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Promega dna t4 ligase
Dna T4 Ligase, supplied by Promega, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/result/dna t4 ligase/product/Promega
Average 93 stars, based on 1 article reviews
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dna t4 ligase - by Bioz Stars, 2020-05
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Amplification:

Article Title: A Plant-Specific TGS1 Homolog Influences Gametophyte Development in Sexual Tetraploid Paspalum notatum Ovules
Article Snippet: .. The F1 fragment and the digested pAct1-gfbsd2 plasmid were then ligated with DNA T4 ligase (PROMEGA) and the resulting construct, pAct1-F1as, was amplified by transforming DH5α competent cells. .. Finally, pAct1-F1as was sequenced at Macrogen Inc. (Seoul, Korea) to check position, sense and integrity of the F1 fragment.

Plasmid Preparation:

Article Title: A Plant-Specific TGS1 Homolog Influences Gametophyte Development in Sexual Tetraploid Paspalum notatum Ovules
Article Snippet: .. The F1 fragment and the digested pAct1-gfbsd2 plasmid were then ligated with DNA T4 ligase (PROMEGA) and the resulting construct, pAct1-F1as, was amplified by transforming DH5α competent cells. .. Finally, pAct1-F1as was sequenced at Macrogen Inc. (Seoul, Korea) to check position, sense and integrity of the F1 fragment.

Construct:

Article Title: A Plant-Specific TGS1 Homolog Influences Gametophyte Development in Sexual Tetraploid Paspalum notatum Ovules
Article Snippet: .. The F1 fragment and the digested pAct1-gfbsd2 plasmid were then ligated with DNA T4 ligase (PROMEGA) and the resulting construct, pAct1-F1as, was amplified by transforming DH5α competent cells. .. Finally, pAct1-F1as was sequenced at Macrogen Inc. (Seoul, Korea) to check position, sense and integrity of the F1 fragment.

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    Promega t4 dna ligase
    Ligation in the presence of DNA-PK requires ATP hydrolysis and an active DNA-PK CS kinase. ( A ) An overall labeled DNA substrate with cohesive ends was incubated with <t>T4</t> DNA ligase, either in the absence (lanes 1–3) or the presence (lanes 4 and 5) of DNA-PK. ATP or AMP-PNP was present as indicated. Ligation products were separated by agarose gel electrophoresis. The nature of the ligation products, identified as intra- or inter-molecular ligation products, was confirmed by exonuclease V digestion. Note that intra-molecular ligation products can be either ligated on one strand (open circular form) or on both strands (covalently closed circular form). ( B ) An overall labeled DNA substrate with cohesive ends was incubated with E.coli DNA ligase, either in the absence (lanes 1–4) or the presence (lanes 5 and 6) of DNA-PK. ATP and/or NAD + were present as indicated. ( C ) An overall labeled DNA substrate with cohesive ends was incubated with T4 DNA ligase, either in the absence (lanes 1 and 2) or the presence (lanes 3 and 4) of DNA-PK. All reaction mixtures contained ATP. The DNA-PK CS kinase inhibitor wortmannin was added in lane 4. Total levels of ligation products in all lanes were decreased in comparison with (A), due to the presence of DMSO in the reaction mixtures. ( D ) Wortmannin inhibits autophosphorylation of DNA-PK CS . Incorporation of radiolabeled phosphate into DNA-PK CS was determined in the absence and presence of 1 or 10 µM wortmannin. Even 1 µM wortmannin completely inhibits DNA-PK CS autophosphorylation.
    T4 Dna Ligase, supplied by Promega, used in various techniques. Bioz Stars score: 99/100, based on 578 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/t4 dna ligase/product/Promega
    Average 99 stars, based on 578 article reviews
    Price from $9.99 to $1999.99
    t4 dna ligase - by Bioz Stars, 2020-05
    99/100 stars
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    Ligation in the presence of DNA-PK requires ATP hydrolysis and an active DNA-PK CS kinase. ( A ) An overall labeled DNA substrate with cohesive ends was incubated with T4 DNA ligase, either in the absence (lanes 1–3) or the presence (lanes 4 and 5) of DNA-PK. ATP or AMP-PNP was present as indicated. Ligation products were separated by agarose gel electrophoresis. The nature of the ligation products, identified as intra- or inter-molecular ligation products, was confirmed by exonuclease V digestion. Note that intra-molecular ligation products can be either ligated on one strand (open circular form) or on both strands (covalently closed circular form). ( B ) An overall labeled DNA substrate with cohesive ends was incubated with E.coli DNA ligase, either in the absence (lanes 1–4) or the presence (lanes 5 and 6) of DNA-PK. ATP and/or NAD + were present as indicated. ( C ) An overall labeled DNA substrate with cohesive ends was incubated with T4 DNA ligase, either in the absence (lanes 1 and 2) or the presence (lanes 3 and 4) of DNA-PK. All reaction mixtures contained ATP. The DNA-PK CS kinase inhibitor wortmannin was added in lane 4. Total levels of ligation products in all lanes were decreased in comparison with (A), due to the presence of DMSO in the reaction mixtures. ( D ) Wortmannin inhibits autophosphorylation of DNA-PK CS . Incorporation of radiolabeled phosphate into DNA-PK CS was determined in the absence and presence of 1 or 10 µM wortmannin. Even 1 µM wortmannin completely inhibits DNA-PK CS autophosphorylation.

