ugi  (New England Biolabs)


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    Name:
    Uracil Glycosylase Inhibitor UGI
    Description:
    Uracil Glycosylase Inhibitor UGI 1 000 units
    Catalog Number:
    m0281l
    Price:
    288
    Size:
    1 000 units
    Category:
    Enzyme Inhibitors
    Buy from Supplier


    Structured Review

    New England Biolabs ugi
    Uracil Glycosylase Inhibitor UGI
    Uracil Glycosylase Inhibitor UGI 1 000 units
    https://www.bioz.com/result/ugi/product/New England Biolabs
    Average 90 stars, based on 5 article reviews
    Price from $9.99 to $1999.99
    ugi - by Bioz Stars, 2020-01
    90/100 stars

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    Related Articles

    Amplification:

    Article Title: Manipulation of Plant Defense Responses by the Tomato Psyllid (Bactericerca cockerelli) and Its Associated Endosymbiont Candidatus Liberibacter Psyllaurous
    Article Snippet: This gene region has never been amplified before with dTTPs in contrast with the IGS primer set in our lab . .. For 1611F and 480R primers, amplifications were performed in a Mastercycler 5331 (Eppendorf, Hamburg, Germany) programmed as: an initial denaturing step of 95°C for 5 min; followed by 38 cycles of 95°C for 30 sec, 60°C for 50 sec, and 72°C for 1.5 min; and a final extension step of 72°C for 10 min. Thermocycler conditions for 50 s rRNA Bop-F and Bop-R primers (using UDG) were: an initial incubation of 37°C for 10 min, an initial denaturing/UDG deactivation step of 95°C for 10 min; followed by 38 cycles of 94°C for 30 sec, 60°C for 45 sec, and 72°C for 45 sec; and a final extension step of 72°C for 5 min. After PCR with 50 S rRNA primers, 0.1 U of uracil glycosylase inhibitor (New England Biolabs, Ipswich, MA, USA) was added to each PCR reaction.

    Filtration:

    Article Title: Timing Facilitated Site Transfer of an Enzyme on DNA
    Article Snippet: Unincorporated [γ33 P] ATP was then removed by gel filtration. .. At each time point 4 μl of the reaction mix was taken out and quenched with Uracil DNA Glycosylase Inhibitor (UGI) at a final concentration of 0.1 Units (New England Biolabs) or 50 nM final concentration of a highly potent duplex DNA inhibitor (2′-fluoro-2′-deoxyuridine paired with 4-methylindole in duplex DNA) , , both of which rapidly and efficiently quenched hUNG activity.

    Article Title: Electrostatic Properties of Complexes along a DNA Glycosylase Damage Search Pathway
    Article Snippet: Unincorporated [γ32 P] and [α32 P]ATP were removed by gel filtration. .. At each time point, an aliquot of the reaction mix was quenched with uracil DNA glycosylase inhibitor (UGI) at a final concentration of 0.1 U (New England Biolabs), which rapidly and efficiently quenched hUNG activity.

    Positive Control:

    Article Title: Manipulation of Plant Defense Responses by the Tomato Psyllid (Bactericerca cockerelli) and Its Associated Endosymbiont Candidatus Liberibacter Psyllaurous
    Article Snippet: For 1611F and 480R primers, amplifications were performed in a Mastercycler 5331 (Eppendorf, Hamburg, Germany) programmed as: an initial denaturing step of 95°C for 5 min; followed by 38 cycles of 95°C for 30 sec, 60°C for 50 sec, and 72°C for 1.5 min; and a final extension step of 72°C for 10 min. Thermocycler conditions for 50 s rRNA Bop-F and Bop-R primers (using UDG) were: an initial incubation of 37°C for 10 min, an initial denaturing/UDG deactivation step of 95°C for 10 min; followed by 38 cycles of 94°C for 30 sec, 60°C for 45 sec, and 72°C for 45 sec; and a final extension step of 72°C for 5 min. After PCR with 50 S rRNA primers, 0.1 U of uracil glycosylase inhibitor (New England Biolabs, Ipswich, MA, USA) was added to each PCR reaction. .. Extraction and PCR negative controls and a PCR positive control were included in each assay.

    Autoradiography:

    Article Title: BCR-ABL1 kinase inhibits uracil DNA glycosylase UNG2 to enhance oxidative DNA damage and stimulate genomic instability
    Article Snippet: Neutralizing antibodies (2μg) and 2U of uracil glycosylase inhibitor (UGI) (New England Biolabs, Inc., MA, USA) ( ) were added when indicated. .. Reactions were separated by electrophoresis in 18% polyacrylamide gel containing 8M urea and visualized by autoradiography.

    Blocking Assay:

    Article Title: Electrostatic Properties of Complexes along a DNA Glycosylase Damage Search Pathway
    Article Snippet: The 5′- and 3′-labeled strands were hybridized by heating to 95 °C in a heating block for 20 min and allowing the block to cool to room temperature. .. At each time point, an aliquot of the reaction mix was quenched with uracil DNA glycosylase inhibitor (UGI) at a final concentration of 0.1 U (New England Biolabs), which rapidly and efficiently quenched hUNG activity.

    Real-time Polymerase Chain Reaction:

    Article Title: A random mutation capture assay to detect genomic point mutations in mouse tissue
    Article Snippet: 95°C 10 min 95°C 30 s 62°C 60 s 74°C 10 s read plate. repeat steps (iii)–(vi) for 47 cycles. run a melting temperature curve to determine relative T(m) of products When the reaction is done, place qPCR plate immediately at −20°C upon completion as UDG will digest products even at 4°C. .. Digest 8 µl of the each PCR reaction with 0.25 µl Taqα 1 (NEB, 100 000 U/ml), 0.5 µl Uracil–DNA glycoslyase inhibitor (UGI, New England Biolabs catalog# M0281L), 0.2 µl NEB4 buffer and 1.0 µl double-deionized water added per reaction.

