XL1 blue cells (Agilent). .. After antibiotic selection a positive clone was sequence verified and propagated for plasmid preparation with the QIAfilter Plasmid Midi kit (Qiagen).

Article Title: A fully synthetic human Fab antibody library based on fixed VH/VL framework pairings with favorable biophysical properties
Article Snippet: Electrocompetent E. coli MC1061F′ cells with a transformation efficiency of at least 1E+10/µg DNA were used for library cloning. .. For all other molecular cloning procedures, E.coli XL1-Blue cells (Agilent Technologies) were used and during antibody selections phage were propagated in E.coli TG1F+ cells (Agilent Technologies).

Article Title: Folding and activity of mutant cystathionine ?-synthase depends on the position and nature of the purification tag: Characterization of the R266K CBS mutant
Article Snippet: .. All constructs were transformed into E. coli XL1-Blue cells (Agilent) and their authenticity was confirmed by DNA sequencing. .. The R266K missense mutation was subsequently introduced into the wild type CBS constructs by the QuikChange site directed mutagenesis kit (Agilent) using forward (5′-cacgggcattgccAGGaagctgaaggag) and reverse (5′-ctccttcagcttCCTggcaatgcccgtg) oligonucleotides.

Article Title: Domain Organization, Catalysis and Regulation of Eukaryotic Cystathionine Beta-Synthases
Article Snippet: .. Both yCBS and dCBS constructs, designated as pGEX-6P1-yCBS and pET28-C-dCBS, respectively, were transformed into E. coli XL1-Blue cells (Agilent) and their authenticity was confirmed by DNA sequencing. .. The pGEX-6P1-yCBS construct was later used as a template for PCR for recloning of full-length yCBS WT and truncated yCBS L345* into a pET-28a vector using the 794 and 795 and 794 and 796 oligonucleotides, respectively.

Article Title: Regulation of the thermoalkaliphilic F1-ATPase from Caldalkalibacillus thermarum
Article Snippet: All cloning was conducted in Escherichia coli XL1-Blue cells (Agilent), and the enzyme used for all PCR was the DNA polymerase from Thermococcus kodakaraensis (Merck Millipore). .. For overexpression of the F1 -ATPase, the plasmids pTrcF1ΔδHisTev and pTrcF1ΔδHisTev-(D89A, R92A) were transformed into a strain of E. coli C41 known as E. coli C41(Δ unc ), where the unc operon, encoding the E. coli F-ATPase complex, had been disrupted by the insertion of a chloramphenicol resistance gene.

Over Expression:

Article Title: Regulation of the thermoalkaliphilic F1-ATPase from Caldalkalibacillus thermarum
Article Snippet: All cloning was conducted in Escherichia coli XL1-Blue cells (Agilent), and the enzyme used for all PCR was the DNA polymerase from Thermococcus kodakaraensis (Merck Millipore). .. For overexpression of the F1 -ATPase, the plasmids pTrcF1ΔδHisTev and pTrcF1ΔδHisTev-(D89A, R92A) were transformed into a strain of E. coli C41 known as E. coli C41(Δ unc ), where the unc operon, encoding the E. coli F-ATPase complex, had been disrupted by the insertion of a chloramphenicol resistance gene.

Chloramphenicol Acetyltransferase Assay:

Article Title: Ubiquitin is a versatile scaffold protein for the generation of molecules with de novo binding and advantageous drug-like properties
Article Snippet: 2.9 Cloning The CXCR4 gene was purchased from Thermo Scientific and PCR amplified with specific primers: pCDNA3-CX forward 5′-GCA TGA ATT CGC CAC CAT GGA GGG GAT CAG TAT ATA C-3′ and pCDNA3-CX reverse 5′-GCA TCT CGA GTT AGC TGG AGT GAA AAC TTG AAG ACT C-3′. .. Ligation reactions were purified with the MinElute Reaction Cleanup kit (Qiagen) and transformed into E. coli XL1 blue cells (Agilent).

Electroporation:

Article Title: Defining the complementarities between antibodies and haptens to refine our understanding and aid the prediction of a successful binding interaction
Article Snippet: .. The ligated product was transformed into E. coli (XL1-Blue) cells (Agilent Technologies, 200228) by electroporation. .. Expressions of scAbs in transformed E. coli XL-1 Blue bacterial cells were carried out in Terrific Broth according to standard protocols [ ].

