talon superflow metal affinity resin  (TaKaRa)

 
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    Name:
    TALON Superflow Metal Affinity Resin
    Description:
    TALON his tag purification resin lets you prepare exceptionally pure his tagged proteins from bacterial mammalian yeast and baculovirus infected cells under native or denaturing conditions TALON is an immobilized metal affinity chromatography IMAC resin charged with cobalt which binds to his tagged proteins with higher specificity than nickel charged resins As a result TALON resin delivers his tagged proteins of the highest purity In addition each cobalt ion is bound to the resin at four sites resulting in low metal ion leakage
    Catalog Number:
    635670
    Price:
    None
    Size:
    250 mL
    Category:
    Cobalt resin FPLC Resin in FPLC cartridges His tagged protein purification Purification products Protein research
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    Structured Review

    TaKaRa talon superflow metal affinity resin
    TALON his tag purification resin lets you prepare exceptionally pure his tagged proteins from bacterial mammalian yeast and baculovirus infected cells under native or denaturing conditions TALON is an immobilized metal affinity chromatography IMAC resin charged with cobalt which binds to his tagged proteins with higher specificity than nickel charged resins As a result TALON resin delivers his tagged proteins of the highest purity In addition each cobalt ion is bound to the resin at four sites resulting in low metal ion leakage
    https://www.bioz.com/result/talon superflow metal affinity resin/product/TaKaRa
    Average 90 stars, based on 30 article reviews
    Price from $9.99 to $1999.99
    talon superflow metal affinity resin - by Bioz Stars, 2020-01
    90/100 stars

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    Related Articles

    Clone Assay:

    Article Title: Structure of a helicase-helicase loader complex reveals insights into the mechanism of bacterial primosome assembly
    Article Snippet: The helicase loader protein DnaI fragment was cloned from B. subtilis 168 genomic DNA (American Type Culture Collection) using PCR with primers (Integrated DNA Technologies): forward 5′-gcgaccatggaaccaatcggccgttcc-3′ and reverse 5′-gcgactcgagtggatgtcggcggttttctc-3′, and inserted into the Nco I–Xho I site of plasmid pET21d(+) (Novagen) and expressed in B834(DE3) (Novagen). .. The supernatant of the lysed cell paste solution was filtered with a 0.22-μM filter unit (Millipore) and loaded onto a 10-ml Talon superflow metal affinity resin (Clontech) mounted in a 5.0 × 20 cm2 Econo column (Bio-rad).

    Article Title: Modular Organization of ?-Toxins from Scorpion Venom Mirrors Domain Structure of Their Targets, Sodium Channels *
    Article Snippet: The toxin DNA was cloned into the expression vector pET-32b (Novagen (Madison, WI)), which was then used to transform Escherichia coli BL21(DE3). .. Trx-BeM9 fusion protein was isolated by affinity chromatography on TALON Superflow Metal Affinity Resin (Clontech, Mountain View, CA) and then hydrolyzed using CNBr as suggested ( ).

    Article Title: Structural Characterization of Proline-rich Tyrosine Kinase 2 (PYK2) Reveals a Unique (DFG-out) Conformation and Enables Inhibitor Design
    Article Snippet: The 854-bp DNA fragment was digested with BamHI and XhoI and cloned into the modified pFastBac vector at these sites to yield an N-terminal His6 -tagged thrombin cleavable fusion with PYK2 catalytic domain residues Pro-416 to Glu-692 based on accession . .. The supernatant was extracted and dispensed into a tube containing 20 ml of TALON Superflow Metal Affinity Resin (BD-Clonetech), pre-equilibrated with buffer B (100 m m KH2 PO4 /K2 HPO4 ; pH 8.0, 200 m m NaCl, 1 m m TCEP, 2 m m phenylmethylsulfonyl fluoride, 5 m m imidazole; pH 7.5).

    Article Title: Gene Expression and Molecular Characterization of a Xylanase from Chicken Cecum Metagenome
    Article Snippet: TALON Superflow Metal Affinity Resin (Clonetech) was purchased from TaKaRa (Otsu, Japan). .. The expression vector pET-32a (Novagen) was used for cloning and expressing the xylanase.

    Centrifugation:

    Article Title: Bacillus subtilis Intramembrane Protease RasP Activity in Escherichia coli and In Vitro
    Article Snippet: The supernatant was mixed with 1 ml Talon Superflow metal affinity resin (Clontech) that had been equilibrated with buffer (PBS containing 1% DM, 5 mM 2-mercaptoethanol, and 10% glycerol). .. The cobalt resin was sedimented by centrifugation at 708 × g for 2 min at 4°C.

    Article Title: Human APE2 protein is mostly localized in the nuclei and to some extent in the mitochondria, while nuclear APE2 is partly associated with proliferating cell nuclear antigen
    Article Snippet: Cell lysates were clarified by centrifugation at 100 000 g for 20 min at room temperature. .. Two milliliters of a 50% suspension of TALON superflow metal affinity resin (Clontech Laboratories) equilibrated with lysis buffer was mixed with each cell lysate and gently rotated for 1 h. Then the resin was washed five times with binding buffer (20 mM sodium phosphate buffer pH 7.4, 0.1% BSA) containing 50 mM NaCl and used for pull-down binding assays.

    Article Title: Rapid antibody selection by mRNA display on a microfluidic chip
    Article Snippet: For purification of proteins, the cells were collected by centrifugation, and resuspended in 20 ml of TBS (20 mM Tris–HCl buffer, pH 7.5, 138 mM NaCl) containing 8 U of DNase I (Invitrogen), 40 µl of EDTA-free protease inhibitor cocktail (Nacalai tesque) and 1 mM 2-mercaptoethanol (Nacalai tesque). .. The supernatants containing the histidine-tagged proteins in denatured form were immobilized on the TALON superflow metal affinity resin (Clontech), and the columns were washed with 10 volumes of TBS containing 10 mM imidazole and 6 M urea, 10 volumes of TBS containing 1 M NaCl and 10 volumes of TBS, to allow refolding of the bound proteins on the columns.

    Article Title: Creation of a type IIS restriction endonuclease with a long recognition sequence
    Article Snippet: Cellular debris was removed by centrifugation for 2 min at 16 000g , 4°C. .. Thirty microliters of 50% slurry of TALON Superflow Metal Affinity Resin (Clontech, Mountain View, CA), pre-equilibrated with 50 mM sodium phosphate, pH 7.0, 0.5 M NaCl and 50 mM MgCl2 , were added to each, and the mixtures were rocked for an hour at 4°C.

    Article Title: Structural Characterization of Proline-rich Tyrosine Kinase 2 (PYK2) Reveals a Unique (DFG-out) Conformation and Enables Inhibitor Design
    Article Snippet: The lysate was clarified by centrifugation at 4 °C for 35 min at 30,000 × g (16,000 rpm) in a Sorval SL-50T (or appropriate) rotor. .. The supernatant was extracted and dispensed into a tube containing 20 ml of TALON Superflow Metal Affinity Resin (BD-Clonetech), pre-equilibrated with buffer B (100 m m KH2 PO4 /K2 HPO4 ; pH 8.0, 200 m m NaCl, 1 m m TCEP, 2 m m phenylmethylsulfonyl fluoride, 5 m m imidazole; pH 7.5).

    Article Title: Generation of a More Immunogenic Measles Vaccine by Increasing Its Hemagglutinin Expression
    Article Snippet: Supernatants and cells were collected at 24-h intervals and lysed by one freeze-thaw cycle, and supernatants were clarified by centrifugation at 3,000 × g for 5 min in an SX4750 rotor. .. Talon Superflow metal affinity resin (Clontech Laboratories, Mountain View, CA) equilibrated in interaction buffer (50 mM NaH2 PO4 [pH 8], 200 mM NaCl, 1% Triton X-100) was added to the clarified supernatants for 2 h of batch interaction.

    Article Title: Interaction of intramembrane metalloprotease SpoIVFB with substrate Pro-σK
    Article Snippet: The rest was mixed with 0.5 mL of Talon superflow metal affinity resin (Clontech) that had been equilibrated with PBS containing 0.1% DM, 5 mM 2-mercaptoethanol, and 10% glycerol. .. The cobalt resin was sedimented by centrifugation at 708 × g for 2 min at 4 °C.

    Article Title: The structural basis of CstF-77 modulation of cleavage and polyadenylation through stimulation of CstF-64 activity
    Article Snippet: .. Lysates were clarified by centrifugation at 16 000 × g for 20 min at 4°C and loaded on home-packed columns either with an Amylose High Flow resin (New England Biolabs) or TALON Superflow Metal Affinity resin (TaKaRa) pre-equilibrated with the corresponding lysis buffers using peristaltic pump. .. Bound protein was washed with about 10 column volumes in the corresponding lysis buffer followed by a wash with about five column volumes in buffer containing 0.3 M NaCl, 20 mM HEPES pH 7.5, 5 mM imidazole, 0.05% sodium azide.