    Incubation:

    Article Title: Manipulation of Plant Defense Responses by the Tomato Psyllid (Bactericerca cockerelli) and Its Associated Endosymbiont Candidatus Liberibacter Psyllaurous
    Article Snippet: .. For 1611F and 480R primers, amplifications were performed in a Mastercycler 5331 (Eppendorf, Hamburg, Germany) programmed as: an initial denaturing step of 95°C for 5 min; followed by 38 cycles of 95°C for 30 sec, 60°C for 50 sec, and 72°C for 1.5 min; and a final extension step of 72°C for 10 min. Thermocycler conditions for 50 s rRNA Bop-F and Bop-R primers (using UDG) were: an initial incubation of 37°C for 10 min, an initial denaturing/UDG deactivation step of 95°C for 10 min; followed by 38 cycles of 94°C for 30 sec, 60°C for 45 sec, and 72°C for 45 sec; and a final extension step of 72°C for 5 min. After PCR with 50 S rRNA primers, 0.1 U of uracil glycosylase inhibitor (New England Biolabs, Ipswich, MA, USA) was added to each PCR reaction. ..

    Article Title: Timing Facilitated Site Transfer of an Enzyme on DNA
    Article Snippet: The reaction was then initiated by the addition of 2 μl of human UNG (hUNG) to a final concentration of 5–20 pM and incubated at 37 °C. .. At each time point 4 μl of the reaction mix was taken out and quenched with Uracil DNA Glycosylase Inhibitor (UGI) at a final concentration of 0.1 Units (New England Biolabs) or 50 nM final concentration of a highly potent duplex DNA inhibitor (2′-fluoro-2′-deoxyuridine paired with 4-methylindole in duplex DNA) , , both of which rapidly and efficiently quenched hUNG activity.

    Article Title: Epstein-Barr virus BORF2 inhibits cellular APOBEC3B to preserve viral genome integrity
    Article Snippet: Activity assays were carried out as described using a 10 minute incubation with a dU-containing oligo (RSH12955 5’-AAA AAA AAA UCG GGA AAA AAA-fluorescein-3’). .. 2 μL of UGI (4 units, NEB M0281S) were used to inhibit UDG.

    Article Title: 3CAPS – a structural AP–site analogue as a tool to investigate DNA base excision repair
    Article Snippet: .. To ensure comparable conditions, also the 3CAPS and THF substrates were incubated with UDG as above and all reactions were stopped by adding uracil glycosylase inhibitor UGI (NEB). ..

    Article Title: Ancient DNA from Chalcolithic Israel reveals the role of population mixture in cultural transformation
    Article Snippet: .. We added 30 μL of extract to the USER treatment mixture (1× Buffer Tango (ThermoFisher), 100 μM dNTP Mix (ThermoFisher), 1 mM ATP (ThermoFisher), 0.06 U/μL USER enzyme (NEB)), and incubated the reaction at 37 °C for 30 min. We inhibited the UDG enzyme by adding Uracil Glycosylase Inhibitor (0.12 U/μL; NEB) to the mix and incubating for a further 30 min at 37 °C. .. We then performed blunt end repair on the samples by adding T4 PNK (0.5 U/μL; ThermoFisher) and T4 Polymerase (90.1 U/μL; ThermoFisher) to the mixture and incubating for 15 min at 25 °C, followed by 5 min at 12 °C.

    Article Title: Electrostatic Properties of Complexes along a DNA Glycosylase Damage Search Pathway
    Article Snippet: Stock solutions of DNA containing either a 5′ or 3′ 32 P end label were generated by incubation of a DNA strand with [γ32 P]ATP (PerkinElmer) and T4 polynucleotide kinase (New England Biolabs) or [α32 P]ATP (PerkinElmer) and terminal transferase (New England Biolabs), respectively. .. At each time point, an aliquot of the reaction mix was quenched with uracil DNA glycosylase inhibitor (UGI) at a final concentration of 0.1 U (New England Biolabs), which rapidly and efficiently quenched hUNG activity.

    Article Title: BCR-ABL1 kinase inhibits uracil DNA glycosylase UNG2 to enhance oxidative DNA damage and stimulate genomic instability
    Article Snippet: Briefly, 10μg of the nuclear extract proteins were incubated in 37°C with 50fM of the [32 P] radiolabeled single-stranded or double-stranded substrate containing 5-OH-U or control in the reaction buffer (20mM Tris-HCl, pH 8.0, 10mM NaCl, 1mM EDTA, 1mM DTT, 0.5mg/ml BSA, 2mM ATP, 40mM creatine phosphate, 2U/μl creatine phosphokinase). .. Neutralizing antibodies (2μg) and 2U of uracil glycosylase inhibitor (UGI) (New England Biolabs, Inc., MA, USA) ( ) were added when indicated.

    Activity Assay:

    Article Title: Timing Facilitated Site Transfer of an Enzyme on DNA
    Article Snippet: .. At each time point 4 μl of the reaction mix was taken out and quenched with Uracil DNA Glycosylase Inhibitor (UGI) at a final concentration of 0.1 Units (New England Biolabs) or 50 nM final concentration of a highly potent duplex DNA inhibitor (2′-fluoro-2′-deoxyuridine paired with 4-methylindole in duplex DNA) , , both of which rapidly and efficiently quenched hUNG activity. ..

    Article Title: Epstein-Barr virus BORF2 inhibits cellular APOBEC3B to preserve viral genome integrity
    Article Snippet: Paragraph title: UDG activity assays. ... 2 μL of UGI (4 units, NEB M0281S) were used to inhibit UDG.

    Article Title: Human Base Excision Repair Creates a Bias Toward -1 Frameshift Mutations *
    Article Snippet: .. Inhibition of U-bulge glycosylase activity was achieved by addition of 0.02 units of uracil glycosylase inhibitor (New England Biolabs) to 0.4 mg/ml of HeLa WCE as described above. .. Inhibition of the glycosylase activity toward I-bulge and ϵA-bulge bulge DNA (10 n m ) was investigated by adding 100 n m unlabeled 25-mer competitor DNA that was either damaged (ϵA·T) or undamaged (A·T).

