talon xtractor buffer  (TaKaRa)

 
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    Name:
    xTractor Buffer
    Description:
    xTractor Buffer is the most flexible and efficient cell lysis buffer for protein extraction from bacterial yeast mammalian and baculovirus infected cells This cell lysis reagent provides fast and efficient 10 min lysis using mild non denaturing conditions xTractor Buffer has been optimized for extraction of his tagged proteins so it is compatible with all IMAC resins
    Catalog Number:
    635671
    Price:
    None
    Size:
    250 mL
    Category:
    xTractor cell lysis buffers Purification products Protein research
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    Structured Review

    TaKaRa talon xtractor buffer
    xTractor Buffer is the most flexible and efficient cell lysis buffer for protein extraction from bacterial yeast mammalian and baculovirus infected cells This cell lysis reagent provides fast and efficient 10 min lysis using mild non denaturing conditions xTractor Buffer has been optimized for extraction of his tagged proteins so it is compatible with all IMAC resins
    https://www.bioz.com/result/talon xtractor buffer/product/TaKaRa
    Average 90 stars, based on 5 article reviews
    Price from $9.99 to $1999.99
    talon xtractor buffer - by Bioz Stars, 2020-01
    90/100 stars

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    Related Articles

    Clone Assay:

    Article Title: Electron Microscopy and 3D Reconstruction of F-actin Decorated with Cardiac Myosin-binding Protein C (cMyBP-C)
    Article Snippet: The Xho1 site was made so that the fragments were cloned in frame with a C-terminal 6-His tag. .. Cells were collected by centrifugation and lysed using the xTractor Buffer supplied in the HisTalon Buffer set (Clontech).

    Article Title: Polymyxin B Resistance and Biofilm Formation in Vibrio cholerae Are Controlled by the Response Regulator CarR
    Article Snippet: The coding region of carR , excluding the stop codon, was amplified by PCR and cloned into the pBAD/Myc-His C bacterial expression vector (Life Technologies, Carlsbad, CA), resulting in an overexpression plasmid encoding a recombinant CarR protein with a C-terminal Myc and 6×His tags (CarR-mycHis). .. Cells were harvested and then lysed with xTractor buffer (20 ml/g) (Clontech Laboratories, Mountain View, CA) for 10 min with orbital agitation at 4°C.

    Article Title: MAP1203 Promotes Mycobacterium avium Subspecies paratuberculosis Binding and Invasion to Bovine Epithelial Cells
    Article Snippet: Fragments were cloned into the HindIII/EcoI restriction sites of the pET6xHN-N-terminal expression vector. .. Bacteria were lysed using the xTractor Buffer (Clontech) as per manufacturer's instructions, separated on the SDS-PAGE protein gel, probed for 6xHN-tagged MAP1203 expression using a goat anti-6xHN primary antibody (1:2,000; Clontech Laboratories, Inc), and visualized with a rabbit anti-goat IR800 secondary antibody (1:10,000) (Licor).

    Article Title: Evolutionarily recent, insertional fission of mitochondrial cox2 into complementary genes in bilaterian Metazoa
    Article Snippet: Upon cleavage with Kpn I and Bam HI, the amplified products were cloned at the Kpn I-Bam HI site of the pProEx THb vector (ThermoFisher, Pittsburgh, PA). .. Bacteria were grown to the exponential phase, at which point the expression of recombinant proteins was induced with 1 mM IPTG at 30 °C for 6 h. Upon harvesting, the cells were disrupted using xTractor Buffer (Clontech).

    Centrifugation:

    Article Title: Citrullination mediated by PPAD constrains biofilm formation in P. gingivalis strain 381
    Article Snippet: For biofilm samples, 24 h biofilms were washed twice in dH2 O and then lysed with scraping in xTractor Buffer containing DNase I, lysozyme, and protease inhibitor (Takara Bio USA Inc.). .. Lysates were cleared by centrifugation at 12,000×g for 20 min at 4 °C and protein concentration of cleared lysates was determined using the PierceTM BCA Protein Assay Kit (Thermo Fisher Scientific).

    Article Title: Electron Microscopy and 3D Reconstruction of F-actin Decorated with Cardiac Myosin-binding Protein C (cMyBP-C)
    Article Snippet: .. Cells were collected by centrifugation and lysed using the xTractor Buffer supplied in the HisTalon Buffer set (Clontech). .. Protein was purified according to the HisTalon protocol, using HisPur Cobalt Resin (Thermo).

    Article Title: Studies of recombinant TWA1 reveal constitutive dimerization
    Article Snippet: At each timepoint, the bacteria were pelleted by centrifugation and stored at −20°C. .. To compare the extractability of TWA1.V5His6 using different methods of bacterial cell lysis, bacterial pellets corresponding to 0 h or 3 h timepoints were either lysed by gentle agitation in 500 μl of Xtractor buffer (Clontech) at room temperature for 1 h or resuspended in 1 ml of lysis buffer (200 mM NaCl, 50 mM TRIS, pH 8.0, 10% glycerol, 25 mM imidazole and 2 mM PMSF) and disrupted by sonication for 2 min (four cycles of 5 s bursts at 25 s intervals) on ice.

    Article Title: Specific Inter-residue Interactions as Determinants of Human Monoacylglycerol Lipase Catalytic Competency
    Article Snippet: After 5 h of induction at 30 °C, the cells were harvested by centrifugation at 5000 × g for 10 min, washed with phosphate-buffered saline, and held at −80 °C. .. For enzyme purification, 3 g (wet weight) of cells were resuspended in 20 ml of lysis buffer (20 m m sodium phosphate, 200 m m NaCl, 1 m m DTT, pH 7.4) and 20 ml of xTractor buffer (Clontech, Mountain View, CA) supplemented with lysozyme (0.1 mg/ml) and DNase I (25 μg/ml) (Fisher).

    Article Title: Citrullination mediated by PPAD constrains biofilm formation in P. gingivalis strain 381
    Article Snippet: Mass spectrometry analysis P. gingivalis stationary-phase cultures were centrifuged, supernatant was removed, and cells were lysed in xTractor Buffer containing DNase I, lysozyme, and protease inhibitor (Takara Bio USA Inc.). .. Lysates were cleared by centrifugation at 12,000×g for 20 min at 4 °C and protein concentration of cleared lysates was determined using the PierceTM BCA Protein Assay Kit (Thermo Fisher Scientific).

    Amplification:

    Article Title: Mating in the closest living relatives of animals is induced by a bacterial chondroitinase
    Article Snippet: The eroS gene was amplified by PCR (Phusion® DNA polymerase, New England Biosciences) from V. fischeri ES114 genomic DNA (Forward primer: 5’-GCCTCTGTCGACGCAAAAAATACCCAAACACCAC; Reverse primer: 5’-AATTAAGCGGCCGCCGTCTTGAATTGTTACTTGGAAAGAATAAG). .. After 24 hours, cells were pelleted and lysed with xTractor™ buffer (Clontech), purified with HisTALON™ gravity columns (Clontech), and the His-tag was released by enterokinase cleavage (Millipore enterokinase cleave capture kit #69067).