    Filtration:

    Article Title: Structure of a helicase-helicase loader complex reveals insights into the mechanism of bacterial primosome assembly
    Article Snippet: The supernatant of the lysed cell paste solution was filtered with a 0.22-μM filter unit (Millipore) and loaded onto a 10-ml Talon superflow metal affinity resin (Clontech) mounted in a 5.0 × 20 cm2 Econo column (Bio-rad). .. The eluate was then loaded onto a 16/60 or 26/60 Superdex G200 prep grade gel filtration column (GE Healthcare) equilibrated in buffer C (20 mM TRIS pH 8.0, 50 mM sodium chloride).

    Construct:

    Article Title: The structural basis of CstF-77 modulation of cleavage and polyadenylation through stimulation of CstF-64 activity
    Article Snippet: One-liter cultures were induced with 0.5 mM isopropyl-β-d-thiogalactopyranoside (IPTG) at an optical density of 0.8 at 37°C for 4 h. Bacterial cells were suspended in either 1 M NaCl, 25 mM Tris–HCl pH 7.5, 1 mM EDTA, 0.05% sodium azide for the constructs containing the maltose binding protein (see above) or 1 M NaCl, 20 mM HEPES pH 7.5, 5 mM Imidazole, 0.05% sodium azide for the RRM. .. Lysates were clarified by centrifugation at 16 000 × g for 20 min at 4°C and loaded on home-packed columns either with an Amylose High Flow resin (New England Biolabs) or TALON Superflow Metal Affinity resin (TaKaRa) pre-equilibrated with the corresponding lysis buffers using peristaltic pump.

    Incubation:

    Article Title: Human APE2 protein is mostly localized in the nuclei and to some extent in the mitochondria, while nuclear APE2 is partly associated with proliferating cell nuclear antigen
    Article Snippet: BL21(DE3) E.coli cells carrying plasmid pET32a(+), pET3228D:PCNA or pET32a(+):APE2, which express His tagged proteins TrxA, His–PCNA and TrxA–APE2, respectively, were cultured in 250 ml of M9ZB medium ( ) until the OD600 reached 0.6 and then incubated with 1 mM isopropyl β- d -thiogalactoside for a further 2 h. Cells were harvested by centrifugation, resuspended in 25 ml of lysis buffer [50 mM sodium phosphate buffer pH 7.0, 300 mM NaCl, 1% Triton X-100, 8 M urea, 1 mM 2-mercaptoethanol (2-ME), 1 mM PMSF, 1 µg/ml leupeptin, 1 µg/ml pepstatin A and 1 µg/ml chymostatin] and disrupted by sonication. .. Two milliliters of a 50% suspension of TALON superflow metal affinity resin (Clontech Laboratories) equilibrated with lysis buffer was mixed with each cell lysate and gently rotated for 1 h. Then the resin was washed five times with binding buffer (20 mM sodium phosphate buffer pH 7.4, 0.1% BSA) containing 50 mM NaCl and used for pull-down binding assays.

    Article Title: Structural Characterization of Proline-rich Tyrosine Kinase 2 (PYK2) Reveals a Unique (DFG-out) Conformation and Enables Inhibitor Design
    Article Snippet: The supernatant was extracted and dispensed into a tube containing 20 ml of TALON Superflow Metal Affinity Resin (BD-Clonetech), pre-equilibrated with buffer B (100 m m KH2 PO4 /K2 HPO4 ; pH 8.0, 200 m m NaCl, 1 m m TCEP, 2 m m phenylmethylsulfonyl fluoride, 5 m m imidazole; pH 7.5). .. The suspension was incubated at 4 °C for 3 h, with gentle rocking, then centrifuged at 700 × g in a swinging bucket rotor for 2-4 min, and the supernatant was extracted.

    Infection:

    Article Title: Generation of a More Immunogenic Measles Vaccine by Increasing Its Hemagglutinin Expression
    Article Snippet: To isolate 6×His-tagged soluble H protein from the supernatants of MVvac2-Hsol-infected cells, 100-mm-diameter dishes were seeded with 1 × 106 Vero/hSLAM cells and infected with MVvac2 or MVvac2-Hsol at an MOI of 0.1. .. Talon Superflow metal affinity resin (Clontech Laboratories, Mountain View, CA) equilibrated in interaction buffer (50 mM NaH2 PO4 [pH 8], 200 mM NaCl, 1% Triton X-100) was added to the clarified supernatants for 2 h of batch interaction.

    Expressing:

    Article Title: RAD18-mediated ubiquitination of PCNA activates the Fanconi anemia DNA repair network
    Article Snippet: .. GST pull-down assays GST pull-down experiments were performed as described previously ( ) using proteins obtained as follows: GST, GST-PCNA, His6 -PCNA, and GST-FANCL (amino acids 1–306) were purified from E. coli using glutathione agarose (Thermo Fisher Scientific) or TALON Superflow metal affinity resin (Takara Bio Inc.) chromatography, S-tagged FANCL was produced with a coupled in vitro transcription/translation system (Promega), and SFB-FANCL was produced by transient expression in K562 cells. .. In vitro ubiquitination assays To prepare in vitro–ubiquitylated PCNA, 100 ng purified GST-PCNA or GST-PCNAK164R was incubated with 50 ng rabbit E1 ubiquitin–activating enzyme (EMD) and 350 ng hRAD18-RAD6 (prepared as described previously in ) with or without 5 µg HA-tagged human ubiquitin (Boston Biochem) in ubiquitination reaction buffer (50 mM Tris, pH 7.4, 5 mM MgCl2 , 2 mM ATP, 2 µM ZnCl2 , 2 mM NaF, and 0.6 mM DTT) at 30°C for 1 h. The GST-tagged PCNA preparations were incubated with glutathione agarose at 4°C for 1 h and washed with 50 mM Tris, pH 7.4, 500 mM NaCl, and 1% NP-40 four times and once with ubiquitination reaction buffer.

    Article Title: Structure of a helicase-helicase loader complex reveals insights into the mechanism of bacterial primosome assembly
    Article Snippet: Paragraph title: Protein expression and purification ... The supernatant of the lysed cell paste solution was filtered with a 0.22-μM filter unit (Millipore) and loaded onto a 10-ml Talon superflow metal affinity resin (Clontech) mounted in a 5.0 × 20 cm2 Econo column (Bio-rad).

    Article Title: Creation of a type IIS restriction endonuclease with a long recognition sequence
    Article Snippet: Paragraph title: Screen for high expression level of I-SceI mutant proteins in E. coli ... Thirty microliters of 50% slurry of TALON Superflow Metal Affinity Resin (Clontech, Mountain View, CA), pre-equilibrated with 50 mM sodium phosphate, pH 7.0, 0.5 M NaCl and 50 mM MgCl2 , were added to each, and the mixtures were rocked for an hour at 4°C.

    Article Title: ADP-dependent Conformational Changes Distinguish Mycobacterium tuberculosis SecA2 from SecA1 *
    Article Snippet: For SecA2 expression, cells were grown similarly, except at mid-log phase the temperature was lowered to 16 °C, induced with isopropyl β- d -thiogalactopyranoside, grown at 16 °C overnight, and harvested. .. The lysates were centrifuged at 26,000 × g for 15 min followed by ultracentrifugation at 162,000 × g for 3 h. The clarified lysate was applied to a column with Talon Superflow metal affinity resin (Clontech).

    Article Title: Modular Organization of ?-Toxins from Scorpion Venom Mirrors Domain Structure of Their Targets, Sodium Channels *
    Article Snippet: Gene expression was induced by 0.2 m m isopropyl β- d -1-thiogalactopyranoside. .. Trx-BeM9 fusion protein was isolated by affinity chromatography on TALON Superflow Metal Affinity Resin (Clontech, Mountain View, CA) and then hydrolyzed using CNBr as suggested ( ).

    Article Title: Structural Characterization of Proline-rich Tyrosine Kinase 2 (PYK2) Reveals a Unique (DFG-out) Conformation and Enables Inhibitor Design
    Article Snippet: Standard expression conditions using the Bac-to-Bac® Baculovirus Expression System from Invitrogen and Sf9 cells were employed for large-scale expression in shake flask and WAVE BioReactors. .. The supernatant was extracted and dispensed into a tube containing 20 ml of TALON Superflow Metal Affinity Resin (BD-Clonetech), pre-equilibrated with buffer B (100 m m KH2 PO4 /K2 HPO4 ; pH 8.0, 200 m m NaCl, 1 m m TCEP, 2 m m phenylmethylsulfonyl fluoride, 5 m m imidazole; pH 7.5).

    Article Title: Gene Expression and Molecular Characterization of a Xylanase from Chicken Cecum Metagenome
    Article Snippet: TALON Superflow Metal Affinity Resin (Clonetech) was purchased from TaKaRa (Otsu, Japan). .. The expression vector pET-32a (Novagen) was used for cloning and expressing the xylanase.