    Journal: Nucleic Acids Research

    Article Title: The role of DNA dependent protein kinase in synapsis of DNA ends

    doi: 10.1093/nar/gkg889

    Figure Lengend Snippet: Ligation in the presence of DNA-PK requires ATP hydrolysis and an active DNA-PK CS kinase. ( A ) An overall labeled DNA substrate with cohesive ends was incubated with T4 DNA ligase, either in the absence (lanes 1–3) or the presence (lanes 4 and 5) of DNA-PK. ATP or AMP-PNP was present as indicated. Ligation products were separated by agarose gel electrophoresis. The nature of the ligation products, identified as intra- or inter-molecular ligation products, was confirmed by exonuclease V digestion. Note that intra-molecular ligation products can be either ligated on one strand (open circular form) or on both strands (covalently closed circular form). ( B ) An overall labeled DNA substrate with cohesive ends was incubated with E.coli DNA ligase, either in the absence (lanes 1–4) or the presence (lanes 5 and 6) of DNA-PK. ATP and/or NAD + were present as indicated. ( C ) An overall labeled DNA substrate with cohesive ends was incubated with T4 DNA ligase, either in the absence (lanes 1 and 2) or the presence (lanes 3 and 4) of DNA-PK. All reaction mixtures contained ATP. The DNA-PK CS kinase inhibitor wortmannin was added in lane 4. Total levels of ligation products in all lanes were decreased in comparison with (A), due to the presence of DMSO in the reaction mixtures. ( D ) Wortmannin inhibits autophosphorylation of DNA-PK CS . Incorporation of radiolabeled phosphate into DNA-PK CS was determined in the absence and presence of 1 or 10 µM wortmannin. Even 1 µM wortmannin completely inhibits DNA-PK CS autophosphorylation.

    Article Snippet: First, we used AMP-PNP, an ATP analog that supports activity of T4 DNA ligase, but cannot function as a cofactor for DNA-PKCS (Fig. A).

    Techniques: Ligation, Labeling, Incubation, Agarose Gel Electrophoresis

    DNA transactions by recombinant AaHMGB1 proteins. (A) Preferential binding of AaHMGB1 protein to supercoiled DNA. An equimolar mixture of supercoiled and linearized plasmid pTZ19R (∼10 nM) was pre-incubated with increasing amounts of AaHMGB1 (0.5–1 µM) and the DNA–protein complexes were resolved on a 1% agarose gel, followed by staining of the gel with ethidium bromide. Form I, supercoiled DNA; L, Linear DNA; Form II, relaxed circular DNA; (B) DNA supercoiling by AaHMGB1 and its truncated forms. Circular relaxed plasmid pTZ19R DNA was incubated in the presence of topoisomerase I (Topo I) and AaHMGB1 recombinant proteins (7–14 µM). Deproteinized DNA topoisomers were resolved on 1% agarose gels, followed by staining of the gel with ethidium bromide. Form I, supercoiled DNA; Form II, relaxed circular DNA. (C) DNA bending by AaHMGB1 and its truncated forms. A 32 P-labeled 123-bp DNA fragment (∼1 nM) was pre-incubated with recombinant proteins (25–50 nM) followed by ligation with T4 DNA ligase. Exonuclease III was used to verify the identity of DNA circles. The deproteinized DNA ligation products were subjected to electrophoresis on 6% non-denaturing polyacrylamide gels and visualized by autoradiography. Lm: linear multimers. Exo III, exonuclease III. These experiments were repeated three to five times each.