    Article Title: Electrostatic Properties of Complexes along a DNA Glycosylase Damage Search Pathway
    Article Snippet: .. At each time point, an aliquot of the reaction mix was quenched with uracil DNA glycosylase inhibitor (UGI) at a final concentration of 0.1 U (New England Biolabs), which rapidly and efficiently quenched hUNG activity. ..

    Mass Spectrometry:

    Article Title: 3CAPS – a structural AP–site analogue as a tool to investigate DNA base excision repair
    Article Snippet: Quality of synthesised oligonucleotides was evaluated by electrospray ionisation mass spectroscopy (ESI-MS). .. To ensure comparable conditions, also the 3CAPS and THF substrates were incubated with UDG as above and all reactions were stopped by adding uracil glycosylase inhibitor UGI (NEB).

    Modification:

    Article Title: Epstein-Barr virus BORF2 inhibits cellular APOBEC3B to preserve viral genome integrity
    Article Snippet: Cells were harvested after 30 hrs and were lysed in 300 μL modified HED buffer (20 mM HEPES, 15 mM EDTA, Roche cOmplete EDTA-free protease inhibitor cocktail tablet, pH 7.4) per 106 cells. .. 2 μL of UGI (4 units, NEB M0281S) were used to inhibit UDG.

    Transformation Assay:

    Article Title: A panel of colorimetric assays to measure enzymatic activity in the base excision DNA repair pathway
    Article Snippet: All other recombinant DNA repair enzymes and uracil glycosylase inhibitor (UGI) were purchased from New England Biolabs (Hitchin, UK). .. SV40 T-antigen transformed wild-type MEFs were a generous gift from Prof. Leona D. Samson (MIT, USA).

    High Performance Liquid Chromatography:

    Article Title: A panel of colorimetric assays to measure enzymatic activity in the base excision DNA repair pathway
    Article Snippet: Synthetic oligonucleotides were purchased from Integrated DNA Technologies (Leuven, Belgium) and Sigma-Aldrich, and were purified by high-performance liquid chromatography. .. All other recombinant DNA repair enzymes and uracil glycosylase inhibitor (UGI) were purchased from New England Biolabs (Hitchin, UK).

    Transfection:

    Article Title: Epstein-Barr virus BORF2 inhibits cellular APOBEC3B to preserve viral genome integrity
    Article Snippet: AGS or AGSΔUNG cells were transfected with 1 μg BKRF3 or vector control using 3 μL TransIT-LT1 in serum free RPMI. .. 2 μL of UGI (4 units, NEB M0281S) were used to inhibit UDG.

    Chromatography:

    Article Title: A panel of colorimetric assays to measure enzymatic activity in the base excision DNA repair pathway
    Article Snippet: Recombinant human POLB , purified by his-tag chromatography using an imidazole gradient elution , was a generous gift from Dr Jason Parsons (University of Liverpool, UK). .. All other recombinant DNA repair enzymes and uracil glycosylase inhibitor (UGI) were purchased from New England Biolabs (Hitchin, UK).

    Protease Inhibitor:

    Article Title: Epstein-Barr virus BORF2 inhibits cellular APOBEC3B to preserve viral genome integrity
    Article Snippet: Cells were harvested after 30 hrs and were lysed in 300 μL modified HED buffer (20 mM HEPES, 15 mM EDTA, Roche cOmplete EDTA-free protease inhibitor cocktail tablet, pH 7.4) per 106 cells. .. 2 μL of UGI (4 units, NEB M0281S) were used to inhibit UDG.

    Article Title: A panel of colorimetric assays to measure enzymatic activity in the base excision DNA repair pathway
    Article Snippet: Nunc® Immobiliser™ amino 96-well plates and Halt™ protease inhibitor cocktail were purchased from ThermoFisher Scientific (Hemel Hemstead, UK). .. All other recombinant DNA repair enzymes and uracil glycosylase inhibitor (UGI) were purchased from New England Biolabs (Hitchin, UK).

    Transferring:

    Article Title: Ancient DNA from Chalcolithic Israel reveals the role of population mixture in cultural transformation
    Article Snippet: We added 30 μL of extract to the USER treatment mixture (1× Buffer Tango (ThermoFisher), 100 μM dNTP Mix (ThermoFisher), 1 mM ATP (ThermoFisher), 0.06 U/μL USER enzyme (NEB)), and incubated the reaction at 37 °C for 30 min. We inhibited the UDG enzyme by adding Uracil Glycosylase Inhibitor (0.12 U/μL; NEB) to the mix and incubating for a further 30 min at 37 °C. .. We cleaned the reactions up using a MinElute PCR purification kit, adding five volumes of PB buffer to the reaction mixture, transferring to a collection tube, and spinning for 30 s at 3300× g .

    Generated:

    Article Title: 3CAPS – a structural AP–site analogue as a tool to investigate DNA base excision repair
    Article Snippet: To ensure comparable conditions, also the 3CAPS and THF substrates were incubated with UDG as above and all reactions were stopped by adding uracil glycosylase inhibitor UGI (NEB). .. The 5–hoU substrate was generated by annealing the 5′–fluorescein-labelled 5′–FAM-d(CGGAATTCGT CTAGGTTTGA GGT- 5–hoU -GACATC GGATCCATGG TACCTCGAGG GCAATGTCTA) and complementary oligonucleotides d(TAGACATTGC CCTCGAGGTA CCATGGATCC GATGTCGACC TCAAACCTAG ACGAATTCCG).

    Article Title: Human Base Excision Repair Creates a Bias Toward -1 Frameshift Mutations *
    Article Snippet: To confirm the identity of the 12-mer reaction product, we generated the authentic 12-mer from a single turnover reaction with 2 μ m AAG and 10 n m I·T DNA. .. Inhibition of U-bulge glycosylase activity was achieved by addition of 0.02 units of uracil glycosylase inhibitor (New England Biolabs) to 0.4 mg/ml of HeLa WCE as described above.