Article Title: Engineered Dengue Virus Domain III Proteins Elicit Cross-Neutralizing Antibody Responses in Mice
Article Snippet: .. EDIII-expressing phage particles were produced by electroporation into Escherichia coli XL1-Blue cells (Agilent Technologies, Santa Clara, CA) and recovered in 2× yeast extract tryptone (YT) medium containing 50 μg/ml carbenicillin and 10 μg/ml tetracycline at 37°C and 220 rpm for 5 h. Phage were coinfected with M13KO7 helper phage (New England BioLabs, Ipswich, MA) at a final concentration of 1010 PFU per ml and incubated for 1 h at 37°C and 220 rpm. .. Phage were harvested by precipitation with 4% (wt/vol) polyethylene glycol (PEG) 8000 and 3% (wt/vol) NaCl after centrifugation to remove cells.

Ligation:

Article Title: Ubiquitin is a versatile scaffold protein for the generation of molecules with de novo binding and advantageous drug-like properties
Article Snippet: .. Ligation reactions were purified with the MinElute Reaction Cleanup kit (Qiagen) and transformed into E. coli XL1 blue cells (Agilent). .. After antibiotic selection a positive clone was sequence verified and propagated for plasmid preparation with the QIAfilter Plasmid Midi kit (Qiagen).

DNA Sequencing:

Article Title: Domain Organization, Catalysis and Regulation of Eukaryotic Cystathionine Beta-Synthases
Article Snippet: .. Both pET28-C-yCBS WT and pET28-C-yCBS L345* constructs were transformed into E. coli XL1-Blue cells (Agilent) and their authenticity and the presence of a C-terminal 6xHis tag was confirmed by DNA sequencing. .. In order to prepare the various truncated forms of yCBS and dCBS lacking the regulatory domain , the translational STOP codons were introduced using a QuikChange II XL site directed mutagenesis kit (Agilent).

Article Title: His224 Alters the R2 Drug Binding Site and Phe218 Influences the Catalytic Efficiency of the Metallo-β-Lactamase VIM-7
Article Snippet: Parental DNA was digested with DpnI, and E. coli XL1-Blue cells (Agilent Biosciences) were transformed with the PCR products for clone selection. .. Mutations were verified by DNA sequencing as described above, using insert flanking T7 primers ( ).

Article Title: TDP-43 Inclusion Bodies Formed in Bacteria Are Structurally Amorphous, Non-Amyloid and Inherently Toxic to Neuroblastoma Cells
Article Snippet: Their correct nucleotide sequence was verified by DNA sequencing. .. Cultures of E. coli XL1 Blue cells (Agilent Technologies, Santa Clara, CA, USA) were transformed with the resulting plasmids containing FL TDP-43 or Ct TDP-43 and were grown overnight at 37°C in LB medium with 100 µg/mL ampicillin under vigorous shaking.

Article Title: Folding and activity of mutant cystathionine ?-synthase depends on the position and nature of the purification tag: Characterization of the R266K CBS mutant
Article Snippet: .. All constructs were transformed into E. coli XL1-Blue cells (Agilent) and their authenticity was confirmed by DNA sequencing. .. The R266K missense mutation was subsequently introduced into the wild type CBS constructs by the QuikChange site directed mutagenesis kit (Agilent) using forward (5′-cacgggcattgccAGGaagctgaaggag) and reverse (5′-ctccttcagcttCCTggcaatgcccgtg) oligonucleotides.

Article Title: Domain Organization, Catalysis and Regulation of Eukaryotic Cystathionine Beta-Synthases
Article Snippet: .. Both yCBS and dCBS constructs, designated as pGEX-6P1-yCBS and pET28-C-dCBS, respectively, were transformed into E. coli XL1-Blue cells (Agilent) and their authenticity was confirmed by DNA sequencing. .. The pGEX-6P1-yCBS construct was later used as a template for PCR for recloning of full-length yCBS WT and truncated yCBS L345* into a pET-28a vector using the 794 and 795 and 794 and 796 oligonucleotides, respectively.

Sequencing:

Article Title: Domain Organization, Catalysis and Regulation of Eukaryotic Cystathionine Beta-Synthases
Article Snippet: The dCBS coding sequence was PCR amplified from a recently reported pGEX-6P1-DMCBS plasmid using the 824 and 825 oligonucleotides. .. Both pET28-C-yCBS WT and pET28-C-yCBS L345* constructs were transformed into E. coli XL1-Blue cells (Agilent) and their authenticity and the presence of a C-terminal 6xHis tag was confirmed by DNA sequencing.

Article Title: Ubiquitin is a versatile scaffold protein for the generation of molecules with de novo binding and advantageous drug-like properties
Article Snippet: Ligation reactions were purified with the MinElute Reaction Cleanup kit (Qiagen) and transformed into E. coli XL1 blue cells (Agilent). .. After antibiotic selection a positive clone was sequence verified and propagated for plasmid preparation with the QIAfilter Plasmid Midi kit (Qiagen).