    Article Title: Polymyxin B Resistance and Biofilm Formation in Vibrio cholerae Are Controlled by the Response Regulator CarR
    Article Snippet: The coding region of carR , excluding the stop codon, was amplified by PCR and cloned into the pBAD/Myc-His C bacterial expression vector (Life Technologies, Carlsbad, CA), resulting in an overexpression plasmid encoding a recombinant CarR protein with a C-terminal Myc and 6×His tags (CarR-mycHis). .. Cells were harvested and then lysed with xTractor buffer (20 ml/g) (Clontech Laboratories, Mountain View, CA) for 10 min with orbital agitation at 4°C.

    Article Title: Chtop (Chromatin target of Prmt1) auto-regulates its expression level via intron retention and nonsense-mediated decay of its own mRNA
    Article Snippet: The amplified DNA was inserted at the Kpn I/Xho I site of pcDNA3.1 to produce HA-N1-pcDNA3.1. .. Cells were extracted with xTractor Buffer (Clontech), and TF-N1 was pulled down using TALON Magnetic Beads (Life Technologies).

    Article Title: Evolutionarily recent, insertional fission of mitochondrial cox2 into complementary genes in bilaterian Metazoa
    Article Snippet: Upon cleavage with Kpn I and Bam HI, the amplified products were cloned at the Kpn I-Bam HI site of the pProEx THb vector (ThermoFisher, Pittsburgh, PA). .. Bacteria were grown to the exponential phase, at which point the expression of recombinant proteins was induced with 1 mM IPTG at 30 °C for 6 h. Upon harvesting, the cells were disrupted using xTractor Buffer (Clontech).

    Expressing:

    Article Title: Mating in the closest living relatives of animals is induced by a bacterial chondroitinase
    Article Snippet: Paragraph title: Protein expression and purification ... After 24 hours, cells were pelleted and lysed with xTractor™ buffer (Clontech), purified with HisTALON™ gravity columns (Clontech), and the His-tag was released by enterokinase cleavage (Millipore enterokinase cleave capture kit #69067).

    Article Title: Electron Microscopy and 3D Reconstruction of F-actin Decorated with Cardiac Myosin-binding Protein C (cMyBP-C)
    Article Snippet: Protein expression was in BL21 (DE3) cells, grown for 4 hours after induction with IPTG. .. Cells were collected by centrifugation and lysed using the xTractor Buffer supplied in the HisTalon Buffer set (Clontech).

    Article Title: Polymyxin B Resistance and Biofilm Formation in Vibrio cholerae Are Controlled by the Response Regulator CarR
    Article Snippet: Briefly, cultures of BL21(DE3) harboring p carR were grown to mid-exponential phase in LB medium supplemented with ampicillin at 37°C, and expression of the recombinant protein was induced for 3 h at 30°C with the addition of 0.2% arabinose. .. Cells were harvested and then lysed with xTractor buffer (20 ml/g) (Clontech Laboratories, Mountain View, CA) for 10 min with orbital agitation at 4°C.

    Article Title: MAP1203 Promotes Mycobacterium avium Subspecies paratuberculosis Binding and Invasion to Bovine Epithelial Cells
    Article Snippet: .. Bacteria were lysed using the xTractor Buffer (Clontech) as per manufacturer's instructions, separated on the SDS-PAGE protein gel, probed for 6xHN-tagged MAP1203 expression using a goat anti-6xHN primary antibody (1:2,000; Clontech Laboratories, Inc), and visualized with a rabbit anti-goat IR800 secondary antibody (1:10,000) (Licor). .. The MAP1203 protein was purified on the nickel columns as per manufacturer's instructions (Clontech).

    Article Title: Chtop (Chromatin target of Prmt1) auto-regulates its expression level via intron retention and nonsense-mediated decay of its own mRNA
    Article Snippet: Paragraph title: Expression of TF-N1 ... Cells were extracted with xTractor Buffer (Clontech), and TF-N1 was pulled down using TALON Magnetic Beads (Life Technologies).

    Article Title: Evolutionarily recent, insertional fission of mitochondrial cox2 into complementary genes in bilaterian Metazoa
    Article Snippet: .. Bacteria were grown to the exponential phase, at which point the expression of recombinant proteins was induced with 1 mM IPTG at 30 °C for 6 h. Upon harvesting, the cells were disrupted using xTractor Buffer (Clontech). .. Recombinant proteins were purified using a CapturemHis-Tagged Purification Kit (Clontech).

    Article Title: Studies of recombinant TWA1 reveal constitutive dimerization
    Article Snippet: Paragraph title: Pilot experiments for TWA1 expression in Escherichia coli ... To compare the extractability of TWA1.V5His6 using different methods of bacterial cell lysis, bacterial pellets corresponding to 0 h or 3 h timepoints were either lysed by gentle agitation in 500 μl of Xtractor buffer (Clontech) at room temperature for 1 h or resuspended in 1 ml of lysis buffer (200 mM NaCl, 50 mM TRIS, pH 8.0, 10% glycerol, 25 mM imidazole and 2 mM PMSF) and disrupted by sonication for 2 min (four cycles of 5 s bursts at 25 s intervals) on ice.

    Article Title: Cloning, Assembly, and Modification of the Primary Human Cytomegalovirus Isolate Toledo by Yeast-Based Transformation-Associated Recombination
    Article Snippet: The culture was then cooled to 20°C, and isopropyl-β-d -thiogalactopyranoside (IPTG) was added to a final concentration of 0.5 mM to induce protein expression. .. Cell pellets were washed once with phosphate-buffered saline (PBS), pH 7.4, and then resuspended in 60-ml volumes of xTractor buffer supplemented with EDTA-free protease inhibitor (Clontech), 2 mg lysozyme (Sigma-Aldrich), and 625 U Benzonase (Millipore).

    Article Title: Specific Inter-residue Interactions as Determinants of Human Monoacylglycerol Lipase Catalytic Competency
    Article Snippet: Paragraph title: hMGL Mutagenesis, Expression, and Purification ... For enzyme purification, 3 g (wet weight) of cells were resuspended in 20 ml of lysis buffer (20 m m sodium phosphate, 200 m m NaCl, 1 m m DTT, pH 7.4) and 20 ml of xTractor buffer (Clontech, Mountain View, CA) supplemented with lysozyme (0.1 mg/ml) and DNase I (25 μg/ml) (Fisher).

    Construct:

    Article Title: MAP1203 Promotes Mycobacterium avium Subspecies paratuberculosis Binding and Invasion to Bovine Epithelial Cells
    Article Snippet: Resulting constructs were transformed into E. coli BL21 DE3 (Life Technologies) and grown on LB agar plates with 100 μg/ml ampicillin. .. Bacteria were lysed using the xTractor Buffer (Clontech) as per manufacturer's instructions, separated on the SDS-PAGE protein gel, probed for 6xHN-tagged MAP1203 expression using a goat anti-6xHN primary antibody (1:2,000; Clontech Laboratories, Inc), and visualized with a rabbit anti-goat IR800 secondary antibody (1:10,000) (Licor).

