vitro protein synthesis kit  (New England Biolabs)


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    Name:
    PURExpress In Vitro Protein Synthesis Kit
    Description:
    PURExpress In Vitro Protein Synthesis Kit 100 rxns
    Catalog Number:
    e6800l
    Price:
    2292
    Size:
    100 rxns
    Category:
    Protein Expression and Purification Kits
    Buy from Supplier


    Structured Review

    New England Biolabs vitro protein synthesis kit
    PURExpress In Vitro Protein Synthesis Kit
    PURExpress In Vitro Protein Synthesis Kit 100 rxns
    https://www.bioz.com/result/vitro protein synthesis kit/product/New England Biolabs
    Average 90 stars, based on 57 article reviews
    Price from $9.99 to $1999.99
    vitro protein synthesis kit - by Bioz Stars, 2020-01
    90/100 stars

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    Related Articles

    Clone Assay:

    Article Title: Foot-and-Mouth Disease (FMD) Virus 3C Protease Mutant L127P: Implications for FMD Vaccine Development
    Article Snippet: Nucleotide sequences for 3C(wt), 3C(L127P), and 3C(C163A) with N-terminal FLAG tags were also cloned into pSNAP-tag (T7)-2 vector (New England BioLabs) using cut sites NdeI and NotI. .. Cell-free protein synthesis used a PURExpress in vitro protein synthesis kit (New England BioLabs) with the modification that two DNA plasmids were added in equimolar amounts.

    Amplification:

    Article Title: A yeast two-hybrid system for the screening and characterization of small-molecule inhibitors of protein–protein interactions identifies a novel putative Mdm2-binding site in p53
    Article Snippet: In vitro transcription–translation The SBP-p53 and HA-Mdm2 ORF’s were amplified using SBP-p53-pET22b (+) and HA-Mdm2-pET22b (+) vectors as templates. .. These PCR products were used as substrates for in vitro translation (IVT) using PURExpress in vitro protein synthesis kit according to the instruction manual (NEB, Ipswich, MA, USA).

    Synthesized:

    Article Title: Roles of AtpI and Two YidC-Type Proteins from Alkaliphilic Bacillus pseudofirmus OF4 in ATP Synthase Assembly and Nonfermentative Growth
    Article Snippet: .. AtpI and/or c -subunit were synthesized in vitro with a PURExpress in vitro protein synthesis kit (New England BioLabs). .. Four hundred nanograms of plasmid DNA was added in a 25-μl reaction mixture of the PURE system.

    Article Title: Foot-and-Mouth Disease (FMD) Virus 3C Protease Mutant L127P: Implications for FMD Vaccine Development
    Article Snippet: The bovine eIF4AI gene (GenBank accession no. 77735406 ) was synthesized by GenScript with the addition of an N-terminal FLAG tag and a C-terminal myc tag and cloned into pSNAP-tag (T7)-2 vector (New England BioLabs) using cut sites NdeI and NotI. .. Cell-free protein synthesis used a PURExpress in vitro protein synthesis kit (New England BioLabs) with the modification that two DNA plasmids were added in equimolar amounts.

    Article Title: Accurate high-throughput screening based on digital protein synthesis in a massively parallel femtoliter droplet array
    Article Snippet: .. We synthesized proteins from single DNA molecules encapsulated in the droplets using a cell-free transcription and translation system (PURExpress In Vitro Protein Synthesis Kit, New England Biolabs). .. The protein synthesis working solution was composed of 4.0 μl of solution A, 3.0 μl of solution B, 0.2 μl of recombinant RNase inhibitor (40 U/μl; Takara-Bio), template DNA with desired concentrations, biliverdin (10 μM; Sigma-Aldrich) as required, disulfide bond enhancer 1 and disulfide bond enhancer 2 (0.4 μl for each, PURExpress Disulfide Bond Enhancer, New England Biolabs) as required, fluorogenic substrate (1.0 mM DiFMUP, Invitrogen; or 1 mM fluorescein diphosphate, AAT Bioquest) as required, and nuclease-free water (Invitrogen) to a final volume of 10 μl (the total volume may be arbitrarily scaled down or up).

    Article Title: PARAGEN 1.0: A Standardized Synthetic Gene Library for Fast Cell-Free Bacteriocin Synthesis
    Article Snippet: A total of 128 bacteriocins were synthesized, 23 belonging to the class IIa (18%), 15 belonging to the class IIb (12%), 15 belonging to the class IIc (12%), 50 belonging to the class IId (39%), and 25 belonging to the class III (20%) ( ). .. *Image from PURExpress® in vitro Protein Synthesis Kit (New England Biolabs).