    Article Title: The structural basis of CstF-77 modulation of cleavage and polyadenylation through stimulation of CstF-64 activity
    Article Snippet: Paragraph title: Bacterial protein expression and purification ... Lysates were clarified by centrifugation at 16 000 × g for 20 min at 4°C and loaded on home-packed columns either with an Amylose High Flow resin (New England Biolabs) or TALON Superflow Metal Affinity resin (TaKaRa) pre-equilibrated with the corresponding lysis buffers using peristaltic pump.

    Modification:

    Article Title: Structural Characterization of Proline-rich Tyrosine Kinase 2 (PYK2) Reveals a Unique (DFG-out) Conformation and Enables Inhibitor Design
    Article Snippet: The 854-bp DNA fragment was digested with BamHI and XhoI and cloned into the modified pFastBac vector at these sites to yield an N-terminal His6 -tagged thrombin cleavable fusion with PYK2 catalytic domain residues Pro-416 to Glu-692 based on accession . .. The supernatant was extracted and dispensed into a tube containing 20 ml of TALON Superflow Metal Affinity Resin (BD-Clonetech), pre-equilibrated with buffer B (100 m m KH2 PO4 /K2 HPO4 ; pH 8.0, 200 m m NaCl, 1 m m TCEP, 2 m m phenylmethylsulfonyl fluoride, 5 m m imidazole; pH 7.5).

    Transformation Assay:

    Article Title: Rapid antibody selection by mRNA display on a microfluidic chip
    Article Snippet: The transformed cells were cultured at 37°C in 400 ml of TB medium containing 100 µg/ml carbenicillin (Sigma) until the OD660 reached 0.5–0.6, then isopropylthio-ß-d -galactoside (Nacalai tesque) was added to a final concentration of 1 mM, and the cells were harvested 4–6 h later. .. The supernatants containing the histidine-tagged proteins in denatured form were immobilized on the TALON superflow metal affinity resin (Clontech), and the columns were washed with 10 volumes of TBS containing 10 mM imidazole and 6 M urea, 10 volumes of TBS containing 1 M NaCl and 10 volumes of TBS, to allow refolding of the bound proteins on the columns.

    Article Title: The structural basis of CstF-77 modulation of cleavage and polyadenylation through stimulation of CstF-64 activity
    Article Snippet: Bacterial protein expression and purification The constructs were transformed in Rosetta™ 2 (DE3) pLysS cell (Novagen). .. Lysates were clarified by centrifugation at 16 000 × g for 20 min at 4°C and loaded on home-packed columns either with an Amylose High Flow resin (New England Biolabs) or TALON Superflow Metal Affinity resin (TaKaRa) pre-equilibrated with the corresponding lysis buffers using peristaltic pump.

    High Performance Liquid Chromatography:

    Article Title: Modular Organization of ?-Toxins from Scorpion Venom Mirrors Domain Structure of Their Targets, Sodium Channels *
    Article Snippet: Trx-BeM9 fusion protein was isolated by affinity chromatography on TALON Superflow Metal Affinity Resin (Clontech, Mountain View, CA) and then hydrolyzed using CNBr as suggested ( ). .. Recombinant BeM9 was purified by reversed-phase HPLC on a Jupiter C5 column (250 × 4.6 mm; Phenomenex, Torrance, CA).

    Transfection:

    Article Title: Commitment and Differentiation of Osteoclast Precursor Cells by the Sequential Expression of C-Fms and Receptor Activator of Nuclear Factor ?b (Rank) Receptors
    Article Snippet: .. The PCR product was digested with HindIII and XhoI and ligated into the HindIII and XhoI sites of the pSecTag2 vector (Invitrogen Corp.) to yield pSecTag2–soluble (s)RANKL containing His6 and myc tags. pSecTag2–sRANKL was transfected into COS7 cells cultured in DMEM (Life Technologies, Inc.) containing 10% FCS, and the supernatant was collected every 4 d for 12 d. sRANKL was purified from the supernatant using TALON and TALON Superflow Metal Affinity Resin (Clontech). .. The supernatant was applied to a chromatography column (Bio-Rad Labs.) filled with affinity resin, and the trapped proteins were eluted in 125 nM imidazol and fractionated.

    Sequencing:

    Article Title: Modular Organization of ?-Toxins from Scorpion Venom Mirrors Domain Structure of Their Targets, Sodium Channels *
    Article Snippet: Because no methionine residues are found in BeM9, we introduced a methionine codon upstream of the toxin-coding sequence and used cyanogen bromide (CNBr) to liberate the toxin from the carrier protein thioredoxin (Trx; see below). .. Trx-BeM9 fusion protein was isolated by affinity chromatography on TALON Superflow Metal Affinity Resin (Clontech, Mountain View, CA) and then hydrolyzed using CNBr as suggested ( ).

    Chromatography:

    Article Title: RAD18-mediated ubiquitination of PCNA activates the Fanconi anemia DNA repair network
    Article Snippet: .. GST pull-down assays GST pull-down experiments were performed as described previously ( ) using proteins obtained as follows: GST, GST-PCNA, His6 -PCNA, and GST-FANCL (amino acids 1–306) were purified from E. coli using glutathione agarose (Thermo Fisher Scientific) or TALON Superflow metal affinity resin (Takara Bio Inc.) chromatography, S-tagged FANCL was produced with a coupled in vitro transcription/translation system (Promega), and SFB-FANCL was produced by transient expression in K562 cells. .. In vitro ubiquitination assays To prepare in vitro–ubiquitylated PCNA, 100 ng purified GST-PCNA or GST-PCNAK164R was incubated with 50 ng rabbit E1 ubiquitin–activating enzyme (EMD) and 350 ng hRAD18-RAD6 (prepared as described previously in ) with or without 5 µg HA-tagged human ubiquitin (Boston Biochem) in ubiquitination reaction buffer (50 mM Tris, pH 7.4, 5 mM MgCl2 , 2 mM ATP, 2 µM ZnCl2 , 2 mM NaF, and 0.6 mM DTT) at 30°C for 1 h. The GST-tagged PCNA preparations were incubated with glutathione agarose at 4°C for 1 h and washed with 50 mM Tris, pH 7.4, 500 mM NaCl, and 1% NP-40 four times and once with ubiquitination reaction buffer.

    Article Title: Human Common Salivary Protein 1 (CSP-1) Promotes Binding of Streptococcus mutans to Experimental Salivary Pellicle and Glucans Formed on Hydroxyapatite Surface
    Article Snippet: .. The secreted rCSP-1 protein was purified at 4 °C by metal chelate chromatography using TALON Superflow metal affinity resin (Clontech, Palo Alto, CA, USA) on a Biologic DuoFlow Maximizer purification system (Bio-Rad, Hercules, CA, USA) as described in Ambatipudi et. al. . ..

    Article Title: Commitment and Differentiation of Osteoclast Precursor Cells by the Sequential Expression of C-Fms and Receptor Activator of Nuclear Factor ?b (Rank) Receptors
    Article Snippet: The PCR product was digested with HindIII and XhoI and ligated into the HindIII and XhoI sites of the pSecTag2 vector (Invitrogen Corp.) to yield pSecTag2–soluble (s)RANKL containing His6 and myc tags. pSecTag2–sRANKL was transfected into COS7 cells cultured in DMEM (Life Technologies, Inc.) containing 10% FCS, and the supernatant was collected every 4 d for 12 d. sRANKL was purified from the supernatant using TALON and TALON Superflow Metal Affinity Resin (Clontech). .. The supernatant was applied to a chromatography column (Bio-Rad Labs.) filled with affinity resin, and the trapped proteins were eluted in 125 nM imidazol and fractionated.

    Protease Inhibitor:

    Article Title: Structure of a helicase-helicase loader complex reveals insights into the mechanism of bacterial primosome assembly
    Article Snippet: The collected cell paste was quickly frozen in liquid nitrogen, resuspended in buffer A (50 mM TRIS pH 8.0, 0.3 M sodium chloride, supplemented with EDTA-free protease inhibitor cocktail tablets (Roche)) and lysed using a microfluidizer. .. The supernatant of the lysed cell paste solution was filtered with a 0.22-μM filter unit (Millipore) and loaded onto a 10-ml Talon superflow metal affinity resin (Clontech) mounted in a 5.0 × 20 cm2 Econo column (Bio-rad).

    Article Title: Rapid antibody selection by mRNA display on a microfluidic chip
    Article Snippet: The precipitates were suspended in 20 ml of TBS containing 8 M urea and 8 U of DNase I, 40 µl of EDTA-free protease inhibitor cocktail and 1 mM 2-mercaptoethanol, and then recentrifuged for 20 min at 8500 r.p.m. .. The supernatants containing the histidine-tagged proteins in denatured form were immobilized on the TALON superflow metal affinity resin (Clontech), and the columns were washed with 10 volumes of TBS containing 10 mM imidazole and 6 M urea, 10 volumes of TBS containing 1 M NaCl and 10 volumes of TBS, to allow refolding of the bound proteins on the columns.