    Article Title: Chtop (Chromatin target of Prmt1) auto-regulates its expression level via intron retention and nonsense-mediated decay of its own mRNA
    Article Snippet: To construct N1-pCold TF DNA, a DNA fragment of HA-N1 was generated by digestion of HA-N1-pcDNA3.1 with Kpn I/Xba I and the fragment inserted at the Kpn I/Xba I site of pCold TF DNA (Takara). .. Cells were extracted with xTractor Buffer (Clontech), and TF-N1 was pulled down using TALON Magnetic Beads (Life Technologies).

    Article Title: Cloning, Assembly, and Modification of the Primary Human Cytomegalovirus Isolate Toledo by Yeast-Based Transformation-Associated Recombination
    Article Snippet: A single colony of E. coli Alpha-Select Gold Efficiency (Bioline) harboring the pJExpress404-FLAG-Cas9-His construct was cultured overnight in 50 ml LB supplemented with 100 μg/ml carbenicillin (Teknova). .. Cell pellets were washed once with phosphate-buffered saline (PBS), pH 7.4, and then resuspended in 60-ml volumes of xTractor buffer supplemented with EDTA-free protease inhibitor (Clontech), 2 mg lysozyme (Sigma-Aldrich), and 625 U Benzonase (Millipore).

    Incubation:

    Article Title: Evolutionarily recent, insertional fission of mitochondrial cox2 into complementary genes in bilaterian Metazoa
    Article Snippet: Bacteria were grown to the exponential phase, at which point the expression of recombinant proteins was induced with 1 mM IPTG at 30 °C for 6 h. Upon harvesting, the cells were disrupted using xTractor Buffer (Clontech). .. For the nuclease activity assay, 20 ng of protein was incubated with 400 ng pGEM-derived plasmid in a 20 μl reaction mixture containing 50 mM Tris-HCl (pH 8.0), 150 mM NaCl, and 2 mM MgCl2 at 37 °C for 2 h. The samples were then electrophoresed in 1% agarose gel with ethidium bromide and analyzed under UV light.

    Activity Assay:

    Article Title: A Target-Based Whole Cell Screen Approach To Identify Potential Inhibitors of Mycobacterium tuberculosis Signal Peptidase
    Article Snippet: One hundred milligrams of Triton X-114 membrane fraction pellet was solubilized in 1 mL of XTractor buffer (Clontech) and transferred to a 2 mL screw-cap tube containing 0.1 mm silica. .. The resulting cell membrane fraction was diluted 1:5 with 50 mM Tris-HCl, pH 8.0, and assayed for SPase activity using the fluorogenic peptide substrate 3-nitrotyrosine (NO)-YFSASALA∼KI-aminobenzoic acid-OH (ABZ) (California Peptide).

    Article Title: Evolutionarily recent, insertional fission of mitochondrial cox2 into complementary genes in bilaterian Metazoa
    Article Snippet: Paragraph title: Subcloning of the qnu gene in the bacterial expression vector and QNU activity assay ... Bacteria were grown to the exponential phase, at which point the expression of recombinant proteins was induced with 1 mM IPTG at 30 °C for 6 h. Upon harvesting, the cells were disrupted using xTractor Buffer (Clontech).

    Mass Spectrometry:

    Article Title: Citrullination mediated by PPAD constrains biofilm formation in P. gingivalis strain 381
    Article Snippet: .. Mass spectrometry analysis P. gingivalis stationary-phase cultures were centrifuged, supernatant was removed, and cells were lysed in xTractor Buffer containing DNase I, lysozyme, and protease inhibitor (Takara Bio USA Inc.). .. Lysates were cleared by centrifugation at 12,000×g for 20 min at 4 °C and protein concentration of cleared lysates was determined using the PierceTM BCA Protein Assay Kit (Thermo Fisher Scientific).

    BIA-KA:

    Article Title: Citrullination mediated by PPAD constrains biofilm formation in P. gingivalis strain 381
    Article Snippet: For biofilm samples, 24 h biofilms were washed twice in dH2 O and then lysed with scraping in xTractor Buffer containing DNase I, lysozyme, and protease inhibitor (Takara Bio USA Inc.). .. Lysates were cleared by centrifugation at 12,000×g for 20 min at 4 °C and protein concentration of cleared lysates was determined using the PierceTM BCA Protein Assay Kit (Thermo Fisher Scientific).

    Article Title: Chtop (Chromatin target of Prmt1) auto-regulates its expression level via intron retention and nonsense-mediated decay of its own mRNA
    Article Snippet: Cells were extracted with xTractor Buffer (Clontech), and TF-N1 was pulled down using TALON Magnetic Beads (Life Technologies). .. The eluted solution containing TF-N1 was dialyzed with Tris-buffered saline for 6 h at 4°C, and purified TF-N1 was quantified with the BCA protein assay (Thermo Scientific).

    Article Title: Citrullination mediated by PPAD constrains biofilm formation in P. gingivalis strain 381
    Article Snippet: Mass spectrometry analysis P. gingivalis stationary-phase cultures were centrifuged, supernatant was removed, and cells were lysed in xTractor Buffer containing DNase I, lysozyme, and protease inhibitor (Takara Bio USA Inc.). .. Lysates were cleared by centrifugation at 12,000×g for 20 min at 4 °C and protein concentration of cleared lysates was determined using the PierceTM BCA Protein Assay Kit (Thermo Fisher Scientific).

    Western Blot:

    Article Title: Adiponectin and Its Receptors Are Differentially Expressed in Human Tissues and Cell Lines of Distinct Origin
    Article Snippet: Paragraph title: Protein Extraction and Western-Blot Analysis ... Proteins were extracted from frozen cell pellets with xTractor buffer (Clontech) supplemented with protease inhibitor cocktail (Sigma-Aldrich, St. Louis, MO, USA).

    Article Title: Citrullination mediated by PPAD constrains biofilm formation in P. gingivalis strain 381
    Article Snippet: Paragraph title: Immunoblots ... For biofilm samples, 24 h biofilms were washed twice in dH2 O and then lysed with scraping in xTractor Buffer containing DNase I, lysozyme, and protease inhibitor (Takara Bio USA Inc.).

    Article Title: Nutrient-driven O-linked N-acetylglucosamine (O-GlcNAc) cycling impacts neurodevelopmental timing and metabolism
    Article Snippet: Paragraph title: SDS-PAGE and Western blotting ... Protein was extracted with Tissue Protein Extraction Reagent (T-PER) tissue protein extraction reagent (Thermo Scientific) or Xtractor buffer (Clontech).

    Transformation Assay:

    Article Title: Polymyxin B Resistance and Biofilm Formation in Vibrio cholerae Are Controlled by the Response Regulator CarR
    Article Snippet: The carR overexpression plasmid (p carR ) was transformed into E. coli strains TOP10 and BL21(DE3) for maintenance and protein production, respectively. .. Cells were harvested and then lysed with xTractor buffer (20 ml/g) (Clontech Laboratories, Mountain View, CA) for 10 min with orbital agitation at 4°C.