    Article Title: Electrostatic Properties of Complexes along a DNA Glycosylase Damage Search Pathway
    Article Snippet: Stock solutions of DNA containing either a 5′ or 3′ 32 P end label were generated by incubation of a DNA strand with [γ32 P]ATP (PerkinElmer) and T4 polynucleotide kinase (New England Biolabs) or [α32 P]ATP (PerkinElmer) and terminal transferase (New England Biolabs), respectively. .. At each time point, an aliquot of the reaction mix was quenched with uracil DNA glycosylase inhibitor (UGI) at a final concentration of 0.1 U (New England Biolabs), which rapidly and efficiently quenched hUNG activity.

    Inhibition:

    Article Title: Human Base Excision Repair Creates a Bias Toward -1 Frameshift Mutations *
    Article Snippet: .. Inhibition of U-bulge glycosylase activity was achieved by addition of 0.02 units of uracil glycosylase inhibitor (New England Biolabs) to 0.4 mg/ml of HeLa WCE as described above. .. Inhibition of the glycosylase activity toward I-bulge and ϵA-bulge bulge DNA (10 n m ) was investigated by adding 100 n m unlabeled 25-mer competitor DNA that was either damaged (ϵA·T) or undamaged (A·T).

    Article Title: Electrostatic Properties of Complexes along a DNA Glycosylase Damage Search Pathway
    Article Snippet: We attribute this affect to competitive inhibition by phosphate dianion. .. At each time point, an aliquot of the reaction mix was quenched with uracil DNA glycosylase inhibitor (UGI) at a final concentration of 0.1 U (New England Biolabs), which rapidly and efficiently quenched hUNG activity.

    Sequencing:

    Article Title: Timing Facilitated Site Transfer of an Enzyme on DNA
    Article Snippet: At each time point 4 μl of the reaction mix was taken out and quenched with Uracil DNA Glycosylase Inhibitor (UGI) at a final concentration of 0.1 Units (New England Biolabs) or 50 nM final concentration of a highly potent duplex DNA inhibitor (2′-fluoro-2′-deoxyuridine paired with 4-methylindole in duplex DNA) , , both of which rapidly and efficiently quenched hUNG activity. .. The fragments were then separated on a 10% non-denaturing polyacrylamide gel (19:1 bis-acrylamide ratio, 0.5 mm thickness, run at 20 watts with 1X TBE running buffer in a model S2 sequencing gel apparatus).

    Recombinant:

    Article Title: Characterisation of the substrate specificity of homogeneous vaccinia virus uracil-DNA glycosylase
    Article Snippet: T4 polynucleotide kinase and uracil-DNA glycosylase inhibitor (UGI) were purchased from New England BioLabs (Saint Quentin Yvelines, France). .. The hUNG recombinant protein lacking the N-terminal 84 amino acids (27 kDa, 230 amino acids) was over-expressed in E.coli MS1021 cells and purified as described ( ).

    Article Title: A panel of colorimetric assays to measure enzymatic activity in the base excision DNA repair pathway
    Article Snippet: .. All other recombinant DNA repair enzymes and uracil glycosylase inhibitor (UGI) were purchased from New England Biolabs (Hitchin, UK). .. L189 mammalian DNA ligase inhibitor was purchased from Bio-Techne (Abingdon, UK).

    Molecular Weight:

    Article Title: A random mutation capture assay to detect genomic point mutations in mouse tissue
    Article Snippet: In order to distinguish between wild-type and mutant products, perform a post-PCR TaqI digestion and agarose gel electrophoresis analysis, using wild-type PCR products with and without TaqI digestion as molecular weight markers. .. Digest 8 µl of the each PCR reaction with 0.25 µl Taqα 1 (NEB, 100 000 U/ml), 0.5 µl Uracil–DNA glycoslyase inhibitor (UGI, New England Biolabs catalog# M0281L), 0.2 µl NEB4 buffer and 1.0 µl double-deionized water added per reaction.

    DNA Extraction:

    Article Title: Ancient DNA from Chalcolithic Israel reveals the role of population mixture in cultural transformation
    Article Snippet: For reattempts of one of the samples, we washed the powder with 1 mL 0.5% bleach (incubating for 15 min), followed by three washes with 1 mL water (incubating 3 min), prior to DNA extraction as described in Korlević et al. (see Supplementary Data ), and prepared libraries using partial UDG treatment (the library protocols varied slightly over the course of data generation, see Supplementary Data ). .. We added 30 μL of extract to the USER treatment mixture (1× Buffer Tango (ThermoFisher), 100 μM dNTP Mix (ThermoFisher), 1 mM ATP (ThermoFisher), 0.06 U/μL USER enzyme (NEB)), and incubated the reaction at 37 °C for 30 min. We inhibited the UDG enzyme by adding Uracil Glycosylase Inhibitor (0.12 U/μL; NEB) to the mix and incubating for a further 30 min at 37 °C.

    Fluorescence:

    Article Title: Epstein-Barr virus BORF2 inhibits cellular APOBEC3B to preserve viral genome integrity
    Article Snippet: 2 μL of UGI (4 units, NEB M0281S) were used to inhibit UDG. .. Separated DNA fragments were visualized on a Typhoon FLA-7000 scanner on fluorescence mode.

    Article Title: Human Base Excision Repair Creates a Bias Toward -1 Frameshift Mutations *
    Article Snippet: The fraction of glycosylase product was determined by dividing the fluorescence intensity of the 12-mer product band by the sum of the 12-mer product and 25-mer substrate bands. .. Inhibition of U-bulge glycosylase activity was achieved by addition of 0.02 units of uracil glycosylase inhibitor (New England Biolabs) to 0.4 mg/ml of HeLa WCE as described above.