Article Title: Folding and activity of mutant cystathionine ?-synthase depends on the position and nature of the purification tag: Characterization of the R266K CBS mutant
Article Snippet: The CBS coding sequence was PCR-amplified using a forward primer with an Apa I site (5′-ccagGGGCCCtctgag) and reverse primer with an Xho I site (5′-gccgCTCGAGtcgactc). .. All constructs were transformed into E. coli XL1-Blue cells (Agilent) and their authenticity was confirmed by DNA sequencing.

Sonication:

Article Title: TDP-43 Inclusion Bodies Formed in Bacteria Are Structurally Amorphous, Non-Amyloid and Inherently Toxic to Neuroblastoma Cells
Article Snippet: Cultures of E. coli XL1 Blue cells (Agilent Technologies, Santa Clara, CA, USA) were transformed with the resulting plasmids containing FL TDP-43 or Ct TDP-43 and were grown overnight at 37°C in LB medium with 100 µg/mL ampicillin under vigorous shaking. .. Cells were harvested by centrifugation, resuspended in PBS buffer (137 mM NaCl, 2.7 mM KCl, 4.3 mM Na2 HPO4 , 1.4 mM KH2 PO4 , 1 mM EDTA, 1 mM β-mercaptoethanol, 0.1 mM PMSF, at pH 7.3) and then lysed by 30 min incubation with 1 mg/mL HEWL in ice, followed by sonication at 40 kHz (five cycles of 30 s each).

Binding Assay:

Article Title: Defining the complementarities between antibodies and haptens to refine our understanding and aid the prediction of a successful binding interaction
Article Snippet: The ligated product was transformed into E. coli (XL1-Blue) cells (Agilent Technologies, 200228) by electroporation. .. The sensitivity of expressed scAb proteins for their target antigen was examined using a scAb binding ELISA [ ] and/or an indirect competition ELISA [ ].

Molecular Cloning:

Article Title: A fully synthetic human Fab antibody library based on fixed VH/VL framework pairings with favorable biophysical properties
Article Snippet: .. For all other molecular cloning procedures, E.coli XL1-Blue cells (Agilent Technologies) were used and during antibody selections phage were propagated in E.coli TG1F+ cells (Agilent Technologies). ..

Mutagenesis:

Article Title: Domain Organization, Catalysis and Regulation of Eukaryotic Cystathionine Beta-Synthases
Article Snippet: Both pET28-C-yCBS WT and pET28-C-yCBS L345* constructs were transformed into E. coli XL1-Blue cells (Agilent) and their authenticity and the presence of a C-terminal 6xHis tag was confirmed by DNA sequencing. .. In order to prepare the various truncated forms of yCBS and dCBS lacking the regulatory domain , the translational STOP codons were introduced using a QuikChange II XL site directed mutagenesis kit (Agilent).

Article Title: His224 Alters the R2 Drug Binding Site and Phe218 Influences the Catalytic Efficiency of the Metallo-β-Lactamase VIM-7
Article Snippet: Paragraph title: Construction of VIM-7 mutants by site-directed mutagenesis. ... Parental DNA was digested with DpnI, and E. coli XL1-Blue cells (Agilent Biosciences) were transformed with the PCR products for clone selection.

Article Title: Folding and activity of mutant cystathionine ?-synthase depends on the position and nature of the purification tag: Characterization of the R266K CBS mutant
Article Snippet: The internal Apa I site at position 1405 was abolished and subsequently the San DI site in the PreScission protease recognition site was mutated into an Apa I site both using the QuikChange site directed mutagenesis kit (Agilent). .. All constructs were transformed into E. coli XL1-Blue cells (Agilent) and their authenticity was confirmed by DNA sequencing.

Article Title: Domain Organization, Catalysis and Regulation of Eukaryotic Cystathionine Beta-Synthases
Article Snippet: Preparation of CBS constructs We used our established constructs for hCBS as models for cloning of yCBS and dCBS into pGEX-6P1 (GE Healthcare) and/or pET-28a (Novagen) vectors and for preparation of their C-terminally truncated forms , . lists all the oligonucleotides used for subcloning and mutagenesis. .. Both yCBS and dCBS constructs, designated as pGEX-6P1-yCBS and pET28-C-dCBS, respectively, were transformed into E. coli XL1-Blue cells (Agilent) and their authenticity was confirmed by DNA sequencing.