    Article Title: MAP1203 Promotes Mycobacterium avium Subspecies paratuberculosis Binding and Invasion to Bovine Epithelial Cells
    Article Snippet: Resulting constructs were transformed into E. coli BL21 DE3 (Life Technologies) and grown on LB agar plates with 100 μg/ml ampicillin. .. Bacteria were lysed using the xTractor Buffer (Clontech) as per manufacturer's instructions, separated on the SDS-PAGE protein gel, probed for 6xHN-tagged MAP1203 expression using a goat anti-6xHN primary antibody (1:2,000; Clontech Laboratories, Inc), and visualized with a rabbit anti-goat IR800 secondary antibody (1:10,000) (Licor).

    Over Expression:

    Article Title: Polymyxin B Resistance and Biofilm Formation in Vibrio cholerae Are Controlled by the Response Regulator CarR
    Article Snippet: The carR overexpression plasmid (p carR ) was transformed into E. coli strains TOP10 and BL21(DE3) for maintenance and protein production, respectively. .. Cells were harvested and then lysed with xTractor buffer (20 ml/g) (Clontech Laboratories, Mountain View, CA) for 10 min with orbital agitation at 4°C.

    Flow Cytometry:

    Article Title: Polymyxin B Resistance and Biofilm Formation in Vibrio cholerae Are Controlled by the Response Regulator CarR
    Article Snippet: Cells were harvested and then lysed with xTractor buffer (20 ml/g) (Clontech Laboratories, Mountain View, CA) for 10 min with orbital agitation at 4°C. .. The protein was purified with a batch/gravity flow protocol at 4°C using preequilibrated Talon resin (equilibration buffer: 50 mM sodium phosphate, 300 mM NaCl; pH 7.4).

    Protease Inhibitor:

    Article Title: Adiponectin and Its Receptors Are Differentially Expressed in Human Tissues and Cell Lines of Distinct Origin
    Article Snippet: .. Proteins were extracted from frozen cell pellets with xTractor buffer (Clontech) supplemented with protease inhibitor cocktail (Sigma-Aldrich, St. Louis, MO, USA). ..

    Article Title: Citrullination mediated by PPAD constrains biofilm formation in P. gingivalis strain 381
    Article Snippet: .. For biofilm samples, 24 h biofilms were washed twice in dH2 O and then lysed with scraping in xTractor Buffer containing DNase I, lysozyme, and protease inhibitor (Takara Bio USA Inc.). .. Lysates were cleared by centrifugation at 12,000×g for 20 min at 4 °C and protein concentration of cleared lysates was determined using the PierceTM BCA Protein Assay Kit (Thermo Fisher Scientific).

    Article Title: Cloning, Assembly, and Modification of the Primary Human Cytomegalovirus Isolate Toledo by Yeast-Based Transformation-Associated Recombination
    Article Snippet: .. Cell pellets were washed once with phosphate-buffered saline (PBS), pH 7.4, and then resuspended in 60-ml volumes of xTractor buffer supplemented with EDTA-free protease inhibitor (Clontech), 2 mg lysozyme (Sigma-Aldrich), and 625 U Benzonase (Millipore). .. Cell suspensions were then gently rotated for 2 h at 4°C prior to mechanical lysis in a Microfluidizer M-110L (Microfluidics).

    Article Title: Citrullination mediated by PPAD constrains biofilm formation in P. gingivalis strain 381
    Article Snippet: .. Mass spectrometry analysis P. gingivalis stationary-phase cultures were centrifuged, supernatant was removed, and cells were lysed in xTractor Buffer containing DNase I, lysozyme, and protease inhibitor (Takara Bio USA Inc.). .. Lysates were cleared by centrifugation at 12,000×g for 20 min at 4 °C and protein concentration of cleared lysates was determined using the PierceTM BCA Protein Assay Kit (Thermo Fisher Scientific).

    Cell Culture:

    Article Title: Cloning, Assembly, and Modification of the Primary Human Cytomegalovirus Isolate Toledo by Yeast-Based Transformation-Associated Recombination
    Article Snippet: A single colony of E. coli Alpha-Select Gold Efficiency (Bioline) harboring the pJExpress404-FLAG-Cas9-His construct was cultured overnight in 50 ml LB supplemented with 100 μg/ml carbenicillin (Teknova). .. Cell pellets were washed once with phosphate-buffered saline (PBS), pH 7.4, and then resuspended in 60-ml volumes of xTractor buffer supplemented with EDTA-free protease inhibitor (Clontech), 2 mg lysozyme (Sigma-Aldrich), and 625 U Benzonase (Millipore).

    Generated:

    Article Title: Electron Microscopy and 3D Reconstruction of F-actin Decorated with Cardiac Myosin-binding Protein C (cMyBP-C)
    Article Snippet: The cDNA fragments were generated by PCR, incorporating compatible restriction enzyme sites for cloning into the pET28a vector, with 5’ Nco1 and a 3’ Xho1 sites. .. Cells were collected by centrifugation and lysed using the xTractor Buffer supplied in the HisTalon Buffer set (Clontech).

    Article Title: Chtop (Chromatin target of Prmt1) auto-regulates its expression level via intron retention and nonsense-mediated decay of its own mRNA
    Article Snippet: To construct N1-pCold TF DNA, a DNA fragment of HA-N1 was generated by digestion of HA-N1-pcDNA3.1 with Kpn I/Xba I and the fragment inserted at the Kpn I/Xba I site of pCold TF DNA (Takara). .. Cells were extracted with xTractor Buffer (Clontech), and TF-N1 was pulled down using TALON Magnetic Beads (Life Technologies).

    Protein Concentration:

    Article Title: Citrullination mediated by PPAD constrains biofilm formation in P. gingivalis strain 381
    Article Snippet: For biofilm samples, 24 h biofilms were washed twice in dH2 O and then lysed with scraping in xTractor Buffer containing DNase I, lysozyme, and protease inhibitor (Takara Bio USA Inc.). .. Lysates were cleared by centrifugation at 12,000×g for 20 min at 4 °C and protein concentration of cleared lysates was determined using the PierceTM BCA Protein Assay Kit (Thermo Fisher Scientific).

    Article Title: Citrullination mediated by PPAD constrains biofilm formation in P. gingivalis strain 381
    Article Snippet: Mass spectrometry analysis P. gingivalis stationary-phase cultures were centrifuged, supernatant was removed, and cells were lysed in xTractor Buffer containing DNase I, lysozyme, and protease inhibitor (Takara Bio USA Inc.). .. Lysates were cleared by centrifugation at 12,000×g for 20 min at 4 °C and protein concentration of cleared lysates was determined using the PierceTM BCA Protein Assay Kit (Thermo Fisher Scientific).

    Sequencing:

    Article Title: Mating in the closest living relatives of animals is induced by a bacterial chondroitinase
    Article Snippet: After 24 hours, cells were pelleted and lysed with xTractor™ buffer (Clontech), purified with HisTALON™ gravity columns (Clontech), and the His-tag was released by enterokinase cleavage (Millipore enterokinase cleave capture kit #69067). .. A gBlock of the eroS gene sequence harboring H278A and Y287A mutations was purchased from Integrated DNA Technologies and ligated into the pET15b vector (Novagen) at NdeI and BamHI restriction sites.