    Mutagenesis:

    Article Title: A random mutation capture assay to detect genomic point mutations in mouse tissue
    Article Snippet: In order to distinguish between wild-type and mutant products, perform a post-PCR TaqI digestion and agarose gel electrophoresis analysis, using wild-type PCR products with and without TaqI digestion as molecular weight markers. .. Digest 8 µl of the each PCR reaction with 0.25 µl Taqα 1 (NEB, 100 000 U/ml), 0.5 µl Uracil–DNA glycoslyase inhibitor (UGI, New England Biolabs catalog# M0281L), 0.2 µl NEB4 buffer and 1.0 µl double-deionized water added per reaction.

    Size-exclusion Chromatography:

    Article Title: Manipulation of Plant Defense Responses by the Tomato Psyllid (Bactericerca cockerelli) and Its Associated Endosymbiont Candidatus Liberibacter Psyllaurous
    Article Snippet: .. For 1611F and 480R primers, amplifications were performed in a Mastercycler 5331 (Eppendorf, Hamburg, Germany) programmed as: an initial denaturing step of 95°C for 5 min; followed by 38 cycles of 95°C for 30 sec, 60°C for 50 sec, and 72°C for 1.5 min; and a final extension step of 72°C for 10 min. Thermocycler conditions for 50 s rRNA Bop-F and Bop-R primers (using UDG) were: an initial incubation of 37°C for 10 min, an initial denaturing/UDG deactivation step of 95°C for 10 min; followed by 38 cycles of 94°C for 30 sec, 60°C for 45 sec, and 72°C for 45 sec; and a final extension step of 72°C for 5 min. After PCR with 50 S rRNA primers, 0.1 U of uracil glycosylase inhibitor (New England Biolabs, Ipswich, MA, USA) was added to each PCR reaction. ..

    Electrophoretic Mobility Shift Assay:

    Article Title: 3CAPS – a structural AP–site analogue as a tool to investigate DNA base excision repair
    Article Snippet: For electrophoretic mobility shift assay (EMSA), incision and reconstituted BER assays, 5′–fluorescein-labelled 5′–FAM-d(CTAGGTTTGA GGT X GACATC GGATCCATGG) and unlabelled 5′–d(CCTCGAGGTA CCATGGATCC GATGTCGACC TCAAACCTAG ACGAATTCCG) oligonucleotides were annealed (X represents either a 3CAPS, tetrahydrofuran or deoxyuridine) to produce 3CAPS, THF and G•U mismatched heteroduplex substrates. .. To ensure comparable conditions, also the 3CAPS and THF substrates were incubated with UDG as above and all reactions were stopped by adding uracil glycosylase inhibitor UGI (NEB).

    Purification:

    Article Title: Ancient DNA from Chalcolithic Israel reveals the role of population mixture in cultural transformation
    Article Snippet: We added 30 μL of extract to the USER treatment mixture (1× Buffer Tango (ThermoFisher), 100 μM dNTP Mix (ThermoFisher), 1 mM ATP (ThermoFisher), 0.06 U/μL USER enzyme (NEB)), and incubated the reaction at 37 °C for 30 min. We inhibited the UDG enzyme by adding Uracil Glycosylase Inhibitor (0.12 U/μL; NEB) to the mix and incubating for a further 30 min at 37 °C. .. We cleaned the reactions up using a MinElute PCR purification kit, adding five volumes of PB buffer to the reaction mixture, transferring to a collection tube, and spinning for 30 s at 3300× g .

    Article Title: Characterisation of the substrate specificity of homogeneous vaccinia virus uracil-DNA glycosylase
    Article Snippet: T4 polynucleotide kinase and uracil-DNA glycosylase inhibitor (UGI) were purchased from New England BioLabs (Saint Quentin Yvelines, France). .. Purification of the E.coli Fpg protein was performed as described ( ).

    Article Title: A panel of colorimetric assays to measure enzymatic activity in the base excision DNA repair pathway
    Article Snippet: Recombinant human POLB , purified by his-tag chromatography using an imidazole gradient elution , was a generous gift from Dr Jason Parsons (University of Liverpool, UK). .. All other recombinant DNA repair enzymes and uracil glycosylase inhibitor (UGI) were purchased from New England Biolabs (Hitchin, UK).

    Polymerase Chain Reaction:

    Article Title: Manipulation of Plant Defense Responses by the Tomato Psyllid (Bactericerca cockerelli) and Its Associated Endosymbiont Candidatus Liberibacter Psyllaurous
    Article Snippet: .. For 1611F and 480R primers, amplifications were performed in a Mastercycler 5331 (Eppendorf, Hamburg, Germany) programmed as: an initial denaturing step of 95°C for 5 min; followed by 38 cycles of 95°C for 30 sec, 60°C for 50 sec, and 72°C for 1.5 min; and a final extension step of 72°C for 10 min. Thermocycler conditions for 50 s rRNA Bop-F and Bop-R primers (using UDG) were: an initial incubation of 37°C for 10 min, an initial denaturing/UDG deactivation step of 95°C for 10 min; followed by 38 cycles of 94°C for 30 sec, 60°C for 45 sec, and 72°C for 45 sec; and a final extension step of 72°C for 5 min. After PCR with 50 S rRNA primers, 0.1 U of uracil glycosylase inhibitor (New England Biolabs, Ipswich, MA, USA) was added to each PCR reaction. ..

    Article Title: Ancient DNA from Chalcolithic Israel reveals the role of population mixture in cultural transformation
    Article Snippet: We added 30 μL of extract to the USER treatment mixture (1× Buffer Tango (ThermoFisher), 100 μM dNTP Mix (ThermoFisher), 1 mM ATP (ThermoFisher), 0.06 U/μL USER enzyme (NEB)), and incubated the reaction at 37 °C for 30 min. We inhibited the UDG enzyme by adding Uracil Glycosylase Inhibitor (0.12 U/μL; NEB) to the mix and incubating for a further 30 min at 37 °C. .. We cleaned the reactions up using a MinElute PCR purification kit, adding five volumes of PB buffer to the reaction mixture, transferring to a collection tube, and spinning for 30 s at 3300× g .