    Article Title: Creation of a type IIS restriction endonuclease with a long recognition sequence
    Article Snippet: The cell pellets were recovered by centrifugation (10 min at 3220g , 4°C), stored frozen at –20°C, then thawed on ice and lysed in 350 μl of 50 mM sodium phosphate, pH 7.0, 0.5 M NaCl, 50 mM MgCl2 , 0.5 mg/ml lysozyme, 0.05 mg/ml DNaseI, and 1× EDTA-free COMPLETE protease inhibitor cocktail (Roche, Indianapolis, IN). .. Thirty microliters of 50% slurry of TALON Superflow Metal Affinity Resin (Clontech, Mountain View, CA), pre-equilibrated with 50 mM sodium phosphate, pH 7.0, 0.5 M NaCl and 50 mM MgCl2 , were added to each, and the mixtures were rocked for an hour at 4°C.

    Article Title: ADP-dependent Conformational Changes Distinguish Mycobacterium tuberculosis SecA2 from SecA1 *
    Article Snippet: Cells were thawed on ice followed by the addition of 1× protease inhibitor mixture, 450 μg/ml each of DNase and RNase and 10 m m MgCl2 (final concentrations). .. The lysates were centrifuged at 26,000 × g for 15 min followed by ultracentrifugation at 162,000 × g for 3 h. The clarified lysate was applied to a column with Talon Superflow metal affinity resin (Clontech).

    Article Title: Structural Characterization of Proline-rich Tyrosine Kinase 2 (PYK2) Reveals a Unique (DFG-out) Conformation and Enables Inhibitor Design
    Article Snippet: Baculovirus cell paste containing the overexpressed His-PYK2 catalytic domain (416-692) recombinant protein was resuspended in 3 volumes of buffer A (100 m m KH2 PO4 /K2 HPO4 ; pH 8.0, 200 m m NaCl, 1 m m TCEP, Complete™ protease inhibitor (1 tablet/50 ml), Roche, 2 m m phenylmethylsulfonyl fluoride, 10% glycerol, 0.5 μl of Benzonase (Sigma) per 50 ml volume). .. The supernatant was extracted and dispensed into a tube containing 20 ml of TALON Superflow Metal Affinity Resin (BD-Clonetech), pre-equilibrated with buffer B (100 m m KH2 PO4 /K2 HPO4 ; pH 8.0, 200 m m NaCl, 1 m m TCEP, 2 m m phenylmethylsulfonyl fluoride, 5 m m imidazole; pH 7.5).

    Cell Culture:

    Article Title: Human APE2 protein is mostly localized in the nuclei and to some extent in the mitochondria, while nuclear APE2 is partly associated with proliferating cell nuclear antigen
    Article Snippet: BL21(DE3) E.coli cells carrying plasmid pET32a(+), pET3228D:PCNA or pET32a(+):APE2, which express His tagged proteins TrxA, His–PCNA and TrxA–APE2, respectively, were cultured in 250 ml of M9ZB medium ( ) until the OD600 reached 0.6 and then incubated with 1 mM isopropyl β- d -thiogalactoside for a further 2 h. Cells were harvested by centrifugation, resuspended in 25 ml of lysis buffer [50 mM sodium phosphate buffer pH 7.0, 300 mM NaCl, 1% Triton X-100, 8 M urea, 1 mM 2-mercaptoethanol (2-ME), 1 mM PMSF, 1 µg/ml leupeptin, 1 µg/ml pepstatin A and 1 µg/ml chymostatin] and disrupted by sonication. .. Two milliliters of a 50% suspension of TALON superflow metal affinity resin (Clontech Laboratories) equilibrated with lysis buffer was mixed with each cell lysate and gently rotated for 1 h. Then the resin was washed five times with binding buffer (20 mM sodium phosphate buffer pH 7.4, 0.1% BSA) containing 50 mM NaCl and used for pull-down binding assays.

    Article Title: Rapid antibody selection by mRNA display on a microfluidic chip
    Article Snippet: The transformed cells were cultured at 37°C in 400 ml of TB medium containing 100 µg/ml carbenicillin (Sigma) until the OD660 reached 0.5–0.6, then isopropylthio-ß-d -galactoside (Nacalai tesque) was added to a final concentration of 1 mM, and the cells were harvested 4–6 h later. .. The supernatants containing the histidine-tagged proteins in denatured form were immobilized on the TALON superflow metal affinity resin (Clontech), and the columns were washed with 10 volumes of TBS containing 10 mM imidazole and 6 M urea, 10 volumes of TBS containing 1 M NaCl and 10 volumes of TBS, to allow refolding of the bound proteins on the columns.

    Article Title: Commitment and Differentiation of Osteoclast Precursor Cells by the Sequential Expression of C-Fms and Receptor Activator of Nuclear Factor ?b (Rank) Receptors
    Article Snippet: .. The PCR product was digested with HindIII and XhoI and ligated into the HindIII and XhoI sites of the pSecTag2 vector (Invitrogen Corp.) to yield pSecTag2–soluble (s)RANKL containing His6 and myc tags. pSecTag2–sRANKL was transfected into COS7 cells cultured in DMEM (Life Technologies, Inc.) containing 10% FCS, and the supernatant was collected every 4 d for 12 d. sRANKL was purified from the supernatant using TALON and TALON Superflow Metal Affinity Resin (Clontech). .. The supernatant was applied to a chromatography column (Bio-Rad Labs.) filled with affinity resin, and the trapped proteins were eluted in 125 nM imidazol and fractionated.

    Polymerase Chain Reaction:

    Article Title: Structure of a helicase-helicase loader complex reveals insights into the mechanism of bacterial primosome assembly
    Article Snippet: The helicase loader protein DnaI fragment was cloned from B. subtilis 168 genomic DNA (American Type Culture Collection) using PCR with primers (Integrated DNA Technologies): forward 5′-gcgaccatggaaccaatcggccgttcc-3′ and reverse 5′-gcgactcgagtggatgtcggcggttttctc-3′, and inserted into the Nco I–Xho I site of plasmid pET21d(+) (Novagen) and expressed in B834(DE3) (Novagen). .. The supernatant of the lysed cell paste solution was filtered with a 0.22-μM filter unit (Millipore) and loaded onto a 10-ml Talon superflow metal affinity resin (Clontech) mounted in a 5.0 × 20 cm2 Econo column (Bio-rad).

    Article Title: Modular Organization of ?-Toxins from Scorpion Venom Mirrors Domain Structure of Their Targets, Sodium Channels *
    Article Snippet: DNA encoding BeM9 from the venom of the scorpion Mesobuthus eupeus ( ) (Uniprot ID ) was assembled from synthetic oligonucleotides ( ) by PCR. .. Trx-BeM9 fusion protein was isolated by affinity chromatography on TALON Superflow Metal Affinity Resin (Clontech, Mountain View, CA) and then hydrolyzed using CNBr as suggested ( ).

    Article Title: Commitment and Differentiation of Osteoclast Precursor Cells by the Sequential Expression of C-Fms and Receptor Activator of Nuclear Factor ?b (Rank) Receptors
    Article Snippet: .. The PCR product was digested with HindIII and XhoI and ligated into the HindIII and XhoI sites of the pSecTag2 vector (Invitrogen Corp.) to yield pSecTag2–soluble (s)RANKL containing His6 and myc tags. pSecTag2–sRANKL was transfected into COS7 cells cultured in DMEM (Life Technologies, Inc.) containing 10% FCS, and the supernatant was collected every 4 d for 12 d. sRANKL was purified from the supernatant using TALON and TALON Superflow Metal Affinity Resin (Clontech). .. The supernatant was applied to a chromatography column (Bio-Rad Labs.) filled with affinity resin, and the trapped proteins were eluted in 125 nM imidazol and fractionated.

    Sonication:

    Article Title: Human APE2 protein is mostly localized in the nuclei and to some extent in the mitochondria, while nuclear APE2 is partly associated with proliferating cell nuclear antigen
    Article Snippet: BL21(DE3) E.coli cells carrying plasmid pET32a(+), pET3228D:PCNA or pET32a(+):APE2, which express His tagged proteins TrxA, His–PCNA and TrxA–APE2, respectively, were cultured in 250 ml of M9ZB medium ( ) until the OD600 reached 0.6 and then incubated with 1 mM isopropyl β- d -thiogalactoside for a further 2 h. Cells were harvested by centrifugation, resuspended in 25 ml of lysis buffer [50 mM sodium phosphate buffer pH 7.0, 300 mM NaCl, 1% Triton X-100, 8 M urea, 1 mM 2-mercaptoethanol (2-ME), 1 mM PMSF, 1 µg/ml leupeptin, 1 µg/ml pepstatin A and 1 µg/ml chymostatin] and disrupted by sonication. .. Two milliliters of a 50% suspension of TALON superflow metal affinity resin (Clontech Laboratories) equilibrated with lysis buffer was mixed with each cell lysate and gently rotated for 1 h. Then the resin was washed five times with binding buffer (20 mM sodium phosphate buffer pH 7.4, 0.1% BSA) containing 50 mM NaCl and used for pull-down binding assays.