Subcloning:

Article Title: Domain Organization, Catalysis and Regulation of Eukaryotic Cystathionine Beta-Synthases
Article Snippet: Preparation of CBS constructs We used our established constructs for hCBS as models for cloning of yCBS and dCBS into pGEX-6P1 (GE Healthcare) and/or pET-28a (Novagen) vectors and for preparation of their C-terminally truncated forms , . lists all the oligonucleotides used for subcloning and mutagenesis. .. Both yCBS and dCBS constructs, designated as pGEX-6P1-yCBS and pET28-C-dCBS, respectively, were transformed into E. coli XL1-Blue cells (Agilent) and their authenticity was confirmed by DNA sequencing.

Purification:

Article Title: Cross Kingdom Functional Conservation of the Core Universally Conserved Threonylcarbamoyladenosine tRNA Synthesis Enzymes
Article Snippet: .. E. coli tRNAAsn(GUU) and tRNALys(UUU) were overexpressed in E. coli XL1-Blue cells (Agilent Technologies) and purified as described previously ( ). .. For purification, RNAs were loaded on an 8 M urea, 12% polyacrylamide gel, and the band corresponding to overexpressed tRNA was cut out. tRNA was eluted from gel in elution buffer (EB; 0.1% SDS, 0.5 M ammonium acetate [NH4 Ac], 10 mM MgAc, 1 mM EDTA), precipitated, and suspended in water.

Article Title: MEHMO syndrome mutation EIF2S3-I259M impairs initiator Met-tRNAiMet binding to eukaryotic translation initiation factor eIF2
Article Snippet: .. His-tagged MetRS was purified from Escherichia coli XL1 Blue cells (Agilent) carrying the E. coli methionyl-tRNA synthetase (MetRS) expression plasmid pRA101 ( ) as described previously ( ). .. Aminoacylation reactions with 5 μM synthesized tRNAi Met , 2 mM adenosine triphosphate-Mg2+ , 0.3 μM [35 S]methionine (Perkin Elmer), 10 mM MgCl2 and 1 μM MetRS were incubated in 1× Reaction Buffer (100 mM HEPES-KOH (pH 7.5), 1 mM DTT and 10 mM KCl) for 30 min at 30°C, and aminoacylated Met-tRNAi Met was purified by phenol/chloroform extraction ( ).

Article Title: Defining the complementarities between antibodies and haptens to refine our understanding and aid the prediction of a successful binding interaction
Article Snippet: Paragraph title: Expression and purification of scAb proteins ... The ligated product was transformed into E. coli (XL1-Blue) cells (Agilent Technologies, 200228) by electroporation.

Article Title: Ubiquitin is a versatile scaffold protein for the generation of molecules with de novo binding and advantageous drug-like properties
Article Snippet: .. Ligation reactions were purified with the MinElute Reaction Cleanup kit (Qiagen) and transformed into E. coli XL1 blue cells (Agilent). .. After antibiotic selection a positive clone was sequence verified and propagated for plasmid preparation with the QIAfilter Plasmid Midi kit (Qiagen).

Article Title: Genotype of a historic strain of Mycobacterium tuberculosis
Article Snippet: PCR products were examined by electrophoresis in 2% agarose gels, and bands were purified using Qiaquick columns (Qiagen). .. DNA was cloned using the CloneJet PCR cloning kit (Fermentas) in Escherichia coli XL1-Blue cells (Agilent).

Article Title: MEHMO syndrome mutation EIF2S3-I259M impairs initiator Met-tRNAiMet binding to eukaryotic translation initiation factor eIF2
Article Snippet: .. His-tagged MetRS was purified from Escherichia coli XL1 Blue cells (Agilent) carrying the E. coli methionyl-tRNA synthetase (MetRS) expression plasmid pRA101 ( ) as described previously ( ). .. Aminoacylation reactions with 5 μM synthesized tRNAi Met , 2 mM adenosine triphosphate-Mg2+ , 0.3 μM [35 S]methionine (Perkin Elmer), 10 mM MgCl2 and 1 μM MetRS were incubated in 1× Reaction Buffer (100 mM HEPES-KOH (pH 7.5), 1 mM DTT and 10 mM KCl) for 30 min at 30°C, and aminoacylated Met-tRNAi Met was purified by phenol/chloroform extraction ( ).

Polymerase Chain Reaction:

Article Title: Domain Organization, Catalysis and Regulation of Eukaryotic Cystathionine Beta-Synthases
Article Snippet: The Nco I- and Xho I-digested, gel-extracted PCR product was ligated into a similarly prepared pET-28a vector using T4 DNA ligase (NEB Biolabs). .. Both pET28-C-yCBS WT and pET28-C-yCBS L345* constructs were transformed into E. coli XL1-Blue cells (Agilent) and their authenticity and the presence of a C-terminal 6xHis tag was confirmed by DNA sequencing.