    Article Title: A random mutation capture assay to detect genomic point mutations in mouse tissue
    Article Snippet: .. Digest 8 µl of the each PCR reaction with 0.25 µl Taqα 1 (NEB, 100 000 U/ml), 0.5 µl Uracil–DNA glycoslyase inhibitor (UGI, New England Biolabs catalog# M0281L), 0.2 µl NEB4 buffer and 1.0 µl double-deionized water added per reaction. .. Incubate for 10 min at 65°C, 1 min at 95°C and then 1 min at 25°C (digestions are easiest to do in a PCR machine).

    Polyacrylamide Gel Electrophoresis:

    Article Title: Epstein-Barr virus BORF2 inhibits cellular APOBEC3B to preserve viral genome integrity
    Article Snippet: 2 μL of UGI (4 units, NEB M0281S) were used to inhibit UDG. .. Products were separated by a 20% TBE-Urea PAGE.

    Article Title: 3CAPS – a structural AP–site analogue as a tool to investigate DNA base excision repair
    Article Snippet: All other oligonucleotides (PAGE-purified grade) were purchased from Microsynth AG (Balgach, Switzerland) or from Sigma-Aldrich (St. Louis, MO, USA). .. To ensure comparable conditions, also the 3CAPS and THF substrates were incubated with UDG as above and all reactions were stopped by adding uracil glycosylase inhibitor UGI (NEB).

    Staining:

    Article Title: Manipulation of Plant Defense Responses by the Tomato Psyllid (Bactericerca cockerelli) and Its Associated Endosymbiont Candidatus Liberibacter Psyllaurous
    Article Snippet: For 1611F and 480R primers, amplifications were performed in a Mastercycler 5331 (Eppendorf, Hamburg, Germany) programmed as: an initial denaturing step of 95°C for 5 min; followed by 38 cycles of 95°C for 30 sec, 60°C for 50 sec, and 72°C for 1.5 min; and a final extension step of 72°C for 10 min. Thermocycler conditions for 50 s rRNA Bop-F and Bop-R primers (using UDG) were: an initial incubation of 37°C for 10 min, an initial denaturing/UDG deactivation step of 95°C for 10 min; followed by 38 cycles of 94°C for 30 sec, 60°C for 45 sec, and 72°C for 45 sec; and a final extension step of 72°C for 5 min. After PCR with 50 S rRNA primers, 0.1 U of uracil glycosylase inhibitor (New England Biolabs, Ipswich, MA, USA) was added to each PCR reaction. .. Amplified DNA was visualized after electrophoresis on a 1% agarose gel stained with ethidium bromide (run at ∼4.9 V/cm for at least 1 hr).

    Plasmid Preparation:

    Article Title: Epstein-Barr virus BORF2 inhibits cellular APOBEC3B to preserve viral genome integrity
    Article Snippet: AGS or AGSΔUNG cells were transfected with 1 μg BKRF3 or vector control using 3 μL TransIT-LT1 in serum free RPMI. .. 2 μL of UGI (4 units, NEB M0281S) were used to inhibit UDG.

    Article Title: Characterisation of the substrate specificity of homogeneous vaccinia virus uracil-DNA glycosylase
    Article Snippet: T4 polynucleotide kinase and uracil-DNA glycosylase inhibitor (UGI) were purchased from New England BioLabs (Saint Quentin Yvelines, France). .. The NdeI–HindIII fragment containing the full-length UNG2 cDNA ( ) was subcloned into the pET28a vector (Novagen).

    Software:

    Article Title: BCR-ABL1 kinase inhibits uracil DNA glycosylase UNG2 to enhance oxidative DNA damage and stimulate genomic instability
    Article Snippet: Neutralizing antibodies (2μg) and 2U of uracil glycosylase inhibitor (UGI) (New England Biolabs, Inc., MA, USA) ( ) were added when indicated. .. The products were quantified using Quantity One software (Bio-Rad, Hercules, CA, USA).

    SYBR Green Assay:

    Article Title: Highly Sensitive Detection of Uracil-DNA Glycosylase Activity Based on Self-Initiating Multiple Rolling Circle Amplification
    Article Snippet: 4.1 Materials and Reagents Uracil-DNA glycosylase (UDG), human alkyladenine glycosylase (hAAG), uracil glycosylase inhibitor (UGI), T4 DNA ligase, exonuclease I (Exo I), exonuclease III (Exo III), endonuclease IV (Endo IV), and 10× NEBuffer 2 (500 mM NaCl, 100 mM Tris-HCl, 100 mM MgCl2 , 10 mM dithiothreitol) were purchased from New England Biolabs (Ipswich, MA, USA). .. 10× SYBR Green I was purchased from Zeesan (Xiamen, China).

    Agarose Gel Electrophoresis:

    Article Title: Manipulation of Plant Defense Responses by the Tomato Psyllid (Bactericerca cockerelli) and Its Associated Endosymbiont Candidatus Liberibacter Psyllaurous
    Article Snippet: For 1611F and 480R primers, amplifications were performed in a Mastercycler 5331 (Eppendorf, Hamburg, Germany) programmed as: an initial denaturing step of 95°C for 5 min; followed by 38 cycles of 95°C for 30 sec, 60°C for 50 sec, and 72°C for 1.5 min; and a final extension step of 72°C for 10 min. Thermocycler conditions for 50 s rRNA Bop-F and Bop-R primers (using UDG) were: an initial incubation of 37°C for 10 min, an initial denaturing/UDG deactivation step of 95°C for 10 min; followed by 38 cycles of 94°C for 30 sec, 60°C for 45 sec, and 72°C for 45 sec; and a final extension step of 72°C for 5 min. After PCR with 50 S rRNA primers, 0.1 U of uracil glycosylase inhibitor (New England Biolabs, Ipswich, MA, USA) was added to each PCR reaction. .. Amplified DNA was visualized after electrophoresis on a 1% agarose gel stained with ethidium bromide (run at ∼4.9 V/cm for at least 1 hr).