    Sonication:

    Article Title: Studies of recombinant TWA1 reveal constitutive dimerization
    Article Snippet: .. To compare the extractability of TWA1.V5His6 using different methods of bacterial cell lysis, bacterial pellets corresponding to 0 h or 3 h timepoints were either lysed by gentle agitation in 500 μl of Xtractor buffer (Clontech) at room temperature for 1 h or resuspended in 1 ml of lysis buffer (200 mM NaCl, 50 mM TRIS, pH 8.0, 10% glycerol, 25 mM imidazole and 2 mM PMSF) and disrupted by sonication for 2 min (four cycles of 5 s bursts at 25 s intervals) on ice. ..

    Article Title: Specific Inter-residue Interactions as Determinants of Human Monoacylglycerol Lipase Catalytic Competency
    Article Snippet: For enzyme purification, 3 g (wet weight) of cells were resuspended in 20 ml of lysis buffer (20 m m sodium phosphate, 200 m m NaCl, 1 m m DTT, pH 7.4) and 20 ml of xTractor buffer (Clontech, Mountain View, CA) supplemented with lysozyme (0.1 mg/ml) and DNase I (25 μg/ml) (Fisher). .. After 20 min the lysate was disrupted on ice by three 1-min sonication cycles, each consisting of 1-s sonication bursts at a 50-watt power level separated by a 5-s interval (Vibra-Cell 500 W, Sonics, Newtown, CT).

    Affinity Purification:

    Article Title: Polymyxin B Resistance and Biofilm Formation in Vibrio cholerae Are Controlled by the Response Regulator CarR
    Article Snippet: CarR-mycHis was purified by metal affinity purification using a Talon resin (Clontech Laboratories, Mountain View, CA) according to the manufacturer's instructions. .. Cells were harvested and then lysed with xTractor buffer (20 ml/g) (Clontech Laboratories, Mountain View, CA) for 10 min with orbital agitation at 4°C.

    Recombinant:

    Article Title: Polymyxin B Resistance and Biofilm Formation in Vibrio cholerae Are Controlled by the Response Regulator CarR
    Article Snippet: Briefly, cultures of BL21(DE3) harboring p carR were grown to mid-exponential phase in LB medium supplemented with ampicillin at 37°C, and expression of the recombinant protein was induced for 3 h at 30°C with the addition of 0.2% arabinose. .. Cells were harvested and then lysed with xTractor buffer (20 ml/g) (Clontech Laboratories, Mountain View, CA) for 10 min with orbital agitation at 4°C.

    Article Title: Evolutionarily recent, insertional fission of mitochondrial cox2 into complementary genes in bilaterian Metazoa
    Article Snippet: .. Bacteria were grown to the exponential phase, at which point the expression of recombinant proteins was induced with 1 mM IPTG at 30 °C for 6 h. Upon harvesting, the cells were disrupted using xTractor Buffer (Clontech). .. Recombinant proteins were purified using a CapturemHis-Tagged Purification Kit (Clontech).

    Article Title: Cloning, Assembly, and Modification of the Primary Human Cytomegalovirus Isolate Toledo by Yeast-Based Transformation-Associated Recombination
    Article Snippet: Cell pellets were washed once with phosphate-buffered saline (PBS), pH 7.4, and then resuspended in 60-ml volumes of xTractor buffer supplemented with EDTA-free protease inhibitor (Clontech), 2 mg lysozyme (Sigma-Aldrich), and 625 U Benzonase (Millipore). .. Cell debris was pelleted at 8,000 × g for 1 h, and recombinant protein was purified from the supernatant using a HisTalon column (Clontech) according to the manufacturer’s instructions.

    Magnetic Beads:

    Article Title: Chtop (Chromatin target of Prmt1) auto-regulates its expression level via intron retention and nonsense-mediated decay of its own mRNA
    Article Snippet: .. Cells were extracted with xTractor Buffer (Clontech), and TF-N1 was pulled down using TALON Magnetic Beads (Life Technologies). .. The eluted solution containing TF-N1 was dialyzed with Tris-buffered saline for 6 h at 4°C, and purified TF-N1 was quantified with the BCA protein assay (Thermo Scientific).

    Mutagenesis:

    Article Title: Mating in the closest living relatives of animals is induced by a bacterial chondroitinase
    Article Snippet: After 24 hours, cells were pelleted and lysed with xTractor™ buffer (Clontech), purified with HisTALON™ gravity columns (Clontech), and the His-tag was released by enterokinase cleavage (Millipore enterokinase cleave capture kit #69067). .. The mutant eroS gene was transformed into OneShot BL21 Star (DE3) cells (Invitrogen) for expression.

    Article Title: Specific Inter-residue Interactions as Determinants of Human Monoacylglycerol Lipase Catalytic Competency
    Article Snippet: Paragraph title: hMGL Mutagenesis, Expression, and Purification ... For enzyme purification, 3 g (wet weight) of cells were resuspended in 20 ml of lysis buffer (20 m m sodium phosphate, 200 m m NaCl, 1 m m DTT, pH 7.4) and 20 ml of xTractor buffer (Clontech, Mountain View, CA) supplemented with lysozyme (0.1 mg/ml) and DNase I (25 μg/ml) (Fisher).

    Isolation:

    Article Title: A Target-Based Whole Cell Screen Approach To Identify Potential Inhibitors of Mycobacterium tuberculosis Signal Peptidase
    Article Snippet: LepB Membrane Assay Cell membrane fractions were isolated from γ-irradiated whole M. tuberculosis H37Rv (NR-14819; BEI resources). .. One hundred milligrams of Triton X-114 membrane fraction pellet was solubilized in 1 mL of XTractor buffer (Clontech) and transferred to a 2 mL screw-cap tube containing 0.1 mm silica.

    Article Title: Specific Inter-residue Interactions as Determinants of Human Monoacylglycerol Lipase Catalytic Competency
    Article Snippet: For enzyme purification, 3 g (wet weight) of cells were resuspended in 20 ml of lysis buffer (20 m m sodium phosphate, 200 m m NaCl, 1 m m DTT, pH 7.4) and 20 ml of xTractor buffer (Clontech, Mountain View, CA) supplemented with lysozyme (0.1 mg/ml) and DNase I (25 μg/ml) (Fisher). .. The resulting cell lysate was centrifuged at 20, 000 × g for 25 min at 4 °C. hMGL was isolated by incubating the resulting supernatant with 4.0 ml (bed volume) pre-equilibrated Talon metal affinity resin (Clontech) for 1 h at 4 °C.

    Subcloning:

    Article Title: Evolutionarily recent, insertional fission of mitochondrial cox2 into complementary genes in bilaterian Metazoa
    Article Snippet: Paragraph title: Subcloning of the qnu gene in the bacterial expression vector and QNU activity assay ... Bacteria were grown to the exponential phase, at which point the expression of recombinant proteins was induced with 1 mM IPTG at 30 °C for 6 h. Upon harvesting, the cells were disrupted using xTractor Buffer (Clontech).

    Purification:

    Article Title: Mating in the closest living relatives of animals is induced by a bacterial chondroitinase
    Article Snippet: .. After 24 hours, cells were pelleted and lysed with xTractor™ buffer (Clontech), purified with HisTALON™ gravity columns (Clontech), and the His-tag was released by enterokinase cleavage (Millipore enterokinase cleave capture kit #69067). .. A gBlock of the eroS gene sequence harboring H278A and Y287A mutations was purchased from Integrated DNA Technologies and ligated into the pET15b vector (Novagen) at NdeI and BamHI restriction sites.