Article Title: His224 Alters the R2 Drug Binding Site and Phe218 Influences the Catalytic Efficiency of the Metallo-β-Lactamase VIM-7
Article Snippet: .. Parental DNA was digested with DpnI, and E. coli XL1-Blue cells (Agilent Biosciences) were transformed with the PCR products for clone selection. .. Mutations were verified by DNA sequencing as described above, using insert flanking T7 primers ( ).

Article Title: TDP-43 Inclusion Bodies Formed in Bacteria Are Structurally Amorphous, Non-Amyloid and Inherently Toxic to Neuroblastoma Cells
Article Snippet: In brief, the sequence coding for FL TDP-43 and that coding for Ct TDP-43 were amplified from the pINCY vector (Thermo Fisher Scientific, Waltham, MA, USA) by PCR, using forward and reverse primers containing the restriction sites for BamHI and EcoRI enzymes, respectively. .. Cultures of E. coli XL1 Blue cells (Agilent Technologies, Santa Clara, CA, USA) were transformed with the resulting plasmids containing FL TDP-43 or Ct TDP-43 and were grown overnight at 37°C in LB medium with 100 µg/mL ampicillin under vigorous shaking.

Article Title: Ubiquitin is a versatile scaffold protein for the generation of molecules with de novo binding and advantageous drug-like properties
Article Snippet: PCR fragments were digested with EcoRI and XhoI (NEB) and ligated into pCDNA3 (Life Technologies) digested with the same enzymes using the Rapid Ligation kit (Roche). .. Ligation reactions were purified with the MinElute Reaction Cleanup kit (Qiagen) and transformed into E. coli XL1 blue cells (Agilent).

Article Title: Genotype of a historic strain of Mycobacterium tuberculosis
Article Snippet: .. DNA was cloned using the CloneJet PCR cloning kit (Fermentas) in Escherichia coli XL1-Blue cells (Agilent). .. Recombinant plasmid DNA was purified using Qiaquick columns and sequenced (GATC Biotech).

Article Title: Folding and activity of mutant cystathionine ?-synthase depends on the position and nature of the purification tag: Characterization of the R266K CBS mutant
Article Snippet: The CBS coding sequence was PCR-amplified using a forward primer with an Apa I site (5′-ccagGGGCCCtctgag) and reverse primer with an Xho I site (5′-gccgCTCGAGtcgactc). .. All constructs were transformed into E. coli XL1-Blue cells (Agilent) and their authenticity was confirmed by DNA sequencing.

Article Title: Domain Organization, Catalysis and Regulation of Eukaryotic Cystathionine Beta-Synthases
Article Snippet: The Nco I- and Hind III-digested, gel-extracted PCR product was ligated into a similarly prepared pET-28a vector using T4 DNA ligase (NEB Biolabs). .. Both yCBS and dCBS constructs, designated as pGEX-6P1-yCBS and pET28-C-dCBS, respectively, were transformed into E. coli XL1-Blue cells (Agilent) and their authenticity was confirmed by DNA sequencing.

Article Title: Regulation of the thermoalkaliphilic F1-ATPase from Caldalkalibacillus thermarum
Article Snippet: .. All cloning was conducted in Escherichia coli XL1-Blue cells (Agilent), and the enzyme used for all PCR was the DNA polymerase from Thermococcus kodakaraensis (Merck Millipore). ..

Affinity Chromatography:

Article Title: Defining the complementarities between antibodies and haptens to refine our understanding and aid the prediction of a successful binding interaction
Article Snippet: The ligated product was transformed into E. coli (XL1-Blue) cells (Agilent Technologies, 200228) by electroporation. .. The expressed scAbs were purified via the hexa-histidine tag using immobilized metal ion chelate affinity chromatography (IMAC).

Recombinant:

Article Title: Genotype of a historic strain of Mycobacterium tuberculosis
Article Snippet: DNA was cloned using the CloneJet PCR cloning kit (Fermentas) in Escherichia coli XL1-Blue cells (Agilent). .. Recombinant plasmid DNA was purified using Qiaquick columns and sequenced (GATC Biotech).