    Article Title: A random mutation capture assay to detect genomic point mutations in mouse tissue
    Article Snippet: In order to distinguish between wild-type and mutant products, perform a post-PCR TaqI digestion and agarose gel electrophoresis analysis, using wild-type PCR products with and without TaqI digestion as molecular weight markers. .. Digest 8 µl of the each PCR reaction with 0.25 µl Taqα 1 (NEB, 100 000 U/ml), 0.5 µl Uracil–DNA glycoslyase inhibitor (UGI, New England Biolabs catalog# M0281L), 0.2 µl NEB4 buffer and 1.0 µl double-deionized water added per reaction.

    Electrophoresis:

    Article Title: Manipulation of Plant Defense Responses by the Tomato Psyllid (Bactericerca cockerelli) and Its Associated Endosymbiont Candidatus Liberibacter Psyllaurous
    Article Snippet: For 1611F and 480R primers, amplifications were performed in a Mastercycler 5331 (Eppendorf, Hamburg, Germany) programmed as: an initial denaturing step of 95°C for 5 min; followed by 38 cycles of 95°C for 30 sec, 60°C for 50 sec, and 72°C for 1.5 min; and a final extension step of 72°C for 10 min. Thermocycler conditions for 50 s rRNA Bop-F and Bop-R primers (using UDG) were: an initial incubation of 37°C for 10 min, an initial denaturing/UDG deactivation step of 95°C for 10 min; followed by 38 cycles of 94°C for 30 sec, 60°C for 45 sec, and 72°C for 45 sec; and a final extension step of 72°C for 5 min. After PCR with 50 S rRNA primers, 0.1 U of uracil glycosylase inhibitor (New England Biolabs, Ipswich, MA, USA) was added to each PCR reaction. .. Amplified DNA was visualized after electrophoresis on a 1% agarose gel stained with ethidium bromide (run at ∼4.9 V/cm for at least 1 hr).

    Article Title: BCR-ABL1 kinase inhibits uracil DNA glycosylase UNG2 to enhance oxidative DNA damage and stimulate genomic instability
    Article Snippet: Neutralizing antibodies (2μg) and 2U of uracil glycosylase inhibitor (UGI) (New England Biolabs, Inc., MA, USA) ( ) were added when indicated. .. Reactions were separated by electrophoresis in 18% polyacrylamide gel containing 8M urea and visualized by autoradiography.

    Concentration Assay:

    Article Title: Manipulation of Plant Defense Responses by the Tomato Psyllid (Bactericerca cockerelli) and Its Associated Endosymbiont Candidatus Liberibacter Psyllaurous
    Article Snippet: For both primer sets, PCR was performed in 25-µL reactions containing 1 µL of DNA template (concentration not determined), 0.2 mM each of dATP, dCTP, and dGTP, 0.4 mM of dTTP mix, 0.2 µM each primer, 1× PCR MgCl2 -free buffer (Roche, Indianapolis, IN, USA), 5 mM MgCl2 , 1 U polymerase (Roche, Indianapolis, IN, USA), and 1 U UDG when 50 s rRNA primers were used. .. For 1611F and 480R primers, amplifications were performed in a Mastercycler 5331 (Eppendorf, Hamburg, Germany) programmed as: an initial denaturing step of 95°C for 5 min; followed by 38 cycles of 95°C for 30 sec, 60°C for 50 sec, and 72°C for 1.5 min; and a final extension step of 72°C for 10 min. Thermocycler conditions for 50 s rRNA Bop-F and Bop-R primers (using UDG) were: an initial incubation of 37°C for 10 min, an initial denaturing/UDG deactivation step of 95°C for 10 min; followed by 38 cycles of 94°C for 30 sec, 60°C for 45 sec, and 72°C for 45 sec; and a final extension step of 72°C for 5 min. After PCR with 50 S rRNA primers, 0.1 U of uracil glycosylase inhibitor (New England Biolabs, Ipswich, MA, USA) was added to each PCR reaction.

    Article Title: Timing Facilitated Site Transfer of an Enzyme on DNA
    Article Snippet: .. At each time point 4 μl of the reaction mix was taken out and quenched with Uracil DNA Glycosylase Inhibitor (UGI) at a final concentration of 0.1 Units (New England Biolabs) or 50 nM final concentration of a highly potent duplex DNA inhibitor (2′-fluoro-2′-deoxyuridine paired with 4-methylindole in duplex DNA) , , both of which rapidly and efficiently quenched hUNG activity. ..

    Article Title: Human Base Excision Repair Creates a Bias Toward -1 Frameshift Mutations *
    Article Snippet: The fraction was converted into concentration by multiplying by the total amount of DNA substrate present. .. Inhibition of U-bulge glycosylase activity was achieved by addition of 0.02 units of uracil glycosylase inhibitor (New England Biolabs) to 0.4 mg/ml of HeLa WCE as described above.

    Article Title: Electrostatic Properties of Complexes along a DNA Glycosylase Damage Search Pathway
    Article Snippet: .. At each time point, an aliquot of the reaction mix was quenched with uracil DNA glycosylase inhibitor (UGI) at a final concentration of 0.1 U (New England Biolabs), which rapidly and efficiently quenched hUNG activity. ..

    Lysis:

    Article Title: Human Base Excision Repair Creates a Bias Toward -1 Frameshift Mutations *
    Article Snippet: Whole cell extracts (WCE) were obtained from Active Motif and were stored at −80 °C in lysis buffer (20 m m NaHEPES, pH 7.5, 350 m m NaCl, 20% glycerol, 1% Igepal-CA630, 1 m m MgCl2 , 0.5 m m EDTA, 0.1 m m EGTA) until immediately before use. .. Inhibition of U-bulge glycosylase activity was achieved by addition of 0.02 units of uracil glycosylase inhibitor (New England Biolabs) to 0.4 mg/ml of HeLa WCE as described above.

    other:

    Article Title: Arabidopsis Uracil DNA Glycosylase (UNG) Is Required for Base Excision Repair of Uracil and Increases Plant Sensitivity to 5-Fluorouracil *
    Article Snippet: E. coli Ung, and uracil DNA glycosylase inhibitor (Ugi) were obtained from New England BioLabs.