    Incubation:

    Article Title: SecA mediates cotranslational targeting and translocation of an inner membrane protein
    Article Snippet: In vitro translocation assay in PURE system Translation was performed at 30°C using a PURExpress in vitro protein synthesis kit (New England Biolabs, Inc.) supplemented with [35 S]methionine (1.5 mCi/ml; PerkinElmer) and indicated concentrations of cytosolic factors (SecA, SecB, Ffh, FtsY, or TF). .. Digestion was stopped by addition of 5 mM PMSF, after which the sample was incubated on ice for 10 min.

    Article Title: Iroquois proteins promote skeletal joint formation by maintaining chondrocytes in an immature state
    Article Snippet: Zebrafish Irx5a and Sox9a and negative control GST proteins were translated using the PURExpress in vitro protein synthesis kit (NEB). .. Proteins and probes in binding buffer were incubated at room temperature for 30 minutes and then run on a 5% TBE polyacrylamide gel.

    Article Title: Ribosome Rescue Inhibitors Kill Actively Growing and Nonreplicating Persister Mycobacterium tuberculosis Cells
    Article Snippet: In Vitro Photolabeling and Click Conjugation Assays were set up using the PURExpress in vitro protein synthesis kit (NEB, Ipswich, MA) according to the manufacturer’s protocols. .. The reactions were performed with no DNA template, a nonstop DHFR template, the full length DHFR gene, full length DHFR gene with 0 bases after the stop codon, or full length DHFR gene with 33 bases after the stop codon., KKL-2098 (final concentration of 1 μM) was added to a mixture of assay components, and the samples were incubated at room temperature for 1 h. Samples were placed on ice, irradiated with 312 nm UV light for 10 min, and used to set up click conjugation assays in the presence of KKL-2107 (final concentration of 0.5 mM).

    Article Title: Identifying the targets of aminoacyl-tRNA synthetase inhibitors by primer extension inhibition
    Article Snippet: Translation The PCR-generated DNA template was expressed in a cell-free transcription-translation system (PURExpress In Vitro Protein Synthesis kit, New England BioLabs) ( ). .. Samples were incubated at 37°C for 20 min. For the reactions that were run at a reduced concentration of amino acids, a modified solution A ( ) was prepared to contain 125 mM Hepes-KOH (pH 7.6), 250 mM potassium glutamate, 15 mM magnesium acetate, 5 mM spermidine, 2.5 mM DTT, 25 µg/ml formyl donor (see later in the text), 50 mM creatine phosphate (Sigma), 5 mg/ml Escherichia coli tRNA (Roche), 15 µM each amino acid, 5 mM ATP, 5 mM GTP, 2.5 mM CTP, 2.5 mM UTP (pH of all the nucleotide triphosphate stocks was previously adjusted to 7.5).

    Article Title: Accurate high-throughput screening based on digital protein synthesis in a massively parallel femtoliter droplet array
    Article Snippet: The flush oil was equilibrated with buffer components of the cell-free system by vortex mixing for 30 s, incubation for 10 min, and centrifugal separation at 2 × 104 g for 5 min. .. We synthesized proteins from single DNA molecules encapsulated in the droplets using a cell-free transcription and translation system (PURExpress In Vitro Protein Synthesis Kit, New England Biolabs).

    Activity Assay:

    Article Title: PARAGEN 1.0: A Standardized Synthetic Gene Library for Fast Cell-Free Bacteriocin Synthesis
    Article Snippet: The remaining bacteriocins (59 out of 128) did not show activity against any of the indicators tested ( ). .. *Image from PURExpress® in vitro Protein Synthesis Kit (New England Biolabs).

    Expressing:

    Article Title: In vitro affinity screening of protein and peptide binders by megavalent bead surface display
    Article Snippet: .. Step 5 and 6—in vitro expression In vitro transcription and translation (IVTT) reactions were carried out using the PURExpress™ in vitro Protein Synthesis Kit (NEB). .. Reactions of 12.5 µl, consisted of 5 µl of solution A, 3.75 µl of solution B and plasmid or ePCR-amplified DNA on beads in water.

    Article Title: Droplet Microfluidics Approach for Single-DNA Molecule Amplification and Condensation into DNA-Magnesium-Pyrophosphate Particles
    Article Snippet: .. Coupled In Vitro Transcription and Translation β-galactosidase enzyme expression was performed using an IVTT system purchased from New England Biolabs (PURExpress® in vitro Protein Synthesis Kit, NEB, Ipswich, MA, USA) in the presence of RNase inhibitor Ribolock (Thermo Fisher Scientific, Vilnius, Lithuania). .. IVTT reaction in 384-well format: The in vitro expression of lacZ in a 384-well format (10 μL/well) was performed by preparing two sets of IVTT reaction mixtures.