Activated Clotting Time Assay:

Article Title: Ubiquitin is a versatile scaffold protein for the generation of molecules with de novo binding and advantageous drug-like properties
Article Snippet: 2.9 Cloning The CXCR4 gene was purchased from Thermo Scientific and PCR amplified with specific primers: pCDNA3-CX forward 5′-GCA TGA ATT CGC CAC CAT GGA GGG GAT CAG TAT ATA C-3′ and pCDNA3-CX reverse 5′-GCA TCT CGA GTT AGC TGG AGT GAA AAC TTG AAG ACT C-3′. .. Ligation reactions were purified with the MinElute Reaction Cleanup kit (Qiagen) and transformed into E. coli XL1 blue cells (Agilent).

Plasmid Preparation:

Article Title: Domain Organization, Catalysis and Regulation of Eukaryotic Cystathionine Beta-Synthases
Article Snippet: The Nco I- and Xho I-digested, gel-extracted PCR product was ligated into a similarly prepared pET-28a vector using T4 DNA ligase (NEB Biolabs). .. Both pET28-C-yCBS WT and pET28-C-yCBS L345* constructs were transformed into E. coli XL1-Blue cells (Agilent) and their authenticity and the presence of a C-terminal 6xHis tag was confirmed by DNA sequencing.

Article Title: MEHMO syndrome mutation EIF2S3-I259M impairs initiator Met-tRNAiMet binding to eukaryotic translation initiation factor eIF2
Article Snippet: .. His-tagged MetRS was purified from Escherichia coli XL1 Blue cells (Agilent) carrying the E. coli methionyl-tRNA synthetase (MetRS) expression plasmid pRA101 ( ) as described previously ( ). .. Aminoacylation reactions with 5 μM synthesized tRNAi Met , 2 mM adenosine triphosphate-Mg2+ , 0.3 μM [35 S]methionine (Perkin Elmer), 10 mM MgCl2 and 1 μM MetRS were incubated in 1× Reaction Buffer (100 mM HEPES-KOH (pH 7.5), 1 mM DTT and 10 mM KCl) for 30 min at 30°C, and aminoacylated Met-tRNAi Met was purified by phenol/chloroform extraction ( ).

Article Title: Defining the complementarities between antibodies and haptens to refine our understanding and aid the prediction of a successful binding interaction
Article Snippet: Expression and purification of scAb proteins The plasmid DNA of the anti-hapten scFv were digested with Nco I and Not I restriction enzymes, and cloned into a similarly digested soluble expression vector pIMS147 [ ]. .. The ligated product was transformed into E. coli (XL1-Blue) cells (Agilent Technologies, 200228) by electroporation.

Article Title: TDP-43 Inclusion Bodies Formed in Bacteria Are Structurally Amorphous, Non-Amyloid and Inherently Toxic to Neuroblastoma Cells
Article Snippet: Each amplified sequence and the pGEX-2T plasmid were digested with BamHI and EcoRI (Thermo Fisher Scientific) and combined with the T4 DNA ligase (Thermo Fisher Scientific) to obtain the constructs coding for the GST/FL TDP-43 and the GST/Ct TDP-43 fusion proteins. .. Cultures of E. coli XL1 Blue cells (Agilent Technologies, Santa Clara, CA, USA) were transformed with the resulting plasmids containing FL TDP-43 or Ct TDP-43 and were grown overnight at 37°C in LB medium with 100 µg/mL ampicillin under vigorous shaking.

Article Title: Ubiquitin is a versatile scaffold protein for the generation of molecules with de novo binding and advantageous drug-like properties
Article Snippet: Ligation reactions were purified with the MinElute Reaction Cleanup kit (Qiagen) and transformed into E. coli XL1 blue cells (Agilent). .. After antibiotic selection a positive clone was sequence verified and propagated for plasmid preparation with the QIAfilter Plasmid Midi kit (Qiagen).

Article Title: FnCpf1: a novel and efficient genome editing tool for Saccharomyces cerevisiae
Article Snippet: .. For plasmid propagation, E. coli XL1-Blue cells (Agilent Technologies, Santa Clara, CA, USA) were cultivated in Lysogeny broth (LB) medium supplied with ampicillin (100 mg l−1 ) or kanamycin (50 mg l−1 ) at 37 °C with 180 rpm agitation. ..

Article Title: Genotype of a historic strain of Mycobacterium tuberculosis
Article Snippet: DNA was cloned using the CloneJet PCR cloning kit (Fermentas) in Escherichia coli XL1-Blue cells (Agilent). .. Recombinant plasmid DNA was purified using Qiaquick columns and sequenced (GATC Biotech).

Article Title: Folding and activity of mutant cystathionine ?-synthase depends on the position and nature of the purification tag: Characterization of the R266K CBS mutant
Article Snippet: Subsequently, the CBS fragment was ligated into an Apa I- Xho I linearized modified pET-47b(+) vector. .. All constructs were transformed into E. coli XL1-Blue cells (Agilent) and their authenticity was confirmed by DNA sequencing.