    Article Title: DIFFERENTIAL ROLE OF BASE EXCISION REPAIR PROTEINS IN MEDIATING CISPLATIN CYTOTOXICITY
    Article Snippet: Uracil DNA glycosylase from E. coli (UDG) 2,000 U/ml, human apurinic/apyridimic endonuclease (APE1) 10,000 U/ml, and Uracil Glycosylase inhibitor (UGI) 2,000 U/ml were from New England Biolabs.

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    New England Biolabs uracil glycosylase inhibitor ugi
    Cisplatin cytotoxicity and effect on <t>glycosylase</t> activity (A) Colony survival assay in MDA-MB-231 cells following UNG and SMUG1 knockdown: shControl (open circles), shUNG (closed triangles), shSMUG1 (closed circles) and shUNG + shSMUG1 (open squares). Results are represented as mean ± SE from 3 independent experiments. Cells were transfected with shRNA directed against UNG and SMUG1. (B) Colony survival assay in MDA-MB-231 cells following MBD4 knockdown with shControl (open circles) and shMBD4 (closed triangles). shRNA transfected cells were treated with increasing doses of cisplatin and cytotoxicity. Results are represented as mean ± SE from 3 independent experiments. (C) In vitro glycosylase assay, DNA (5nM) was incubated with either pure enzyme or HeLa extract. Lane 1, undamaged DNA alone.; lane 2, undamaged DNA treated with UDG and APE1 to generate a 19 mer product; lane 3, undamaged DNA substrate treated with UDG, APE1 and 1 unit of <t>UGI;</t> lane 4, undamaged DNA incubated with HeLa extract; lane 5 reactions in which HeLa extract was preincubated with 1 unit of UGI before adding the undamaged DNA substrate. Lanes 6–10 follow the same set up as lanes 1–5, but with ICL DNA substrate. Both undamaged and ICL substrates contain a central uracil and a 3′ Cy3 label. M is a 21-nt marker.
    Uracil Glycosylase Inhibitor Ugi, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Cisplatin cytotoxicity and effect on glycosylase activity (A) Colony survival assay in MDA-MB-231 cells following UNG and SMUG1 knockdown: shControl (open circles), shUNG (closed triangles), shSMUG1 (closed circles) and shUNG + shSMUG1 (open squares). Results are represented as mean ± SE from 3 independent experiments. Cells were transfected with shRNA directed against UNG and SMUG1. (B) Colony survival assay in MDA-MB-231 cells following MBD4 knockdown with shControl (open circles) and shMBD4 (closed triangles). shRNA transfected cells were treated with increasing doses of cisplatin and cytotoxicity. Results are represented as mean ± SE from 3 independent experiments. (C) In vitro glycosylase assay, DNA (5nM) was incubated with either pure enzyme or HeLa extract. Lane 1, undamaged DNA alone.; lane 2, undamaged DNA treated with UDG and APE1 to generate a 19 mer product; lane 3, undamaged DNA substrate treated with UDG, APE1 and 1 unit of UGI; lane 4, undamaged DNA incubated with HeLa extract; lane 5 reactions in which HeLa extract was preincubated with 1 unit of UGI before adding the undamaged DNA substrate. Lanes 6–10 follow the same set up as lanes 1–5, but with ICL DNA substrate. Both undamaged and ICL substrates contain a central uracil and a 3′ Cy3 label. M is a 21-nt marker.

    Journal: DNA repair

    Article Title: DIFFERENTIAL ROLE OF BASE EXCISION REPAIR PROTEINS IN MEDIATING CISPLATIN CYTOTOXICITY

    doi: 10.1016/j.dnarep.2017.01.002

    Figure Lengend Snippet: Cisplatin cytotoxicity and effect on glycosylase activity (A) Colony survival assay in MDA-MB-231 cells following UNG and SMUG1 knockdown: shControl (open circles), shUNG (closed triangles), shSMUG1 (closed circles) and shUNG + shSMUG1 (open squares). Results are represented as mean ± SE from 3 independent experiments. Cells were transfected with shRNA directed against UNG and SMUG1. (B) Colony survival assay in MDA-MB-231 cells following MBD4 knockdown with shControl (open circles) and shMBD4 (closed triangles). shRNA transfected cells were treated with increasing doses of cisplatin and cytotoxicity. Results are represented as mean ± SE from 3 independent experiments. (C) In vitro glycosylase assay, DNA (5nM) was incubated with either pure enzyme or HeLa extract. Lane 1, undamaged DNA alone.; lane 2, undamaged DNA treated with UDG and APE1 to generate a 19 mer product; lane 3, undamaged DNA substrate treated with UDG, APE1 and 1 unit of UGI; lane 4, undamaged DNA incubated with HeLa extract; lane 5 reactions in which HeLa extract was preincubated with 1 unit of UGI before adding the undamaged DNA substrate. Lanes 6–10 follow the same set up as lanes 1–5, but with ICL DNA substrate. Both undamaged and ICL substrates contain a central uracil and a 3′ Cy3 label. M is a 21-nt marker.

    Article Snippet: Uracil DNA glycosylase from E. coli (UDG) 2,000 U/ml, human apurinic/apyridimic endonuclease (APE1) 10,000 U/ml, and Uracil Glycosylase inhibitor (UGI) 2,000 U/ml were from New England Biolabs.

    Techniques: Activity Assay, Clonogenic Cell Survival Assay, Multiple Displacement Amplification, Transfection, shRNA, In Vitro, Incubation, Marker