    Modification:

    Article Title: Iroquois proteins promote skeletal joint formation by maintaining chondrocytes in an immature state
    Article Snippet: Zebrafish Irx5a and Sox9a and negative control GST proteins were translated using the PURExpress in vitro protein synthesis kit (NEB). .. Biotin-labeled probes were obtained from IDT (See , Sequences of modified R2 enhancers and probes for EMSA).

    Article Title: Identifying the targets of aminoacyl-tRNA synthetase inhibitors by primer extension inhibition
    Article Snippet: Translation The PCR-generated DNA template was expressed in a cell-free transcription-translation system (PURExpress In Vitro Protein Synthesis kit, New England BioLabs) ( ). .. Samples were incubated at 37°C for 20 min. For the reactions that were run at a reduced concentration of amino acids, a modified solution A ( ) was prepared to contain 125 mM Hepes-KOH (pH 7.6), 250 mM potassium glutamate, 15 mM magnesium acetate, 5 mM spermidine, 2.5 mM DTT, 25 µg/ml formyl donor (see later in the text), 50 mM creatine phosphate (Sigma), 5 mg/ml Escherichia coli tRNA (Roche), 15 µM each amino acid, 5 mM ATP, 5 mM GTP, 2.5 mM CTP, 2.5 mM UTP (pH of all the nucleotide triphosphate stocks was previously adjusted to 7.5).

    Article Title: Foot-and-Mouth Disease (FMD) Virus 3C Protease Mutant L127P: Implications for FMD Vaccine Development
    Article Snippet: .. Cell-free protein synthesis used a PURExpress in vitro protein synthesis kit (New England BioLabs) with the modification that two DNA plasmids were added in equimolar amounts. .. Western blots of cell-free synthesis products used anti-eIF4AI (ab31217, Abcam) antibody to detect eIF4AI and anti-DYKDDDDK (635691; TaKaRa) antibody to detect FLAG-tagged 3C.

    Western Blot:

    Article Title: Foot-and-Mouth Disease (FMD) Virus 3C Protease Mutant L127P: Implications for FMD Vaccine Development
    Article Snippet: Paragraph title: Cell-free protein synthesis and Western blotting. ... Cell-free protein synthesis used a PURExpress in vitro protein synthesis kit (New England BioLabs) with the modification that two DNA plasmids were added in equimolar amounts.

    Translocation Assay:

    Article Title: SecA mediates cotranslational targeting and translocation of an inner membrane protein
    Article Snippet: .. In vitro translocation assay in PURE system Translation was performed at 30°C using a PURExpress in vitro protein synthesis kit (New England Biolabs, Inc.) supplemented with [35 S]methionine (1.5 mCi/ml; PerkinElmer) and indicated concentrations of cytosolic factors (SecA, SecB, Ffh, FtsY, or TF). ..

    Conjugation Assay:

    Article Title: Ribosome Rescue Inhibitors Kill Actively Growing and Nonreplicating Persister Mycobacterium tuberculosis Cells
    Article Snippet: .. In Vitro Photolabeling and Click Conjugation Assays were set up using the PURExpress in vitro protein synthesis kit (NEB, Ipswich, MA) according to the manufacturer’s protocols. .. The reactions were performed with no DNA template, a nonstop DHFR template, the full length DHFR gene, full length DHFR gene with 0 bases after the stop codon, or full length DHFR gene with 33 bases after the stop codon., KKL-2098 (final concentration of 1 μM) was added to a mixture of assay components, and the samples were incubated at room temperature for 1 h. Samples were placed on ice, irradiated with 312 nm UV light for 10 min, and used to set up click conjugation assays in the presence of KKL-2107 (final concentration of 0.5 mM).

    Concentration Assay:

    Article Title: Ribosome Rescue Inhibitors Kill Actively Growing and Nonreplicating Persister Mycobacterium tuberculosis Cells
    Article Snippet: In Vitro Photolabeling and Click Conjugation Assays were set up using the PURExpress in vitro protein synthesis kit (NEB, Ipswich, MA) according to the manufacturer’s protocols. .. The reactions were performed with no DNA template, a nonstop DHFR template, the full length DHFR gene, full length DHFR gene with 0 bases after the stop codon, or full length DHFR gene with 33 bases after the stop codon., KKL-2098 (final concentration of 1 μM) was added to a mixture of assay components, and the samples were incubated at room temperature for 1 h. Samples were placed on ice, irradiated with 312 nm UV light for 10 min, and used to set up click conjugation assays in the presence of KKL-2107 (final concentration of 0.5 mM).