    Article Title: Electron Microscopy and 3D Reconstruction of F-actin Decorated with Cardiac Myosin-binding Protein C (cMyBP-C)
    Article Snippet: F-actin was purified from chicken pectoralis muscle according to Pardee et al. . cMyBP-C fragments were bacterially expressed from mouse cardiac cDNA using the pET expression system (Novagen, Madison, WI). .. Cells were collected by centrifugation and lysed using the xTractor Buffer supplied in the HisTalon Buffer set (Clontech).

    Article Title: Polymyxin B Resistance and Biofilm Formation in Vibrio cholerae Are Controlled by the Response Regulator CarR
    Article Snippet: Paragraph title: Protein production and purification. ... Cells were harvested and then lysed with xTractor buffer (20 ml/g) (Clontech Laboratories, Mountain View, CA) for 10 min with orbital agitation at 4°C.

    Article Title: MAP1203 Promotes Mycobacterium avium Subspecies paratuberculosis Binding and Invasion to Bovine Epithelial Cells
    Article Snippet: Paragraph title: Purification of MAP1203 ... Bacteria were lysed using the xTractor Buffer (Clontech) as per manufacturer's instructions, separated on the SDS-PAGE protein gel, probed for 6xHN-tagged MAP1203 expression using a goat anti-6xHN primary antibody (1:2,000; Clontech Laboratories, Inc), and visualized with a rabbit anti-goat IR800 secondary antibody (1:10,000) (Licor).

    Article Title: Chtop (Chromatin target of Prmt1) auto-regulates its expression level via intron retention and nonsense-mediated decay of its own mRNA
    Article Snippet: Cells were extracted with xTractor Buffer (Clontech), and TF-N1 was pulled down using TALON Magnetic Beads (Life Technologies). .. The eluted solution containing TF-N1 was dialyzed with Tris-buffered saline for 6 h at 4°C, and purified TF-N1 was quantified with the BCA protein assay (Thermo Scientific).

    Article Title: Evolutionarily recent, insertional fission of mitochondrial cox2 into complementary genes in bilaterian Metazoa
    Article Snippet: Bacteria were grown to the exponential phase, at which point the expression of recombinant proteins was induced with 1 mM IPTG at 30 °C for 6 h. Upon harvesting, the cells were disrupted using xTractor Buffer (Clontech). .. Recombinant proteins were purified using a CapturemHis-Tagged Purification Kit (Clontech).

    Article Title: Cloning, Assembly, and Modification of the Primary Human Cytomegalovirus Isolate Toledo by Yeast-Based Transformation-Associated Recombination
    Article Snippet: Paragraph title: Synthesis and purification of Cas9 and chimeric synthetic guide RNAs. ... Cell pellets were washed once with phosphate-buffered saline (PBS), pH 7.4, and then resuspended in 60-ml volumes of xTractor buffer supplemented with EDTA-free protease inhibitor (Clontech), 2 mg lysozyme (Sigma-Aldrich), and 625 U Benzonase (Millipore).

    Article Title: Specific Inter-residue Interactions as Determinants of Human Monoacylglycerol Lipase Catalytic Competency
    Article Snippet: .. For enzyme purification, 3 g (wet weight) of cells were resuspended in 20 ml of lysis buffer (20 m m sodium phosphate, 200 m m NaCl, 1 m m DTT, pH 7.4) and 20 ml of xTractor buffer (Clontech, Mountain View, CA) supplemented with lysozyme (0.1 mg/ml) and DNase I (25 μg/ml) (Fisher). .. After 20 min the lysate was disrupted on ice by three 1-min sonication cycles, each consisting of 1-s sonication bursts at a 50-watt power level separated by a 5-s interval (Vibra-Cell 500 W, Sonics, Newtown, CT).

    Protein Purification:

    Article Title: Human Adenosine A2A Receptor Binds Calmodulin with High Affinity in a Calcium-Dependent Manner
    Article Snippet: Paragraph title: Protein purification ... BL21(DE3) cells with either A2A-ctL or A2A-ctS were suspended in xTractor Buffer (Takara Bio Europe/Clontech, Saint-Germain-en-Laye, France), containing complete EDTA-free protease inhibitors (Roche, Basel, Switzerland).

    Polymerase Chain Reaction:

    Article Title: Mating in the closest living relatives of animals is induced by a bacterial chondroitinase
    Article Snippet: The eroS gene was amplified by PCR (Phusion® DNA polymerase, New England Biosciences) from V. fischeri ES114 genomic DNA (Forward primer: 5’-GCCTCTGTCGACGCAAAAAATACCCAAACACCAC; Reverse primer: 5’-AATTAAGCGGCCGCCGTCTTGAATTGTTACTTGGAAAGAATAAG). .. After 24 hours, cells were pelleted and lysed with xTractor™ buffer (Clontech), purified with HisTALON™ gravity columns (Clontech), and the His-tag was released by enterokinase cleavage (Millipore enterokinase cleave capture kit #69067).

    Article Title: Electron Microscopy and 3D Reconstruction of F-actin Decorated with Cardiac Myosin-binding Protein C (cMyBP-C)
    Article Snippet: The cDNA fragments were generated by PCR, incorporating compatible restriction enzyme sites for cloning into the pET28a vector, with 5’ Nco1 and a 3’ Xho1 sites. .. Cells were collected by centrifugation and lysed using the xTractor Buffer supplied in the HisTalon Buffer set (Clontech).

    Article Title: Polymyxin B Resistance and Biofilm Formation in Vibrio cholerae Are Controlled by the Response Regulator CarR
    Article Snippet: The coding region of carR , excluding the stop codon, was amplified by PCR and cloned into the pBAD/Myc-His C bacterial expression vector (Life Technologies, Carlsbad, CA), resulting in an overexpression plasmid encoding a recombinant CarR protein with a C-terminal Myc and 6×His tags (CarR-mycHis). .. Cells were harvested and then lysed with xTractor buffer (20 ml/g) (Clontech Laboratories, Mountain View, CA) for 10 min with orbital agitation at 4°C.

    Article Title: Evolutionarily recent, insertional fission of mitochondrial cox2 into complementary genes in bilaterian Metazoa
    Article Snippet: Subcloning of the qnu gene in the bacterial expression vector and QNU activity assay The full-length qnu and its truncated allele, Δqnu , missing the N-terminal 30% encoding the DNA-binding QN domain, were PCR amplified using the following primers: qnuR (5’-GAGGTACCTGGATTAATATAATTTTATGGTCGAGGAG-3’), and qnuF1 (5’- AGGGATCCATGAATTATAATCATACTGGTCAATATAAAAC-3’) or qnuF2 (5’- AGGGATCCATGTTACCTCATAATAATAATCTTCCTAATTT-3’), respectively. .. Bacteria were grown to the exponential phase, at which point the expression of recombinant proteins was induced with 1 mM IPTG at 30 °C for 6 h. Upon harvesting, the cells were disrupted using xTractor Buffer (Clontech).