    Article Title: Rapid antibody selection by mRNA display on a microfluidic chip
    Article Snippet: The cells were lyzed by sonication using a Bioruptor UCW-201 (Cosmo Bio) twice for 15 min at 30-s intervals. .. The supernatants containing the histidine-tagged proteins in denatured form were immobilized on the TALON superflow metal affinity resin (Clontech), and the columns were washed with 10 volumes of TBS containing 10 mM imidazole and 6 M urea, 10 volumes of TBS containing 1 M NaCl and 10 volumes of TBS, to allow refolding of the bound proteins on the columns.

    Affinity Purification:

    Article Title: Interaction of intramembrane metalloprotease SpoIVFB with substrate Pro-σK
    Article Snippet: Paragraph title: Cobalt Affinity Purification (Pull-Down Assays). ... The rest was mixed with 0.5 mL of Talon superflow metal affinity resin (Clontech) that had been equilibrated with PBS containing 0.1% DM, 5 mM 2-mercaptoethanol, and 10% glycerol.

    Binding Assay:

    Article Title: Human APE2 protein is mostly localized in the nuclei and to some extent in the mitochondria, while nuclear APE2 is partly associated with proliferating cell nuclear antigen
    Article Snippet: .. Two milliliters of a 50% suspension of TALON superflow metal affinity resin (Clontech Laboratories) equilibrated with lysis buffer was mixed with each cell lysate and gently rotated for 1 h. Then the resin was washed five times with binding buffer (20 mM sodium phosphate buffer pH 7.4, 0.1% BSA) containing 50 mM NaCl and used for pull-down binding assays. .. Purity and amount of protein fixed on each resin were determined by SDS–PAGE with a combination of GelCode Blue Stain Reagent (Pierce Chemical Co.) and BSA as the standard.

    Article Title: The structural basis of CstF-77 modulation of cleavage and polyadenylation through stimulation of CstF-64 activity
    Article Snippet: One-liter cultures were induced with 0.5 mM isopropyl-β-d-thiogalactopyranoside (IPTG) at an optical density of 0.8 at 37°C for 4 h. Bacterial cells were suspended in either 1 M NaCl, 25 mM Tris–HCl pH 7.5, 1 mM EDTA, 0.05% sodium azide for the constructs containing the maltose binding protein (see above) or 1 M NaCl, 20 mM HEPES pH 7.5, 5 mM Imidazole, 0.05% sodium azide for the RRM. .. Lysates were clarified by centrifugation at 16 000 × g for 20 min at 4°C and loaded on home-packed columns either with an Amylose High Flow resin (New England Biolabs) or TALON Superflow Metal Affinity resin (TaKaRa) pre-equilibrated with the corresponding lysis buffers using peristaltic pump.

    DNA Extraction:

    Article Title: Gene Expression and Molecular Characterization of a Xylanase from Chicken Cecum Metagenome
    Article Snippet: Plasmid DNA extraction and purification kit was purchased from GE Healthcare, UK. .. TALON Superflow Metal Affinity Resin (Clonetech) was purchased from TaKaRa (Otsu, Japan).

    Mutagenesis:

    Article Title: Creation of a type IIS restriction endonuclease with a long recognition sequence
    Article Snippet: Paragraph title: Screen for high expression level of I-SceI mutant proteins in E. coli ... Thirty microliters of 50% slurry of TALON Superflow Metal Affinity Resin (Clontech, Mountain View, CA), pre-equilibrated with 50 mM sodium phosphate, pH 7.0, 0.5 M NaCl and 50 mM MgCl2 , were added to each, and the mixtures were rocked for an hour at 4°C.

    Isolation:

    Article Title: Modular Organization of ?-Toxins from Scorpion Venom Mirrors Domain Structure of Their Targets, Sodium Channels *
    Article Snippet: .. Trx-BeM9 fusion protein was isolated by affinity chromatography on TALON Superflow Metal Affinity Resin (Clontech, Mountain View, CA) and then hydrolyzed using CNBr as suggested ( ). .. Recombinant BeM9 was purified by reversed-phase HPLC on a Jupiter C5 column (250 × 4.6 mm; Phenomenex, Torrance, CA).

    Flow Cytometry:

    Article Title: Rapid antibody selection by mRNA display on a microfluidic chip
    Article Snippet: The supernatants containing the histidine-tagged proteins in denatured form were immobilized on the TALON superflow metal affinity resin (Clontech), and the columns were washed with 10 volumes of TBS containing 10 mM imidazole and 6 M urea, 10 volumes of TBS containing 1 M NaCl and 10 volumes of TBS, to allow refolding of the bound proteins on the columns. .. Subsequently, the proteins were separated by size exclusion chromatography using Sephadex G-75 10/300 GL (Amersham Biosciences) on an AKTA explorer 10S (Amersham Biosciences) equilibrated with HBS-EP buffer (10 mM HEPES–NaOH, pH 7.4, 150 mM NaCl, 3 mM EDTA and 0.005% Tween-20) at a flow rate of 0.5 ml/min.

    Article Title: The structural basis of CstF-77 modulation of cleavage and polyadenylation through stimulation of CstF-64 activity
    Article Snippet: .. Lysates were clarified by centrifugation at 16 000 × g for 20 min at 4°C and loaded on home-packed columns either with an Amylose High Flow resin (New England Biolabs) or TALON Superflow Metal Affinity resin (TaKaRa) pre-equilibrated with the corresponding lysis buffers using peristaltic pump. .. Bound protein was washed with about 10 column volumes in the corresponding lysis buffer followed by a wash with about five column volumes in buffer containing 0.3 M NaCl, 20 mM HEPES pH 7.5, 5 mM imidazole, 0.05% sodium azide.

    Size-exclusion Chromatography:

    Article Title: Rapid antibody selection by mRNA display on a microfluidic chip
    Article Snippet: The supernatants containing the histidine-tagged proteins in denatured form were immobilized on the TALON superflow metal affinity resin (Clontech), and the columns were washed with 10 volumes of TBS containing 10 mM imidazole and 6 M urea, 10 volumes of TBS containing 1 M NaCl and 10 volumes of TBS, to allow refolding of the bound proteins on the columns. .. Subsequently, the proteins were separated by size exclusion chromatography using Sephadex G-75 10/300 GL (Amersham Biosciences) on an AKTA explorer 10S (Amersham Biosciences) equilibrated with HBS-EP buffer (10 mM HEPES–NaOH, pH 7.4, 150 mM NaCl, 3 mM EDTA and 0.005% Tween-20) at a flow rate of 0.5 ml/min.

    Protein Purification:

    Article Title: Bacillus subtilis Intramembrane Protease RasP Activity in Escherichia coli and In Vitro
    Article Snippet: Paragraph title: Protein purification. ... The supernatant was mixed with 1 ml Talon Superflow metal affinity resin (Clontech) that had been equilibrated with buffer (PBS containing 1% DM, 5 mM 2-mercaptoethanol, and 10% glycerol).

    Article Title: ADP-dependent Conformational Changes Distinguish Mycobacterium tuberculosis SecA2 from SecA1 *
    Article Snippet: Paragraph title: Protein Purification ... The lysates were centrifuged at 26,000 × g for 15 min followed by ultracentrifugation at 162,000 × g for 3 h. The clarified lysate was applied to a column with Talon Superflow metal affinity resin (Clontech).

    Reverse Transcription Polymerase Chain Reaction:

    Article Title: Commitment and Differentiation of Osteoclast Precursor Cells by the Sequential Expression of C-Fms and Receptor Activator of Nuclear Factor ?b (Rank) Receptors
    Article Snippet: A DNA fragment encoding the extracellular domain (Asp76 –Asp316 ) of RANKL was prepared using reverse transcriptase (RT)-PCR from total RNA of ST2 stromal cells that were cultured in the presence of 1,25-(OH)2 D3 (10−8 M) for 4 d. PCR primers were as follows: 5′-CCGCTCGAGCGTCTATGTCCTGAACTTTGA-3′ (sense) and 5′-CCCAAGCTTGATCCTAACAGAATATCAGAAGACA-3′ (antisense). .. The PCR product was digested with HindIII and XhoI and ligated into the HindIII and XhoI sites of the pSecTag2 vector (Invitrogen Corp.) to yield pSecTag2–soluble (s)RANKL containing His6 and myc tags. pSecTag2–sRANKL was transfected into COS7 cells cultured in DMEM (Life Technologies, Inc.) containing 10% FCS, and the supernatant was collected every 4 d for 12 d. sRANKL was purified from the supernatant using TALON and TALON Superflow Metal Affinity Resin (Clontech).

    Positron Emission Tomography:

    Article Title: Modular Organization of ?-Toxins from Scorpion Venom Mirrors Domain Structure of Their Targets, Sodium Channels *
    Article Snippet: The toxin DNA was cloned into the expression vector pET-32b (Novagen (Madison, WI)), which was then used to transform Escherichia coli BL21(DE3). .. Trx-BeM9 fusion protein was isolated by affinity chromatography on TALON Superflow Metal Affinity Resin (Clontech, Mountain View, CA) and then hydrolyzed using CNBr as suggested ( ).