    Article Title: Identifying the targets of aminoacyl-tRNA synthetase inhibitors by primer extension inhibition
    Article Snippet: Translation The PCR-generated DNA template was expressed in a cell-free transcription-translation system (PURExpress In Vitro Protein Synthesis kit, New England BioLabs) ( ). .. Samples were incubated at 37°C for 20 min. For the reactions that were run at a reduced concentration of amino acids, a modified solution A ( ) was prepared to contain 125 mM Hepes-KOH (pH 7.6), 250 mM potassium glutamate, 15 mM magnesium acetate, 5 mM spermidine, 2.5 mM DTT, 25 µg/ml formyl donor (see later in the text), 50 mM creatine phosphate (Sigma), 5 mg/ml Escherichia coli tRNA (Roche), 15 µM each amino acid, 5 mM ATP, 5 mM GTP, 2.5 mM CTP, 2.5 mM UTP (pH of all the nucleotide triphosphate stocks was previously adjusted to 7.5).

    Transferring:

    Article Title: Accurate high-throughput screening based on digital protein synthesis in a massively parallel femtoliter droplet array
    Article Snippet: We synthesized proteins from single DNA molecules encapsulated in the droplets using a cell-free transcription and translation system (PURExpress In Vitro Protein Synthesis Kit, New England Biolabs). .. The reaction solution was injected into the microchannel by pipette, and a sharp chilling process was applied for a few seconds to remove air from the microchambers.

    Generated:

    Article Title: A yeast two-hybrid system for the screening and characterization of small-molecule inhibitors of protein–protein interactions identifies a novel putative Mdm2-binding site in p53
    Article Snippet: Truncated (1–52, 1–116, 1–143, and 1–160) and mutated p53 were generated by overlap PCR. .. These PCR products were used as substrates for in vitro translation (IVT) using PURExpress in vitro protein synthesis kit according to the instruction manual (NEB, Ipswich, MA, USA).

    Polymerase Chain Reaction:

    Article Title: A yeast two-hybrid system for the screening and characterization of small-molecule inhibitors of protein–protein interactions identifies a novel putative Mdm2-binding site in p53
    Article Snippet: .. These PCR products were used as substrates for in vitro translation (IVT) using PURExpress in vitro protein synthesis kit according to the instruction manual (NEB, Ipswich, MA, USA). .. Immunoprecipitation IVT-expressed wild-type, truncated, or mutant p53 were pre-cleared with Protein G beads (Invitrogen, Carlsbad, CA, USA), followed by incubation with IVT-expressed Mdm2 at room temperature for 1 h. Anti-HA 3 F10 antibody (Roche Life Science, USA) at 1:200 dilution was added into the p53/Mdm2 mixture and incubated at 4 °C for 1 h. Protein G beads were pre-blocked with 3% BSA/PBS and then added into the p53-Mdm2-anti-HA 3 F10 mixture.

    Article Title: Identifying the targets of aminoacyl-tRNA synthetase inhibitors by primer extension inhibition
    Article Snippet: .. Translation The PCR-generated DNA template was expressed in a cell-free transcription-translation system (PURExpress In Vitro Protein Synthesis kit, New England BioLabs) ( ). .. For a typical reaction, 2 µl of solution A (kit), 1 µl of solution B (kit), 0.5 µl of DNA template (0.2 pmol/µl), 0.5 µl of radioactive primer (1 pmol), 0.2 µl of Ribolock RNAse inhibitor (40 U/µl, Thermo Scientific), 0.5 µl of the compound to be tested (10X solution in H2 O) and 0.3 µl of H2 O were combined in the reaction tube chilled on ice.

    Article Title: A combined cryo-EM and molecular dynamics approach reveals the mechanism of ErmBL-mediated translation arrest
    Article Snippet: Toe-printing assay The position of the ribosome on the mRNA was monitored with a toe-printing assay based on an in vitro -coupled transcription–translation system with the PURExpress in vitro Protein Synthesis Kit (NEB) . .. All templates were synthetized via PCR using overlapping forward and reverse primers containing the designated mutation in the Lys position of the ermBL motif.

    Sonication:

    Article Title: Roles of AtpI and Two YidC-Type Proteins from Alkaliphilic Bacillus pseudofirmus OF4 in ATP Synthase Assembly and Nonfermentative Growth
    Article Snippet: AtpI and/or c -subunit were synthesized in vitro with a PURExpress in vitro protein synthesis kit (New England BioLabs). .. The procedures were carried out according to reference and included 0.6 mg/ml liposomes made from soybean phosphatidylcholine (Type II-S; Sigma) by sonication and freeze-thaw.

    Injection:

    Article Title: Accurate high-throughput screening based on digital protein synthesis in a massively parallel femtoliter droplet array
    Article Snippet: We synthesized proteins from single DNA molecules encapsulated in the droplets using a cell-free transcription and translation system (PURExpress In Vitro Protein Synthesis Kit, New England Biolabs). .. The reaction solution was injected into the microchannel by pipette, and a sharp chilling process was applied for a few seconds to remove air from the microchambers.