    Protein Extraction:

    Article Title: Adiponectin and Its Receptors Are Differentially Expressed in Human Tissues and Cell Lines of Distinct Origin
    Article Snippet: Paragraph title: Protein Extraction and Western-Blot Analysis ... Proteins were extracted from frozen cell pellets with xTractor buffer (Clontech) supplemented with protease inhibitor cocktail (Sigma-Aldrich, St. Louis, MO, USA).

    Article Title: Nutrient-driven O-linked N-acetylglucosamine (O-GlcNAc) cycling impacts neurodevelopmental timing and metabolism
    Article Snippet: .. Protein was extracted with Tissue Protein Extraction Reagent (T-PER) tissue protein extraction reagent (Thermo Scientific) or Xtractor buffer (Clontech). ..

    Positron Emission Tomography:

    Article Title: Electron Microscopy and 3D Reconstruction of F-actin Decorated with Cardiac Myosin-binding Protein C (cMyBP-C)
    Article Snippet: F-actin was purified from chicken pectoralis muscle according to Pardee et al. . cMyBP-C fragments were bacterially expressed from mouse cardiac cDNA using the pET expression system (Novagen, Madison, WI). .. Cells were collected by centrifugation and lysed using the xTractor Buffer supplied in the HisTalon Buffer set (Clontech).

    Polyacrylamide Gel Electrophoresis:

    Article Title: Human Adenosine A2A Receptor Binds Calmodulin with High Affinity in a Calcium-Dependent Manner
    Article Snippet: BL21(DE3) cells with either A2A-ctL or A2A-ctS were suspended in xTractor Buffer (Takara Bio Europe/Clontech, Saint-Germain-en-Laye, France), containing complete EDTA-free protease inhibitors (Roche, Basel, Switzerland). .. Bound proteins were eluted with elution buffer (50 mM HEPES, 500 mM NaCl, 10% glycerol, 500 mM imidazole pH 7.4), and eluates were analyzed by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE).

    Lysis:

    Article Title: Studies of recombinant TWA1 reveal constitutive dimerization
    Article Snippet: .. To compare the extractability of TWA1.V5His6 using different methods of bacterial cell lysis, bacterial pellets corresponding to 0 h or 3 h timepoints were either lysed by gentle agitation in 500 μl of Xtractor buffer (Clontech) at room temperature for 1 h or resuspended in 1 ml of lysis buffer (200 mM NaCl, 50 mM TRIS, pH 8.0, 10% glycerol, 25 mM imidazole and 2 mM PMSF) and disrupted by sonication for 2 min (four cycles of 5 s bursts at 25 s intervals) on ice. ..

    Article Title: Cloning, Assembly, and Modification of the Primary Human Cytomegalovirus Isolate Toledo by Yeast-Based Transformation-Associated Recombination
    Article Snippet: Cell pellets were washed once with phosphate-buffered saline (PBS), pH 7.4, and then resuspended in 60-ml volumes of xTractor buffer supplemented with EDTA-free protease inhibitor (Clontech), 2 mg lysozyme (Sigma-Aldrich), and 625 U Benzonase (Millipore). .. Cell suspensions were then gently rotated for 2 h at 4°C prior to mechanical lysis in a Microfluidizer M-110L (Microfluidics).

    Article Title: Specific Inter-residue Interactions as Determinants of Human Monoacylglycerol Lipase Catalytic Competency
    Article Snippet: .. For enzyme purification, 3 g (wet weight) of cells were resuspended in 20 ml of lysis buffer (20 m m sodium phosphate, 200 m m NaCl, 1 m m DTT, pH 7.4) and 20 ml of xTractor buffer (Clontech, Mountain View, CA) supplemented with lysozyme (0.1 mg/ml) and DNase I (25 μg/ml) (Fisher). .. After 20 min the lysate was disrupted on ice by three 1-min sonication cycles, each consisting of 1-s sonication bursts at a 50-watt power level separated by a 5-s interval (Vibra-Cell 500 W, Sonics, Newtown, CT).

    SDS Page:

    Article Title: Citrullination mediated by PPAD constrains biofilm formation in P. gingivalis strain 381
    Article Snippet: For biofilm samples, 24 h biofilms were washed twice in dH2 O and then lysed with scraping in xTractor Buffer containing DNase I, lysozyme, and protease inhibitor (Takara Bio USA Inc.). .. Planktonic or biofilm samples were mixed with 2X SDS sample buffer and denatured by heating at 100 °C for 5 min. Planktonic samples (10 µl) or biofilm samples (5 µg) were th en electrophoresed on a 12% polyacrylamide SDS–PAGE gel.

    Article Title: Nutrient-driven O-linked N-acetylglucosamine (O-GlcNAc) cycling impacts neurodevelopmental timing and metabolism
    Article Snippet: Paragraph title: SDS-PAGE and Western blotting ... Protein was extracted with Tissue Protein Extraction Reagent (T-PER) tissue protein extraction reagent (Thermo Scientific) or Xtractor buffer (Clontech).

    Article Title: Human Adenosine A2A Receptor Binds Calmodulin with High Affinity in a Calcium-Dependent Manner
    Article Snippet: BL21(DE3) cells with either A2A-ctL or A2A-ctS were suspended in xTractor Buffer (Takara Bio Europe/Clontech, Saint-Germain-en-Laye, France), containing complete EDTA-free protease inhibitors (Roche, Basel, Switzerland). .. Bound proteins were eluted with elution buffer (50 mM HEPES, 500 mM NaCl, 10% glycerol, 500 mM imidazole pH 7.4), and eluates were analyzed by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE).

    Article Title: MAP1203 Promotes Mycobacterium avium Subspecies paratuberculosis Binding and Invasion to Bovine Epithelial Cells
    Article Snippet: .. Bacteria were lysed using the xTractor Buffer (Clontech) as per manufacturer's instructions, separated on the SDS-PAGE protein gel, probed for 6xHN-tagged MAP1203 expression using a goat anti-6xHN primary antibody (1:2,000; Clontech Laboratories, Inc), and visualized with a rabbit anti-goat IR800 secondary antibody (1:10,000) (Licor). .. The MAP1203 protein was purified on the nickel columns as per manufacturer's instructions (Clontech).

    Article Title: Studies of recombinant TWA1 reveal constitutive dimerization
    Article Snippet: To compare the extractability of TWA1.V5His6 using different methods of bacterial cell lysis, bacterial pellets corresponding to 0 h or 3 h timepoints were either lysed by gentle agitation in 500 μl of Xtractor buffer (Clontech) at room temperature for 1 h or resuspended in 1 ml of lysis buffer (200 mM NaCl, 50 mM TRIS, pH 8.0, 10% glycerol, 25 mM imidazole and 2 mM PMSF) and disrupted by sonication for 2 min (four cycles of 5 s bursts at 25 s intervals) on ice. .. Pellets were resuspended in 100 μl 2× SDS/PAGE sample buffer containing 100 mM DTT and supernatants were mixed 1:1 with the same buffer.