    Journal: PLoS ONE

    Article Title: The Dengue Vector Aedes aegypti Contains a Functional High Mobility Group Box 1 (HMGB1) Protein with a Unique Regulatory C-Terminus

    doi: 10.1371/journal.pone.0040192

    Figure Lengend Snippet: DNA transactions by recombinant AaHMGB1 proteins. (A) Preferential binding of AaHMGB1 protein to supercoiled DNA. An equimolar mixture of supercoiled and linearized plasmid pTZ19R (∼10 nM) was pre-incubated with increasing amounts of AaHMGB1 (0.5–1 µM) and the DNA–protein complexes were resolved on a 1% agarose gel, followed by staining of the gel with ethidium bromide. Form I, supercoiled DNA; L, Linear DNA; Form II, relaxed circular DNA; (B) DNA supercoiling by AaHMGB1 and its truncated forms. Circular relaxed plasmid pTZ19R DNA was incubated in the presence of topoisomerase I (Topo I) and AaHMGB1 recombinant proteins (7–14 µM). Deproteinized DNA topoisomers were resolved on 1% agarose gels, followed by staining of the gel with ethidium bromide. Form I, supercoiled DNA; Form II, relaxed circular DNA. (C) DNA bending by AaHMGB1 and its truncated forms. A 32 P-labeled 123-bp DNA fragment (∼1 nM) was pre-incubated with recombinant proteins (25–50 nM) followed by ligation with T4 DNA ligase. Exonuclease III was used to verify the identity of DNA circles. The deproteinized DNA ligation products were subjected to electrophoresis on 6% non-denaturing polyacrylamide gels and visualized by autoradiography. Lm: linear multimers. Exo III, exonuclease III. These experiments were repeated three to five times each.

    Article Snippet: The DNA was then ligated with T4 DNA ligase (0.6 unit/reaction; Promega) at 30°C for 30 min, and the ligation reactions were terminated by incubation of samples at 65°C for 15 min.

    Techniques: Recombinant, Binding Assay, Plasmid Preparation, Incubation, Agarose Gel Electrophoresis, Staining, Labeling, Ligation, DNA Ligation, Electrophoresis, Autoradiography

    DNA bending assays by posphorylated AaHMGB1. A 32 P-labelled 123-bp DNA fragment (∼1 nM) was pre-incubated with 50 ng of AaHMGB1 that were phosphorylated by PKA (panels A and B, lanes 5 and 2, respectively) or not (panels A and B, lanes 4 and 3, respectively), or by PKC (panels C and D, lanes 5 and 2, respectively) or not (panels C and D, lanes 4 and 3, respectively), followed by ligation with T4 DNA ligase. Exonuclease III was used to verify the identity of DNA circles. The deproteinized DNA ligation products were subjected to electrophoresis on 6% non-denaturing polyacrylamide gels and visualized by autoradiography. Lm: linear multimers. These experiments were repeated five times.

    Journal: PLoS ONE

    Article Title: The Dengue Vector Aedes aegypti Contains a Functional High Mobility Group Box 1 (HMGB1) Protein with a Unique Regulatory C-Terminus

    doi: 10.1371/journal.pone.0040192

    Figure Lengend Snippet: DNA bending assays by posphorylated AaHMGB1. A 32 P-labelled 123-bp DNA fragment (∼1 nM) was pre-incubated with 50 ng of AaHMGB1 that were phosphorylated by PKA (panels A and B, lanes 5 and 2, respectively) or not (panels A and B, lanes 4 and 3, respectively), or by PKC (panels C and D, lanes 5 and 2, respectively) or not (panels C and D, lanes 4 and 3, respectively), followed by ligation with T4 DNA ligase. Exonuclease III was used to verify the identity of DNA circles. The deproteinized DNA ligation products were subjected to electrophoresis on 6% non-denaturing polyacrylamide gels and visualized by autoradiography. Lm: linear multimers. These experiments were repeated five times.

    Article Snippet: The DNA was then ligated with T4 DNA ligase (0.6 unit/reaction; Promega) at 30°C for 30 min, and the ligation reactions were terminated by incubation of samples at 65°C for 15 min.

    Techniques: Incubation, Ligation, DNA Ligation, Electrophoresis, Autoradiography

    DNA cleavage is coordinated at 3′Dβ and Jβ RSSs. (A) Schematic of the Jβ1 ω , Jβ1 M2 , and Jβ1 M4 alleles as described in the legend to Fig. 1 with the BW linker ligated to cleaved 3′ Dβ1 and Jβ1.2 signal ends. The position of the oligonucleotide primers used to amplify linker ligated to the 3′ Dβ1 (BW-1H, 3A, and 3B) and Jβ1.2 (BWJ, JA, and JB) signal ends are shown as is the position of the oligonucleotide (P1) used to probe these PCR products. Genomic DNA from Jβ1 M4/ω or Jβ1 M2/ω DN thymocytes was incubated with the BW linker in the presence (+) or absence (−) or T4 DNA ligase and heminested LMPCRs performed to detect 3′ Dβ1 (B) and Jβ1.2 (C) signal ends as described in the Materials and Methods section. Analyses of Jβ1.2 signal ends was performed on ligated thymocyte DNA that was serially diluted into nonligated DNA keeping the total amount of DNA constant. Cell equivalents of ligated template DNA are indicated. Products from ligation of the BW linker to signal ends from the Jβ1 ω (ω), Jβ1 M4 (M4), and Jβ1 M2 (M2) alleles are indicated. Molecular weight markers are shown. Also shown is a RAG-2 gene PCR performed on ligated and nonligated template DNA and probed with the R2P oligonucleotide.