    Binding Assay:

    Article Title: Iroquois proteins promote skeletal joint formation by maintaining chondrocytes in an immature state
    Article Snippet: Zebrafish Irx5a and Sox9a and negative control GST proteins were translated using the PURExpress in vitro protein synthesis kit (NEB). .. Proteins and probes in binding buffer were incubated at room temperature for 30 minutes and then run on a 5% TBE polyacrylamide gel.

    Mutagenesis:

    Article Title: A combined cryo-EM and molecular dynamics approach reveals the mechanism of ErmBL-mediated translation arrest
    Article Snippet: Toe-printing assay The position of the ribosome on the mRNA was monitored with a toe-printing assay based on an in vitro -coupled transcription–translation system with the PURExpress in vitro Protein Synthesis Kit (NEB) . .. All templates were synthetized via PCR using overlapping forward and reverse primers containing the designated mutation in the Lys position of the ermBL motif.

    Electrophoretic Mobility Shift Assay:

    Article Title: Iroquois proteins promote skeletal joint formation by maintaining chondrocytes in an immature state
    Article Snippet: Paragraph title: Electrophoretic Mobility Shift Assay ... Zebrafish Irx5a and Sox9a and negative control GST proteins were translated using the PURExpress in vitro protein synthesis kit (NEB).

    Purification:

    Article Title: Droplet Microfluidics Approach for Single-DNA Molecule Amplification and Condensation into DNA-Magnesium-Pyrophosphate Particles
    Article Snippet: Coupled In Vitro Transcription and Translation β-galactosidase enzyme expression was performed using an IVTT system purchased from New England Biolabs (PURExpress® in vitro Protein Synthesis Kit, NEB, Ipswich, MA, USA) in the presence of RNase inhibitor Ribolock (Thermo Fisher Scientific, Vilnius, Lithuania). .. The second set of reactions contained purified DNA particles diluted down to 46,000, 23,000 and 5000 particles per well.

    SDS Page:

    Article Title: Roles of AtpI and Two YidC-Type Proteins from Alkaliphilic Bacillus pseudofirmus OF4 in ATP Synthase Assembly and Nonfermentative Growth
    Article Snippet: AtpI and/or c -subunit were synthesized in vitro with a PURExpress in vitro protein synthesis kit (New England BioLabs). .. After the reaction, the synthesized proteins were directly analyzed by SDS-PAGE or treated with proteinase K and RNase in order to remove ribosomes and all of the soluble proteins and then precipitated with 50 mM MgCl2 before SDS-PAGE analysis ( ).

    Article Title: Protein Synthesis Using A Reconstituted Cell-Free System
    Article Snippet: .. A PURExpress In vitro Protein Synthesis kit (New England Biolabs, ) containing: Solution A (yellow tube) Solution B (red tube) DHFR control template PURExpress Disulfide Bond Enhancer (New England Biolabs, ) containing: Disulfide Bond Enhancer Solution 1 Disulfide Bond Enhancer Solution 2 microcentrifuge tubes or microtiter plate Nuclease-free H2 O (Integrated DNA technologies) Template DNA (See Support Protocol 1 and 2) or mRNA (See Support Protocol 3) Murine RNase inhibitor (40 U/µl, New England Biolabs) or RNasin Ribonuclease inhibitor (20–40 U/µl, Promega) Microcentrifuge Air incubator set at 37°C 3x SDS-PAGE loading buffer (New England Biolabs, CPMB Chapter X) SDS-PAGE gel (4–20% Tris-glycine, Life Technologies, CPMB Chapter X) ..

    Article Title: Protein Synthesis Using A Reconstituted Cell-Free System
    Article Snippet: .. PURExpress In vitro Protein Synthesis kit (New England Biolabs) containing: Solution A (yellow tube) Solution B (red tube) DHFR control template Nuclease free microcentrifuge tubes or microtiter plates Nuclease free H2 O Template DNA (See Support Protocol 1 and 2) or mRNA (See Support Protocol 3) Murine RNase inhibitor (40 U/µl, New England Biolabs) [35 S]-L-Methionine (15 mCi/ml, 1000 Ci/mmol) (PerkinElmer) Microcentrifuge Air incubator set at 37°C 3x SDS-PAGE loading buffer (New England Biolabs) SDS-PAGE gel (4–20 % Tris-glycine, Life Technologies) Filter paper (Whatman) Vacuum gel dryer X-ray film or phosphorimager ..

    Plasmid Preparation:

    Article Title: Roles of AtpI and Two YidC-Type Proteins from Alkaliphilic Bacillus pseudofirmus OF4 in ATP Synthase Assembly and Nonfermentative Growth
    Article Snippet: AtpI and/or c -subunit were synthesized in vitro with a PURExpress in vitro protein synthesis kit (New England BioLabs). .. Four hundred nanograms of plasmid DNA was added in a 25-μl reaction mixture of the PURE system.