    Article Title: Gene Expression and Molecular Characterization of a Xylanase from Chicken Cecum Metagenome
    Article Snippet: TALON Superflow Metal Affinity Resin (Clonetech) was purchased from TaKaRa (Otsu, Japan). .. The expression vector pET-32a (Novagen) was used for cloning and expressing the xylanase.

    Fast Protein Liquid Chromatography:

    Article Title: Structural Characterization of Proline-rich Tyrosine Kinase 2 (PYK2) Reveals a Unique (DFG-out) Conformation and Enables Inhibitor Design
    Article Snippet: The supernatant was extracted and dispensed into a tube containing 20 ml of TALON Superflow Metal Affinity Resin (BD-Clonetech), pre-equilibrated with buffer B (100 m m KH2 PO4 /K2 HPO4 ; pH 8.0, 200 m m NaCl, 1 m m TCEP, 2 m m phenylmethylsulfonyl fluoride, 5 m m imidazole; pH 7.5). .. The resin was then transferred to an XK-26 column (Amersham Biosciences) connected to a FPLC™ and washed with buffer B until the OD reached 0.05 units.

    Recombinant:

    Article Title: Human Common Salivary Protein 1 (CSP-1) Promotes Binding of Streptococcus mutans to Experimental Salivary Pellicle and Glucans Formed on Hydroxyapatite Surface
    Article Snippet: Paragraph title: Purification of Recombinant Common Salivary Protein-1 (Rcsp-1) by Metal Affinity Followed by Con A Lectin Affinity Chromatography ... The secreted rCSP-1 protein was purified at 4 °C by metal chelate chromatography using TALON Superflow metal affinity resin (Clontech, Palo Alto, CA, USA) on a Biologic DuoFlow Maximizer purification system (Bio-Rad, Hercules, CA, USA) as described in Ambatipudi et. al. .

    Article Title: Modular Organization of ?-Toxins from Scorpion Venom Mirrors Domain Structure of Their Targets, Sodium Channels *
    Article Snippet: Paragraph title: Production of Recombinant Toxin BeM9 ... Trx-BeM9 fusion protein was isolated by affinity chromatography on TALON Superflow Metal Affinity Resin (Clontech, Mountain View, CA) and then hydrolyzed using CNBr as suggested ( ).

    Article Title: Structural Characterization of Proline-rich Tyrosine Kinase 2 (PYK2) Reveals a Unique (DFG-out) Conformation and Enables Inhibitor Design
    Article Snippet: Baculovirus cell paste containing the overexpressed His-PYK2 catalytic domain (416-692) recombinant protein was resuspended in 3 volumes of buffer A (100 m m KH2 PO4 /K2 HPO4 ; pH 8.0, 200 m m NaCl, 1 m m TCEP, Complete™ protease inhibitor (1 tablet/50 ml), Roche, 2 m m phenylmethylsulfonyl fluoride, 10% glycerol, 0.5 μl of Benzonase (Sigma) per 50 ml volume). .. The supernatant was extracted and dispensed into a tube containing 20 ml of TALON Superflow Metal Affinity Resin (BD-Clonetech), pre-equilibrated with buffer B (100 m m KH2 PO4 /K2 HPO4 ; pH 8.0, 200 m m NaCl, 1 m m TCEP, 2 m m phenylmethylsulfonyl fluoride, 5 m m imidazole; pH 7.5).

    Lysis:

    Article Title: Human APE2 protein is mostly localized in the nuclei and to some extent in the mitochondria, while nuclear APE2 is partly associated with proliferating cell nuclear antigen
    Article Snippet: .. Two milliliters of a 50% suspension of TALON superflow metal affinity resin (Clontech Laboratories) equilibrated with lysis buffer was mixed with each cell lysate and gently rotated for 1 h. Then the resin was washed five times with binding buffer (20 mM sodium phosphate buffer pH 7.4, 0.1% BSA) containing 50 mM NaCl and used for pull-down binding assays. .. Purity and amount of protein fixed on each resin were determined by SDS–PAGE with a combination of GelCode Blue Stain Reagent (Pierce Chemical Co.) and BSA as the standard.

    Article Title: Interaction of intramembrane metalloprotease SpoIVFB with substrate Pro-σK
    Article Snippet: Cells were harvested, lysed, and centrifuged at low speed as described ( ) except 20 mL of lysis buffer was used. .. The rest was mixed with 0.5 mL of Talon superflow metal affinity resin (Clontech) that had been equilibrated with PBS containing 0.1% DM, 5 mM 2-mercaptoethanol, and 10% glycerol.

    Article Title: The structural basis of CstF-77 modulation of cleavage and polyadenylation through stimulation of CstF-64 activity
    Article Snippet: .. Lysates were clarified by centrifugation at 16 000 × g for 20 min at 4°C and loaded on home-packed columns either with an Amylose High Flow resin (New England Biolabs) or TALON Superflow Metal Affinity resin (TaKaRa) pre-equilibrated with the corresponding lysis buffers using peristaltic pump. .. Bound protein was washed with about 10 column volumes in the corresponding lysis buffer followed by a wash with about five column volumes in buffer containing 0.3 M NaCl, 20 mM HEPES pH 7.5, 5 mM imidazole, 0.05% sodium azide.

    Purification:

    Article Title: RAD18-mediated ubiquitination of PCNA activates the Fanconi anemia DNA repair network
    Article Snippet: .. GST pull-down assays GST pull-down experiments were performed as described previously ( ) using proteins obtained as follows: GST, GST-PCNA, His6 -PCNA, and GST-FANCL (amino acids 1–306) were purified from E. coli using glutathione agarose (Thermo Fisher Scientific) or TALON Superflow metal affinity resin (Takara Bio Inc.) chromatography, S-tagged FANCL was produced with a coupled in vitro transcription/translation system (Promega), and SFB-FANCL was produced by transient expression in K562 cells. .. In vitro ubiquitination assays To prepare in vitro–ubiquitylated PCNA, 100 ng purified GST-PCNA or GST-PCNAK164R was incubated with 50 ng rabbit E1 ubiquitin–activating enzyme (EMD) and 350 ng hRAD18-RAD6 (prepared as described previously in ) with or without 5 µg HA-tagged human ubiquitin (Boston Biochem) in ubiquitination reaction buffer (50 mM Tris, pH 7.4, 5 mM MgCl2 , 2 mM ATP, 2 µM ZnCl2 , 2 mM NaF, and 0.6 mM DTT) at 30°C for 1 h. The GST-tagged PCNA preparations were incubated with glutathione agarose at 4°C for 1 h and washed with 50 mM Tris, pH 7.4, 500 mM NaCl, and 1% NP-40 four times and once with ubiquitination reaction buffer.

    Article Title: Structure of a helicase-helicase loader complex reveals insights into the mechanism of bacterial primosome assembly
    Article Snippet: Paragraph title: Protein expression and purification ... The supernatant of the lysed cell paste solution was filtered with a 0.22-μM filter unit (Millipore) and loaded onto a 10-ml Talon superflow metal affinity resin (Clontech) mounted in a 5.0 × 20 cm2 Econo column (Bio-rad).

    Article Title: Rapid antibody selection by mRNA display on a microfluidic chip
    Article Snippet: For purification of proteins, the cells were collected by centrifugation, and resuspended in 20 ml of TBS (20 mM Tris–HCl buffer, pH 7.5, 138 mM NaCl) containing 8 U of DNase I (Invitrogen), 40 µl of EDTA-free protease inhibitor cocktail (Nacalai tesque) and 1 mM 2-mercaptoethanol (Nacalai tesque). .. The supernatants containing the histidine-tagged proteins in denatured form were immobilized on the TALON superflow metal affinity resin (Clontech), and the columns were washed with 10 volumes of TBS containing 10 mM imidazole and 6 M urea, 10 volumes of TBS containing 1 M NaCl and 10 volumes of TBS, to allow refolding of the bound proteins on the columns.

    Article Title: Human Common Salivary Protein 1 (CSP-1) Promotes Binding of Streptococcus mutans to Experimental Salivary Pellicle and Glucans Formed on Hydroxyapatite Surface
    Article Snippet: .. The secreted rCSP-1 protein was purified at 4 °C by metal chelate chromatography using TALON Superflow metal affinity resin (Clontech, Palo Alto, CA, USA) on a Biologic DuoFlow Maximizer purification system (Bio-Rad, Hercules, CA, USA) as described in Ambatipudi et. al. . ..

    Article Title: Modular Organization of ?-Toxins from Scorpion Venom Mirrors Domain Structure of Their Targets, Sodium Channels *
    Article Snippet: Trx-BeM9 fusion protein was isolated by affinity chromatography on TALON Superflow Metal Affinity Resin (Clontech, Mountain View, CA) and then hydrolyzed using CNBr as suggested ( ). .. Recombinant BeM9 was purified by reversed-phase HPLC on a Jupiter C5 column (250 × 4.6 mm; Phenomenex, Torrance, CA).

    Article Title: Gene Expression and Molecular Characterization of a Xylanase from Chicken Cecum Metagenome
    Article Snippet: Plasmid DNA extraction and purification kit was purchased from GE Healthcare, UK. .. TALON Superflow Metal Affinity Resin (Clonetech) was purchased from TaKaRa (Otsu, Japan).