Article Title: MEHMO syndrome mutation EIF2S3-I259M impairs initiator Met-tRNAiMet binding to eukaryotic translation initiation factor eIF2
Article Snippet: .. His-tagged MetRS was purified from Escherichia coli XL1 Blue cells (Agilent) carrying the E. coli methionyl-tRNA synthetase (MetRS) expression plasmid pRA101 ( ) as described previously ( ). .. Aminoacylation reactions with 5 μM synthesized tRNAi Met , 2 mM adenosine triphosphate-Mg2+ , 0.3 μM [35 S]methionine (Perkin Elmer), 10 mM MgCl2 and 1 μM MetRS were incubated in 1× Reaction Buffer (100 mM HEPES-KOH (pH 7.5), 1 mM DTT and 10 mM KCl) for 30 min at 30°C, and aminoacylated Met-tRNAi Met was purified by phenol/chloroform extraction ( ).

Article Title: Domain Organization, Catalysis and Regulation of Eukaryotic Cystathionine Beta-Synthases
Article Snippet: The Nco I- and Hind III-digested, gel-extracted PCR product was ligated into a similarly prepared pET-28a vector using T4 DNA ligase (NEB Biolabs). .. Both yCBS and dCBS constructs, designated as pGEX-6P1-yCBS and pET28-C-dCBS, respectively, were transformed into E. coli XL1-Blue cells (Agilent) and their authenticity was confirmed by DNA sequencing.

Article Title: Regulation of the thermoalkaliphilic F1-ATPase from Caldalkalibacillus thermarum
Article Snippet: The product was cloned into the NotI and SalI sites in pTrcF1ΔδHisTev, generating the plasmid pTrcF1ΔδHisTev-(D89A, R92A). .. All cloning was conducted in Escherichia coli XL1-Blue cells (Agilent), and the enzyme used for all PCR was the DNA polymerase from Thermococcus kodakaraensis (Merck Millipore).

Electrophoresis:

Article Title: Genotype of a historic strain of Mycobacterium tuberculosis
Article Snippet: PCR products were examined by electrophoresis in 2% agarose gels, and bands were purified using Qiaquick columns (Qiagen). .. DNA was cloned using the CloneJet PCR cloning kit (Fermentas) in Escherichia coli XL1-Blue cells (Agilent).

Selection:

Article Title: His224 Alters the R2 Drug Binding Site and Phe218 Influences the Catalytic Efficiency of the Metallo-β-Lactamase VIM-7
Article Snippet: .. Parental DNA was digested with DpnI, and E. coli XL1-Blue cells (Agilent Biosciences) were transformed with the PCR products for clone selection. .. Mutations were verified by DNA sequencing as described above, using insert flanking T7 primers ( ).

Article Title: Ubiquitin is a versatile scaffold protein for the generation of molecules with de novo binding and advantageous drug-like properties
Article Snippet: Ligation reactions were purified with the MinElute Reaction Cleanup kit (Qiagen) and transformed into E. coli XL1 blue cells (Agilent). .. After antibiotic selection a positive clone was sequence verified and propagated for plasmid preparation with the QIAfilter Plasmid Midi kit (Qiagen).

Article Title: FnCpf1: a novel and efficient genome editing tool for Saccharomyces cerevisiae
Article Snippet: When selection on SMG with G418 was required (NH4 )2 SO4 was replaced with 3 g l−1 K2 SO4 and 2.3 g l−1 filter-sterilized urea to maintain a stable pH ( ). .. For plasmid propagation, E. coli XL1-Blue cells (Agilent Technologies, Santa Clara, CA, USA) were cultivated in Lysogeny broth (LB) medium supplied with ampicillin (100 mg l−1 ) or kanamycin (50 mg l−1 ) at 37 °C with 180 rpm agitation.

Agarose Gel Electrophoresis:

Article Title: Folding and activity of mutant cystathionine ?-synthase depends on the position and nature of the purification tag: Characterization of the R266K CBS mutant
Article Snippet: After cleavage with Apa I and Xho I (NEB Biolabs), the CBS fragment was cut out from a 1% agarose gel and cleaned up using the QIAquick gel extraction kit (Qiagen). .. All constructs were transformed into E. coli XL1-Blue cells (Agilent) and their authenticity was confirmed by DNA sequencing.