    Article Title: In vitro affinity screening of protein and peptide binders by megavalent bead surface display
    Article Snippet: Step 5 and 6—in vitro expression In vitro transcription and translation (IVTT) reactions were carried out using the PURExpress™ in vitro Protein Synthesis Kit (NEB). .. Reactions of 12.5 µl, consisted of 5 µl of solution A, 3.75 µl of solution B and plasmid or ePCR-amplified DNA on beads in water.

    Article Title: Foot-and-Mouth Disease (FMD) Virus 3C Protease Mutant L127P: Implications for FMD Vaccine Development
    Article Snippet: Nucleotide sequences for 3C(wt), 3C(L127P), and 3C(C163A) with N-terminal FLAG tags were also cloned into pSNAP-tag (T7)-2 vector (New England BioLabs) using cut sites NdeI and NotI. .. Cell-free protein synthesis used a PURExpress in vitro protein synthesis kit (New England BioLabs) with the modification that two DNA plasmids were added in equimolar amounts.

    Article Title: Droplet Microfluidics Approach for Single-DNA Molecule Amplification and Condensation into DNA-Magnesium-Pyrophosphate Particles
    Article Snippet: Coupled In Vitro Transcription and Translation β-galactosidase enzyme expression was performed using an IVTT system purchased from New England Biolabs (PURExpress® in vitro Protein Synthesis Kit, NEB, Ipswich, MA, USA) in the presence of RNase inhibitor Ribolock (Thermo Fisher Scientific, Vilnius, Lithuania). .. The first set of reactions was supplemented with 0.5 to 500 ng of pIVEX2.2-lacZ-his plasmid, which translated into 1011 –108 copies of free DNA molecules per 10 μL.

    Irradiation:

    Article Title: Ribosome Rescue Inhibitors Kill Actively Growing and Nonreplicating Persister Mycobacterium tuberculosis Cells
    Article Snippet: In Vitro Photolabeling and Click Conjugation Assays were set up using the PURExpress in vitro protein synthesis kit (NEB, Ipswich, MA) according to the manufacturer’s protocols. .. The reactions were performed with no DNA template, a nonstop DHFR template, the full length DHFR gene, full length DHFR gene with 0 bases after the stop codon, or full length DHFR gene with 33 bases after the stop codon., KKL-2098 (final concentration of 1 μM) was added to a mixture of assay components, and the samples were incubated at room temperature for 1 h. Samples were placed on ice, irradiated with 312 nm UV light for 10 min, and used to set up click conjugation assays in the presence of KKL-2107 (final concentration of 0.5 mM).

    Negative Control:

    Article Title: Iroquois proteins promote skeletal joint formation by maintaining chondrocytes in an immature state
    Article Snippet: .. Zebrafish Irx5a and Sox9a and negative control GST proteins were translated using the PURExpress in vitro protein synthesis kit (NEB). .. Biotin-labeled probes were obtained from IDT (See , Sequences of modified R2 enhancers and probes for EMSA).

    Recombinant:

    Article Title: Accurate high-throughput screening based on digital protein synthesis in a massively parallel femtoliter droplet array
    Article Snippet: We synthesized proteins from single DNA molecules encapsulated in the droplets using a cell-free transcription and translation system (PURExpress In Vitro Protein Synthesis Kit, New England Biolabs). .. The protein synthesis working solution was composed of 4.0 μl of solution A, 3.0 μl of solution B, 0.2 μl of recombinant RNase inhibitor (40 U/μl; Takara-Bio), template DNA with desired concentrations, biliverdin (10 μM; Sigma-Aldrich) as required, disulfide bond enhancer 1 and disulfide bond enhancer 2 (0.4 μl for each, PURExpress Disulfide Bond Enhancer, New England Biolabs) as required, fluorogenic substrate (1.0 mM DiFMUP, Invitrogen; or 1 mM fluorescein diphosphate, AAT Bioquest) as required, and nuclease-free water (Invitrogen) to a final volume of 10 μl (the total volume may be arbitrarily scaled down or up).

    In Vitro:

    Article Title: A yeast two-hybrid system for the screening and characterization of small-molecule inhibitors of protein–protein interactions identifies a novel putative Mdm2-binding site in p53
    Article Snippet: .. These PCR products were used as substrates for in vitro translation (IVT) using PURExpress in vitro protein synthesis kit according to the instruction manual (NEB, Ipswich, MA, USA). .. Immunoprecipitation IVT-expressed wild-type, truncated, or mutant p53 were pre-cleared with Protein G beads (Invitrogen, Carlsbad, CA, USA), followed by incubation with IVT-expressed Mdm2 at room temperature for 1 h. Anti-HA 3 F10 antibody (Roche Life Science, USA) at 1:200 dilution was added into the p53/Mdm2 mixture and incubated at 4 °C for 1 h. Protein G beads were pre-blocked with 3% BSA/PBS and then added into the p53-Mdm2-anti-HA 3 F10 mixture.