    Article Title: Commitment and Differentiation of Osteoclast Precursor Cells by the Sequential Expression of C-Fms and Receptor Activator of Nuclear Factor ?b (Rank) Receptors
    Article Snippet: .. The PCR product was digested with HindIII and XhoI and ligated into the HindIII and XhoI sites of the pSecTag2 vector (Invitrogen Corp.) to yield pSecTag2–soluble (s)RANKL containing His6 and myc tags. pSecTag2–sRANKL was transfected into COS7 cells cultured in DMEM (Life Technologies, Inc.) containing 10% FCS, and the supernatant was collected every 4 d for 12 d. sRANKL was purified from the supernatant using TALON and TALON Superflow Metal Affinity Resin (Clontech). .. The supernatant was applied to a chromatography column (Bio-Rad Labs.) filled with affinity resin, and the trapped proteins were eluted in 125 nM imidazol and fractionated.

    Article Title: The structural basis of CstF-77 modulation of cleavage and polyadenylation through stimulation of CstF-64 activity
    Article Snippet: Paragraph title: Bacterial protein expression and purification ... Lysates were clarified by centrifugation at 16 000 × g for 20 min at 4°C and loaded on home-packed columns either with an Amylose High Flow resin (New England Biolabs) or TALON Superflow Metal Affinity resin (TaKaRa) pre-equilibrated with the corresponding lysis buffers using peristaltic pump.

    SDS Page:

    Article Title: Human APE2 protein is mostly localized in the nuclei and to some extent in the mitochondria, while nuclear APE2 is partly associated with proliferating cell nuclear antigen
    Article Snippet: Two milliliters of a 50% suspension of TALON superflow metal affinity resin (Clontech Laboratories) equilibrated with lysis buffer was mixed with each cell lysate and gently rotated for 1 h. Then the resin was washed five times with binding buffer (20 mM sodium phosphate buffer pH 7.4, 0.1% BSA) containing 50 mM NaCl and used for pull-down binding assays. .. Purity and amount of protein fixed on each resin were determined by SDS–PAGE with a combination of GelCode Blue Stain Reagent (Pierce Chemical Co.) and BSA as the standard.

    Article Title: Rapid antibody selection by mRNA display on a microfluidic chip
    Article Snippet: The supernatants containing the histidine-tagged proteins in denatured form were immobilized on the TALON superflow metal affinity resin (Clontech), and the columns were washed with 10 volumes of TBS containing 10 mM imidazole and 6 M urea, 10 volumes of TBS containing 1 M NaCl and 10 volumes of TBS, to allow refolding of the bound proteins on the columns. .. The purified proteins were analyzed by SDS–PAGE followed by staining with SimplyBlue™ (Invitrogen).

    Article Title: Creation of a type IIS restriction endonuclease with a long recognition sequence
    Article Snippet: Thirty microliters of 50% slurry of TALON Superflow Metal Affinity Resin (Clontech, Mountain View, CA), pre-equilibrated with 50 mM sodium phosphate, pH 7.0, 0.5 M NaCl and 50 mM MgCl2 , were added to each, and the mixtures were rocked for an hour at 4°C. .. The resuspended resin was combined with reducing SDS–PAGE buffer, boiled at 99°C for 5 min, separated on 4–12% Bis/Tris gradient gels in MES-based buffer (Invitrogen), and detected with GelCode Blue (Pierce, Rockford, IL) (Figure 1S).

    Article Title: ADP-dependent Conformational Changes Distinguish Mycobacterium tuberculosis SecA2 from SecA1 *
    Article Snippet: The lysates were centrifuged at 26,000 × g for 15 min followed by ultracentrifugation at 162,000 × g for 3 h. The clarified lysate was applied to a column with Talon Superflow metal affinity resin (Clontech). .. The tagged proteins were eluted with 250 m m imidazole, pH 8.0, 20 m m Na2 HPO4 , 300 m m NaCl, and fractions were analyzed by SDS-PAGE.

    Plasmid Preparation:

    Article Title: Structure of a helicase-helicase loader complex reveals insights into the mechanism of bacterial primosome assembly
    Article Snippet: The helicase loader protein DnaI fragment was cloned from B. subtilis 168 genomic DNA (American Type Culture Collection) using PCR with primers (Integrated DNA Technologies): forward 5′-gcgaccatggaaccaatcggccgttcc-3′ and reverse 5′-gcgactcgagtggatgtcggcggttttctc-3′, and inserted into the Nco I–Xho I site of plasmid pET21d(+) (Novagen) and expressed in B834(DE3) (Novagen). .. The supernatant of the lysed cell paste solution was filtered with a 0.22-μM filter unit (Millipore) and loaded onto a 10-ml Talon superflow metal affinity resin (Clontech) mounted in a 5.0 × 20 cm2 Econo column (Bio-rad).

    Article Title: Human APE2 protein is mostly localized in the nuclei and to some extent in the mitochondria, while nuclear APE2 is partly associated with proliferating cell nuclear antigen
    Article Snippet: BL21(DE3) E.coli cells carrying plasmid pET32a(+), pET3228D:PCNA or pET32a(+):APE2, which express His tagged proteins TrxA, His–PCNA and TrxA–APE2, respectively, were cultured in 250 ml of M9ZB medium ( ) until the OD600 reached 0.6 and then incubated with 1 mM isopropyl β- d -thiogalactoside for a further 2 h. Cells were harvested by centrifugation, resuspended in 25 ml of lysis buffer [50 mM sodium phosphate buffer pH 7.0, 300 mM NaCl, 1% Triton X-100, 8 M urea, 1 mM 2-mercaptoethanol (2-ME), 1 mM PMSF, 1 µg/ml leupeptin, 1 µg/ml pepstatin A and 1 µg/ml chymostatin] and disrupted by sonication. .. Two milliliters of a 50% suspension of TALON superflow metal affinity resin (Clontech Laboratories) equilibrated with lysis buffer was mixed with each cell lysate and gently rotated for 1 h. Then the resin was washed five times with binding buffer (20 mM sodium phosphate buffer pH 7.4, 0.1% BSA) containing 50 mM NaCl and used for pull-down binding assays.

    Article Title: Modular Organization of ?-Toxins from Scorpion Venom Mirrors Domain Structure of Their Targets, Sodium Channels *
    Article Snippet: The toxin DNA was cloned into the expression vector pET-32b (Novagen (Madison, WI)), which was then used to transform Escherichia coli BL21(DE3). .. Trx-BeM9 fusion protein was isolated by affinity chromatography on TALON Superflow Metal Affinity Resin (Clontech, Mountain View, CA) and then hydrolyzed using CNBr as suggested ( ).

    Article Title: Structural Characterization of Proline-rich Tyrosine Kinase 2 (PYK2) Reveals a Unique (DFG-out) Conformation and Enables Inhibitor Design
    Article Snippet: The 854-bp DNA fragment was digested with BamHI and XhoI and cloned into the modified pFastBac vector at these sites to yield an N-terminal His6 -tagged thrombin cleavable fusion with PYK2 catalytic domain residues Pro-416 to Glu-692 based on accession . .. The supernatant was extracted and dispensed into a tube containing 20 ml of TALON Superflow Metal Affinity Resin (BD-Clonetech), pre-equilibrated with buffer B (100 m m KH2 PO4 /K2 HPO4 ; pH 8.0, 200 m m NaCl, 1 m m TCEP, 2 m m phenylmethylsulfonyl fluoride, 5 m m imidazole; pH 7.5).

    Article Title: Gene Expression and Molecular Characterization of a Xylanase from Chicken Cecum Metagenome
    Article Snippet: Plasmid DNA extraction and purification kit was purchased from GE Healthcare, UK. .. TALON Superflow Metal Affinity Resin (Clonetech) was purchased from TaKaRa (Otsu, Japan).

    Article Title: Commitment and Differentiation of Osteoclast Precursor Cells by the Sequential Expression of C-Fms and Receptor Activator of Nuclear Factor ?b (Rank) Receptors
    Article Snippet: .. The PCR product was digested with HindIII and XhoI and ligated into the HindIII and XhoI sites of the pSecTag2 vector (Invitrogen Corp.) to yield pSecTag2–soluble (s)RANKL containing His6 and myc tags. pSecTag2–sRANKL was transfected into COS7 cells cultured in DMEM (Life Technologies, Inc.) containing 10% FCS, and the supernatant was collected every 4 d for 12 d. sRANKL was purified from the supernatant using TALON and TALON Superflow Metal Affinity Resin (Clontech). .. The supernatant was applied to a chromatography column (Bio-Rad Labs.) filled with affinity resin, and the trapped proteins were eluted in 125 nM imidazol and fractionated.