    Article Title: Citrullination mediated by PPAD constrains biofilm formation in P. gingivalis strain 381
    Article Snippet: Mass spectrometry analysis P. gingivalis stationary-phase cultures were centrifuged, supernatant was removed, and cells were lysed in xTractor Buffer containing DNase I, lysozyme, and protease inhibitor (Takara Bio USA Inc.). .. Thirty micrograms of total protein was run on a 12% SDS–PAGE gel until the dye front was 5–10 mm down the resolving gel.

    Plasmid Preparation:

    Article Title: Mating in the closest living relatives of animals is induced by a bacterial chondroitinase
    Article Snippet: After digestion with SalI -HF and NotI -HF (New England Biolabs), the eroS gene was ligated in-frame into a pET6xHN-N vector (Clontech) for fusion to a His-tag and transformed into OneShot BL21(DE3) cells (Invitrogen) for expression. .. After 24 hours, cells were pelleted and lysed with xTractor™ buffer (Clontech), purified with HisTALON™ gravity columns (Clontech), and the His-tag was released by enterokinase cleavage (Millipore enterokinase cleave capture kit #69067).

    Article Title: Electron Microscopy and 3D Reconstruction of F-actin Decorated with Cardiac Myosin-binding Protein C (cMyBP-C)
    Article Snippet: The cDNA fragments were generated by PCR, incorporating compatible restriction enzyme sites for cloning into the pET28a vector, with 5’ Nco1 and a 3’ Xho1 sites. .. Cells were collected by centrifugation and lysed using the xTractor Buffer supplied in the HisTalon Buffer set (Clontech).

    Article Title: Polymyxin B Resistance and Biofilm Formation in Vibrio cholerae Are Controlled by the Response Regulator CarR
    Article Snippet: The carR overexpression plasmid (p carR ) was transformed into E. coli strains TOP10 and BL21(DE3) for maintenance and protein production, respectively. .. Cells were harvested and then lysed with xTractor buffer (20 ml/g) (Clontech Laboratories, Mountain View, CA) for 10 min with orbital agitation at 4°C.

    Article Title: MAP1203 Promotes Mycobacterium avium Subspecies paratuberculosis Binding and Invasion to Bovine Epithelial Cells
    Article Snippet: Fragments were cloned into the HindIII/EcoI restriction sites of the pET6xHN-N-terminal expression vector. .. Bacteria were lysed using the xTractor Buffer (Clontech) as per manufacturer's instructions, separated on the SDS-PAGE protein gel, probed for 6xHN-tagged MAP1203 expression using a goat anti-6xHN primary antibody (1:2,000; Clontech Laboratories, Inc), and visualized with a rabbit anti-goat IR800 secondary antibody (1:10,000) (Licor).

    Article Title: Evolutionarily recent, insertional fission of mitochondrial cox2 into complementary genes in bilaterian Metazoa
    Article Snippet: Paragraph title: Subcloning of the qnu gene in the bacterial expression vector and QNU activity assay ... Bacteria were grown to the exponential phase, at which point the expression of recombinant proteins was induced with 1 mM IPTG at 30 °C for 6 h. Upon harvesting, the cells were disrupted using xTractor Buffer (Clontech).

    Binding Assay:

    Article Title: Human Adenosine A2A Receptor Binds Calmodulin with High Affinity in a Calcium-Dependent Manner
    Article Snippet: BL21(DE3) cells with either A2A-ctL or A2A-ctS were suspended in xTractor Buffer (Takara Bio Europe/Clontech, Saint-Germain-en-Laye, France), containing complete EDTA-free protease inhibitors (Roche, Basel, Switzerland). .. After 1 h of binding, the mixture was transferred into an empty column, and unbound proteins were washed away using the wash buffer.

    Agarose Gel Electrophoresis:

    Article Title: Evolutionarily recent, insertional fission of mitochondrial cox2 into complementary genes in bilaterian Metazoa
    Article Snippet: Bacteria were grown to the exponential phase, at which point the expression of recombinant proteins was induced with 1 mM IPTG at 30 °C for 6 h. Upon harvesting, the cells were disrupted using xTractor Buffer (Clontech). .. For the nuclease activity assay, 20 ng of protein was incubated with 400 ng pGEM-derived plasmid in a 20 μl reaction mixture containing 50 mM Tris-HCl (pH 8.0), 150 mM NaCl, and 2 mM MgCl2 at 37 °C for 2 h. The samples were then electrophoresed in 1% agarose gel with ethidium bromide and analyzed under UV light.

    Concentration Assay:

    Article Title: Studies of recombinant TWA1 reveal constitutive dimerization
    Article Snippet: Expression of TWA1.V5His6 was induced in one of these by the addition of IPTG to a final concentration of 1 mM. .. To compare the extractability of TWA1.V5His6 using different methods of bacterial cell lysis, bacterial pellets corresponding to 0 h or 3 h timepoints were either lysed by gentle agitation in 500 μl of Xtractor buffer (Clontech) at room temperature for 1 h or resuspended in 1 ml of lysis buffer (200 mM NaCl, 50 mM TRIS, pH 8.0, 10% glycerol, 25 mM imidazole and 2 mM PMSF) and disrupted by sonication for 2 min (four cycles of 5 s bursts at 25 s intervals) on ice.

    Article Title: Cloning, Assembly, and Modification of the Primary Human Cytomegalovirus Isolate Toledo by Yeast-Based Transformation-Associated Recombination
    Article Snippet: The culture was then cooled to 20°C, and isopropyl-β-d -thiogalactopyranoside (IPTG) was added to a final concentration of 0.5 mM to induce protein expression. .. Cell pellets were washed once with phosphate-buffered saline (PBS), pH 7.4, and then resuspended in 60-ml volumes of xTractor buffer supplemented with EDTA-free protease inhibitor (Clontech), 2 mg lysozyme (Sigma-Aldrich), and 625 U Benzonase (Millipore).

    Article Title: Specific Inter-residue Interactions as Determinants of Human Monoacylglycerol Lipase Catalytic Competency
    Article Snippet: Expression was induced by adding isopropyl-β- d -thiogalactopyranoside (Fisher) to a final concentration of 1 m m . .. For enzyme purification, 3 g (wet weight) of cells were resuspended in 20 ml of lysis buffer (20 m m sodium phosphate, 200 m m NaCl, 1 m m DTT, pH 7.4) and 20 ml of xTractor buffer (Clontech, Mountain View, CA) supplemented with lysozyme (0.1 mg/ml) and DNase I (25 μg/ml) (Fisher).

    Staining:

    Article Title: Citrullination mediated by PPAD constrains biofilm formation in P. gingivalis strain 381
    Article Snippet: Mass spectrometry analysis P. gingivalis stationary-phase cultures were centrifuged, supernatant was removed, and cells were lysed in xTractor Buffer containing DNase I, lysozyme, and protease inhibitor (Takara Bio USA Inc.). .. The gel was stained with Bio-Safe Coomassie G-250 (BioRad) and the entirety of stained protein was carefully cut from the gel and then cut into 1–2 mm cubes.

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    TaKaRa talon xtractor buffer
    Talon Xtractor Buffer, supplied by TaKaRa, used in various techniques. Bioz Stars score: 90/100, based on 5 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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