    Article Title: SecA mediates cotranslational targeting and translocation of an inner membrane protein
    Article Snippet: .. In vitro translocation assay in PURE system Translation was performed at 30°C using a PURExpress in vitro protein synthesis kit (New England Biolabs, Inc.) supplemented with [35 S]methionine (1.5 mCi/ml; PerkinElmer) and indicated concentrations of cytosolic factors (SecA, SecB, Ffh, FtsY, or TF). ..

    Article Title: Roles of AtpI and Two YidC-Type Proteins from Alkaliphilic Bacillus pseudofirmus OF4 in ATP Synthase Assembly and Nonfermentative Growth
    Article Snippet: .. AtpI and/or c -subunit were synthesized in vitro with a PURExpress in vitro protein synthesis kit (New England BioLabs). .. Four hundred nanograms of plasmid DNA was added in a 25-μl reaction mixture of the PURE system.

    Article Title: Iroquois proteins promote skeletal joint formation by maintaining chondrocytes in an immature state
    Article Snippet: .. Zebrafish Irx5a and Sox9a and negative control GST proteins were translated using the PURExpress in vitro protein synthesis kit (NEB). .. Biotin-labeled probes were obtained from IDT (See , Sequences of modified R2 enhancers and probes for EMSA).

    Article Title: Ribosome Rescue Inhibitors Kill Actively Growing and Nonreplicating Persister Mycobacterium tuberculosis Cells
    Article Snippet: .. In Vitro Photolabeling and Click Conjugation Assays were set up using the PURExpress in vitro protein synthesis kit (NEB, Ipswich, MA) according to the manufacturer’s protocols. .. The reactions were performed with no DNA template, a nonstop DHFR template, the full length DHFR gene, full length DHFR gene with 0 bases after the stop codon, or full length DHFR gene with 33 bases after the stop codon., KKL-2098 (final concentration of 1 μM) was added to a mixture of assay components, and the samples were incubated at room temperature for 1 h. Samples were placed on ice, irradiated with 312 nm UV light for 10 min, and used to set up click conjugation assays in the presence of KKL-2107 (final concentration of 0.5 mM).

    Article Title: In vitro affinity screening of protein and peptide binders by megavalent bead surface display
    Article Snippet: .. Step 5 and 6—in vitro expression In vitro transcription and translation (IVTT) reactions were carried out using the PURExpress™ in vitro Protein Synthesis Kit (NEB). .. Reactions of 12.5 µl, consisted of 5 µl of solution A, 3.75 µl of solution B and plasmid or ePCR-amplified DNA on beads in water.

    Article Title: Identifying the targets of aminoacyl-tRNA synthetase inhibitors by primer extension inhibition
    Article Snippet: .. Translation The PCR-generated DNA template was expressed in a cell-free transcription-translation system (PURExpress In Vitro Protein Synthesis kit, New England BioLabs) ( ). .. For a typical reaction, 2 µl of solution A (kit), 1 µl of solution B (kit), 0.5 µl of DNA template (0.2 pmol/µl), 0.5 µl of radioactive primer (1 pmol), 0.2 µl of Ribolock RNAse inhibitor (40 U/µl, Thermo Scientific), 0.5 µl of the compound to be tested (10X solution in H2 O) and 0.3 µl of H2 O were combined in the reaction tube chilled on ice.

    Article Title: Protein Synthesis Using A Reconstituted Cell-Free System
    Article Snippet: .. A PURExpress In vitro Protein Synthesis kit (New England Biolabs, ) containing: Solution A (yellow tube) Solution B (red tube) DHFR control template PURExpress Disulfide Bond Enhancer (New England Biolabs, ) containing: Disulfide Bond Enhancer Solution 1 Disulfide Bond Enhancer Solution 2 microcentrifuge tubes or microtiter plate Nuclease-free H2 O (Integrated DNA technologies) Template DNA (See Support Protocol 1 and 2) or mRNA (See Support Protocol 3) Murine RNase inhibitor (40 U/µl, New England Biolabs) or RNasin Ribonuclease inhibitor (20–40 U/µl, Promega) Microcentrifuge Air incubator set at 37°C 3x SDS-PAGE loading buffer (New England Biolabs, CPMB Chapter X) SDS-PAGE gel (4–20% Tris-glycine, Life Technologies, CPMB Chapter X) ..