In Vitro:

Article Title: MEHMO syndrome mutation EIF2S3-I259M impairs initiator Met-tRNAiMet binding to eukaryotic translation initiation factor eIF2
Article Snippet: Yeast tRNAi Met was synthesized by in vitro transcription with T7 polymerase as described previously ( ). .. His-tagged MetRS was purified from Escherichia coli XL1 Blue cells (Agilent) carrying the E. coli methionyl-tRNA synthetase (MetRS) expression plasmid pRA101 ( ) as described previously ( ).

Article Title: MEHMO syndrome mutation EIF2S3-I259M impairs initiator Met-tRNAiMet binding to eukaryotic translation initiation factor eIF2
Article Snippet: Synthesis and aminoacylation of tRNAi Met Yeast tRNAi Met was synthesized by in vitro transcription with T7 polymerase as described previously ( ). .. His-tagged MetRS was purified from Escherichia coli XL1 Blue cells (Agilent) carrying the E. coli methionyl-tRNA synthetase (MetRS) expression plasmid pRA101 ( ) as described previously ( ).

Produced:

Article Title: Engineered Dengue Virus Domain III Proteins Elicit Cross-Neutralizing Antibody Responses in Mice
Article Snippet: .. EDIII-expressing phage particles were produced by electroporation into Escherichia coli XL1-Blue cells (Agilent Technologies, Santa Clara, CA) and recovered in 2× yeast extract tryptone (YT) medium containing 50 μg/ml carbenicillin and 10 μg/ml tetracycline at 37°C and 220 rpm for 5 h. Phage were coinfected with M13KO7 helper phage (New England BioLabs, Ipswich, MA) at a final concentration of 1010 PFU per ml and incubated for 1 h at 37°C and 220 rpm. .. Phage were harvested by precipitation with 4% (wt/vol) polyethylene glycol (PEG) 8000 and 3% (wt/vol) NaCl after centrifugation to remove cells.

Concentration Assay:

Article Title: Engineered Dengue Virus Domain III Proteins Elicit Cross-Neutralizing Antibody Responses in Mice
Article Snippet: .. EDIII-expressing phage particles were produced by electroporation into Escherichia coli XL1-Blue cells (Agilent Technologies, Santa Clara, CA) and recovered in 2× yeast extract tryptone (YT) medium containing 50 μg/ml carbenicillin and 10 μg/ml tetracycline at 37°C and 220 rpm for 5 h. Phage were coinfected with M13KO7 helper phage (New England BioLabs, Ipswich, MA) at a final concentration of 1010 PFU per ml and incubated for 1 h at 37°C and 220 rpm. .. Phage were harvested by precipitation with 4% (wt/vol) polyethylene glycol (PEG) 8000 and 3% (wt/vol) NaCl after centrifugation to remove cells.

Gel Extraction:

Article Title: Folding and activity of mutant cystathionine ?-synthase depends on the position and nature of the purification tag: Characterization of the R266K CBS mutant
Article Snippet: After cleavage with Apa I and Xho I (NEB Biolabs), the CBS fragment was cut out from a 1% agarose gel and cleaned up using the QIAquick gel extraction kit (Qiagen). .. All constructs were transformed into E. coli XL1-Blue cells (Agilent) and their authenticity was confirmed by DNA sequencing.

Positron Emission Tomography:

Article Title: Domain Organization, Catalysis and Regulation of Eukaryotic Cystathionine Beta-Synthases
Article Snippet: The Nco I- and Xho I-digested, gel-extracted PCR product was ligated into a similarly prepared pET-28a vector using T4 DNA ligase (NEB Biolabs). .. Both pET28-C-yCBS WT and pET28-C-yCBS L345* constructs were transformed into E. coli XL1-Blue cells (Agilent) and their authenticity and the presence of a C-terminal 6xHis tag was confirmed by DNA sequencing.

Article Title: Folding and activity of mutant cystathionine ?-synthase depends on the position and nature of the purification tag: Characterization of the R266K CBS mutant
Article Snippet: Subsequently, the CBS fragment was ligated into an Apa I- Xho I linearized modified pET-47b(+) vector. .. All constructs were transformed into E. coli XL1-Blue cells (Agilent) and their authenticity was confirmed by DNA sequencing.

Article Title: Domain Organization, Catalysis and Regulation of Eukaryotic Cystathionine Beta-Synthases
Article Snippet: The Nco I- and Hind III-digested, gel-extracted PCR product was ligated into a similarly prepared pET-28a vector using T4 DNA ligase (NEB Biolabs). .. Both yCBS and dCBS constructs, designated as pGEX-6P1-yCBS and pET28-C-dCBS, respectively, were transformed into E. coli XL1-Blue cells (Agilent) and their authenticity was confirmed by DNA sequencing.

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