    Affinity Chromatography:

    Article Title: Human Common Salivary Protein 1 (CSP-1) Promotes Binding of Streptococcus mutans to Experimental Salivary Pellicle and Glucans Formed on Hydroxyapatite Surface
    Article Snippet: Paragraph title: Purification of Recombinant Common Salivary Protein-1 (Rcsp-1) by Metal Affinity Followed by Con A Lectin Affinity Chromatography ... The secreted rCSP-1 protein was purified at 4 °C by metal chelate chromatography using TALON Superflow metal affinity resin (Clontech, Palo Alto, CA, USA) on a Biologic DuoFlow Maximizer purification system (Bio-Rad, Hercules, CA, USA) as described in Ambatipudi et. al. .

    Article Title: Modular Organization of ?-Toxins from Scorpion Venom Mirrors Domain Structure of Their Targets, Sodium Channels *
    Article Snippet: .. Trx-BeM9 fusion protein was isolated by affinity chromatography on TALON Superflow Metal Affinity Resin (Clontech, Mountain View, CA) and then hydrolyzed using CNBr as suggested ( ). .. Recombinant BeM9 was purified by reversed-phase HPLC on a Jupiter C5 column (250 × 4.6 mm; Phenomenex, Torrance, CA).

    Article Title: Generation of a More Immunogenic Measles Vaccine by Increasing Its Hemagglutinin Expression
    Article Snippet: Paragraph title: Immobilized cobalt ion affinity chromatography. ... Talon Superflow metal affinity resin (Clontech Laboratories, Mountain View, CA) equilibrated in interaction buffer (50 mM NaH2 PO4 [pH 8], 200 mM NaCl, 1% Triton X-100) was added to the clarified supernatants for 2 h of batch interaction.

    In Vitro:

    Article Title: RAD18-mediated ubiquitination of PCNA activates the Fanconi anemia DNA repair network
    Article Snippet: .. GST pull-down assays GST pull-down experiments were performed as described previously ( ) using proteins obtained as follows: GST, GST-PCNA, His6 -PCNA, and GST-FANCL (amino acids 1–306) were purified from E. coli using glutathione agarose (Thermo Fisher Scientific) or TALON Superflow metal affinity resin (Takara Bio Inc.) chromatography, S-tagged FANCL was produced with a coupled in vitro transcription/translation system (Promega), and SFB-FANCL was produced by transient expression in K562 cells. .. In vitro ubiquitination assays To prepare in vitro–ubiquitylated PCNA, 100 ng purified GST-PCNA or GST-PCNAK164R was incubated with 50 ng rabbit E1 ubiquitin–activating enzyme (EMD) and 350 ng hRAD18-RAD6 (prepared as described previously in ) with or without 5 µg HA-tagged human ubiquitin (Boston Biochem) in ubiquitination reaction buffer (50 mM Tris, pH 7.4, 5 mM MgCl2 , 2 mM ATP, 2 µM ZnCl2 , 2 mM NaF, and 0.6 mM DTT) at 30°C for 1 h. The GST-tagged PCNA preparations were incubated with glutathione agarose at 4°C for 1 h and washed with 50 mM Tris, pH 7.4, 500 mM NaCl, and 1% NP-40 four times and once with ubiquitination reaction buffer.

    Produced:

    Article Title: RAD18-mediated ubiquitination of PCNA activates the Fanconi anemia DNA repair network
    Article Snippet: .. GST pull-down assays GST pull-down experiments were performed as described previously ( ) using proteins obtained as follows: GST, GST-PCNA, His6 -PCNA, and GST-FANCL (amino acids 1–306) were purified from E. coli using glutathione agarose (Thermo Fisher Scientific) or TALON Superflow metal affinity resin (Takara Bio Inc.) chromatography, S-tagged FANCL was produced with a coupled in vitro transcription/translation system (Promega), and SFB-FANCL was produced by transient expression in K562 cells. .. In vitro ubiquitination assays To prepare in vitro–ubiquitylated PCNA, 100 ng purified GST-PCNA or GST-PCNAK164R was incubated with 50 ng rabbit E1 ubiquitin–activating enzyme (EMD) and 350 ng hRAD18-RAD6 (prepared as described previously in ) with or without 5 µg HA-tagged human ubiquitin (Boston Biochem) in ubiquitination reaction buffer (50 mM Tris, pH 7.4, 5 mM MgCl2 , 2 mM ATP, 2 µM ZnCl2 , 2 mM NaF, and 0.6 mM DTT) at 30°C for 1 h. The GST-tagged PCNA preparations were incubated with glutathione agarose at 4°C for 1 h and washed with 50 mM Tris, pH 7.4, 500 mM NaCl, and 1% NP-40 four times and once with ubiquitination reaction buffer.

    Concentration Assay:

    Article Title: Rapid antibody selection by mRNA display on a microfluidic chip
    Article Snippet: The transformed cells were cultured at 37°C in 400 ml of TB medium containing 100 µg/ml carbenicillin (Sigma) until the OD660 reached 0.5–0.6, then isopropylthio-ß-d -galactoside (Nacalai tesque) was added to a final concentration of 1 mM, and the cells were harvested 4–6 h later. .. The supernatants containing the histidine-tagged proteins in denatured form were immobilized on the TALON superflow metal affinity resin (Clontech), and the columns were washed with 10 volumes of TBS containing 10 mM imidazole and 6 M urea, 10 volumes of TBS containing 1 M NaCl and 10 volumes of TBS, to allow refolding of the bound proteins on the columns.

    Article Title: Commitment and Differentiation of Osteoclast Precursor Cells by the Sequential Expression of C-Fms and Receptor Activator of Nuclear Factor ?b (Rank) Receptors
    Article Snippet: The PCR product was digested with HindIII and XhoI and ligated into the HindIII and XhoI sites of the pSecTag2 vector (Invitrogen Corp.) to yield pSecTag2–soluble (s)RANKL containing His6 and myc tags. pSecTag2–sRANKL was transfected into COS7 cells cultured in DMEM (Life Technologies, Inc.) containing 10% FCS, and the supernatant was collected every 4 d for 12 d. sRANKL was purified from the supernatant using TALON and TALON Superflow Metal Affinity Resin (Clontech). .. The high concentration fraction was collected and applied to a PD-10 column (Amersham Pharmacia Biotech, Inc.), eluted in PBS to eliminate the imidazol, and fractionated.

    BAC Assay:

    Article Title: Structural Characterization of Proline-rich Tyrosine Kinase 2 (PYK2) Reveals a Unique (DFG-out) Conformation and Enables Inhibitor Design
    Article Snippet: Standard expression conditions using the Bac-to-Bac® Baculovirus Expression System from Invitrogen and Sf9 cells were employed for large-scale expression in shake flask and WAVE BioReactors. .. The supernatant was extracted and dispensed into a tube containing 20 ml of TALON Superflow Metal Affinity Resin (BD-Clonetech), pre-equilibrated with buffer B (100 m m KH2 PO4 /K2 HPO4 ; pH 8.0, 200 m m NaCl, 1 m m TCEP, 2 m m phenylmethylsulfonyl fluoride, 5 m m imidazole; pH 7.5).

    Staining:

    Article Title: Human APE2 protein is mostly localized in the nuclei and to some extent in the mitochondria, while nuclear APE2 is partly associated with proliferating cell nuclear antigen
    Article Snippet: Two milliliters of a 50% suspension of TALON superflow metal affinity resin (Clontech Laboratories) equilibrated with lysis buffer was mixed with each cell lysate and gently rotated for 1 h. Then the resin was washed five times with binding buffer (20 mM sodium phosphate buffer pH 7.4, 0.1% BSA) containing 50 mM NaCl and used for pull-down binding assays. .. Purity and amount of protein fixed on each resin were determined by SDS–PAGE with a combination of GelCode Blue Stain Reagent (Pierce Chemical Co.) and BSA as the standard.

    Article Title: Rapid antibody selection by mRNA display on a microfluidic chip
    Article Snippet: The supernatants containing the histidine-tagged proteins in denatured form were immobilized on the TALON superflow metal affinity resin (Clontech), and the columns were washed with 10 volumes of TBS containing 10 mM imidazole and 6 M urea, 10 volumes of TBS containing 1 M NaCl and 10 volumes of TBS, to allow refolding of the bound proteins on the columns. .. The purified proteins were analyzed by SDS–PAGE followed by staining with SimplyBlue™ (Invitrogen).

    Article Title: Generation of a More Immunogenic Measles Vaccine by Increasing Its Hemagglutinin Expression
    Article Snippet: Talon Superflow metal affinity resin (Clontech Laboratories, Mountain View, CA) equilibrated in interaction buffer (50 mM NaH2 PO4 [pH 8], 200 mM NaCl, 1% Triton X-100) was added to the clarified supernatants for 2 h of batch interaction. .. The electrophoretically separated proteins were visualized by staining with Coomassie brilliant blue G-250.

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  • 90
    TaKaRa talon superflow metal affinity resin
    Talon Superflow Metal Affinity Resin, supplied by TaKaRa, used in various techniques. Bioz Stars score: 90/100, based on 30 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/talon superflow metal affinity resin/product/TaKaRa
    Average 90 stars, based on 30 article reviews
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    talon superflow metal affinity resin - by Bioz Stars, 2020-01
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