    Article Title: Protein Synthesis Using A Reconstituted Cell-Free System
    Article Snippet: .. PURExpress In vitro Protein Synthesis kit (New England Biolabs) containing: Solution A (yellow tube) Solution B (red tube) DHFR control template Nuclease free microcentrifuge tubes or microtiter plates Nuclease free H2 O Template DNA (See Support Protocol 1 and 2) or mRNA (See Support Protocol 3) Murine RNase inhibitor (40 U/µl, New England Biolabs) [35 S]-L-Methionine (15 mCi/ml, 1000 Ci/mmol) (PerkinElmer) Microcentrifuge Air incubator set at 37°C 3x SDS-PAGE loading buffer (New England Biolabs) SDS-PAGE gel (4–20 % Tris-glycine, Life Technologies) Filter paper (Whatman) Vacuum gel dryer X-ray film or phosphorimager ..

    Article Title: A combined cryo-EM and molecular dynamics approach reveals the mechanism of ErmBL-mediated translation arrest
    Article Snippet: .. Toe-printing assay The position of the ribosome on the mRNA was monitored with a toe-printing assay based on an in vitro -coupled transcription–translation system with the PURExpress in vitro Protein Synthesis Kit (NEB) . ..

    Article Title: Foot-and-Mouth Disease (FMD) Virus 3C Protease Mutant L127P: Implications for FMD Vaccine Development
    Article Snippet: .. Cell-free protein synthesis used a PURExpress in vitro protein synthesis kit (New England BioLabs) with the modification that two DNA plasmids were added in equimolar amounts. .. Western blots of cell-free synthesis products used anti-eIF4AI (ab31217, Abcam) antibody to detect eIF4AI and anti-DYKDDDDK (635691; TaKaRa) antibody to detect FLAG-tagged 3C.

    Article Title: Accurate high-throughput screening based on digital protein synthesis in a massively parallel femtoliter droplet array
    Article Snippet: .. We synthesized proteins from single DNA molecules encapsulated in the droplets using a cell-free transcription and translation system (PURExpress In Vitro Protein Synthesis Kit, New England Biolabs). .. The protein synthesis working solution was composed of 4.0 μl of solution A, 3.0 μl of solution B, 0.2 μl of recombinant RNase inhibitor (40 U/μl; Takara-Bio), template DNA with desired concentrations, biliverdin (10 μM; Sigma-Aldrich) as required, disulfide bond enhancer 1 and disulfide bond enhancer 2 (0.4 μl for each, PURExpress Disulfide Bond Enhancer, New England Biolabs) as required, fluorogenic substrate (1.0 mM DiFMUP, Invitrogen; or 1 mM fluorescein diphosphate, AAT Bioquest) as required, and nuclease-free water (Invitrogen) to a final volume of 10 μl (the total volume may be arbitrarily scaled down or up).

    Article Title: Protein Synthesis Using A Reconstituted Cell-Free System
    Article Snippet: .. A PURExpress In vitro Protein Synthesis kit (New England Biolabs, ) 10 mM magnesium acetate Amicon Ultracel −0.5 ml-100 K MW cut off spin concentrator (Millipore) Microcentrifuge at 4°C Ni-NTA agarose (Qiagen) Microcentrifuge tubes Bio-Rad micro-spin column (Bio-Rad) ..

    Article Title: PARAGEN 1.0: A Standardized Synthetic Gene Library for Fast Cell-Free Bacteriocin Synthesis
    Article Snippet: .. *Image from PURExpress® in vitro Protein Synthesis Kit (New England Biolabs). .. Strains used as indicators to test the antimicrobial activity of the bacteriocins from PARAGEN 1.0.

    Article Title: Droplet Microfluidics Approach for Single-DNA Molecule Amplification and Condensation into DNA-Magnesium-Pyrophosphate Particles
    Article Snippet: .. Coupled In Vitro Transcription and Translation β-galactosidase enzyme expression was performed using an IVTT system purchased from New England Biolabs (PURExpress® in vitro Protein Synthesis Kit, NEB, Ipswich, MA, USA) in the presence of RNase inhibitor Ribolock (Thermo Fisher Scientific, Vilnius, Lithuania). .. IVTT reaction in 384-well format: The in vitro expression of lacZ in a 384-well format (10 μL/well) was performed by preparing two sets of IVTT reaction mixtures.

    FLAG-tag:

    Article Title: Foot-and-Mouth Disease (FMD) Virus 3C Protease Mutant L127P: Implications for FMD Vaccine Development
    Article Snippet: The bovine eIF4AI gene (GenBank accession no. 77735406 ) was synthesized by GenScript with the addition of an N-terminal FLAG tag and a C-terminal myc tag and cloned into pSNAP-tag (T7)-2 vector (New England BioLabs) using cut sites NdeI and NotI. .. Cell-free protein synthesis used a PURExpress in vitro protein synthesis kit (New England BioLabs) with the modification that two DNA plasmids were added in equimolar amounts.

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    New England Biolabs purexpress in vitro protein synthesis kit
    Purexpress In Vitro Protein Synthesis Kit, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 90/100, based on 66 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Average 90 stars, based on 66 article reviews
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