    Journal: The Journal of Experimental Medicine

    Article Title: Restrictions Limiting the Generation of DNA Double Strand Breaks during Chromosomal V(D)J Recombination

    doi: 10.1084/jem.20011803

    Figure Lengend Snippet: DNA cleavage is coordinated at 3′Dβ and Jβ RSSs. (A) Schematic of the Jβ1 ω , Jβ1 M2 , and Jβ1 M4 alleles as described in the legend to Fig. 1 with the BW linker ligated to cleaved 3′ Dβ1 and Jβ1.2 signal ends. The position of the oligonucleotide primers used to amplify linker ligated to the 3′ Dβ1 (BW-1H, 3A, and 3B) and Jβ1.2 (BWJ, JA, and JB) signal ends are shown as is the position of the oligonucleotide (P1) used to probe these PCR products. Genomic DNA from Jβ1 M4/ω or Jβ1 M2/ω DN thymocytes was incubated with the BW linker in the presence (+) or absence (−) or T4 DNA ligase and heminested LMPCRs performed to detect 3′ Dβ1 (B) and Jβ1.2 (C) signal ends as described in the Materials and Methods section. Analyses of Jβ1.2 signal ends was performed on ligated thymocyte DNA that was serially diluted into nonligated DNA keeping the total amount of DNA constant. Cell equivalents of ligated template DNA are indicated. Products from ligation of the BW linker to signal ends from the Jβ1 ω (ω), Jβ1 M4 (M4), and Jβ1 M2 (M2) alleles are indicated. Molecular weight markers are shown. Also shown is a RAG-2 gene PCR performed on ligated and nonligated template DNA and probed with the R2P oligonucleotide.

    Article Snippet: Purified thymocyte genomic DNA (2–3 μg) was ligated to 100 pmoles of the BW linker in a volume of 50–60 μl with 1–3 units T4 DNA Ligase (Promega) at 16°C for 12–14 h. Ligated samples were extracted with phenol and chloroform before PCR analysis.

    Techniques: Polymerase Chain Reaction, Incubation, Ligation, Molecular Weight

    Time-dependent ligation reactions. The optimum reaction conditions were used, which are 30 mM Tris–HCl (pH 7.5), 10 mM DTT, 3 mM MgCl 2 , 10 µM ATP, 20% DMSO, 2.5 pmol acceptor molecule, 70 fmol donor molecule, 3.125 pmol C4 template and 3 U T4 DNA ligase at 22°C. The graph shows the plots and fitted curves of each of the four ligation reactions analyzed (see inset for oligonucleotide sequences). The observed rate constants ( k obs ) for each reaction are given on the graph. The data points are the average of two independent assays.

    Journal: Nucleic Acids Research

    Article Title: Canonical nucleosides can be utilized by T4 DNA ligase as universal template bases at ligation junctions

    doi:

    Figure Lengend Snippet: Time-dependent ligation reactions. The optimum reaction conditions were used, which are 30 mM Tris–HCl (pH 7.5), 10 mM DTT, 3 mM MgCl 2 , 10 µM ATP, 20% DMSO, 2.5 pmol acceptor molecule, 70 fmol donor molecule, 3.125 pmol C4 template and 3 U T4 DNA ligase at 22°C. The graph shows the plots and fitted curves of each of the four ligation reactions analyzed (see inset for oligonucleotide sequences). The observed rate constants ( k obs ) for each reaction are given on the graph. The data points are the average of two independent assays.

    Article Snippet: Harada K. and Orgei,L.E. (1993) Unexpected substrate specificity of T4 DNA ligase revealed by in vitro selection.

    Techniques: Ligation

    Optimization of ligation reactions. ( A and B ) ATP and MgCl 2 concentration-dependent ligations using (c + p c)/T2. The MgCl 2 concentration was first optimized under standard reaction conditions of 30 mM Tris–HCl (pH 7.5), 10 mM DTT, 1 mM ATP, 2.5 pmol acceptor molecule, 70 fmol donor molecule, 3.125 pmol template and 3 U T4 DNA ligase at 30°C for 18 h. An optimum concentration of 3 mM MgCl 2 was found, which was then used in the subsequent ATP-dependent assay. ( C ) DMSO-dependent ligation reaction using (c + p c)/C2 in 3 mM MgCl 2 and 10 µM ATP. ( D ) Time-dependent ligation reaction using (c + p c)/T2 in 3 mM MgCl 2 , 10 µM ATP and 20% DMSO. In each case, product is represented by squares and intermediate by circles.

    Journal: Nucleic Acids Research

    Article Title: Canonical nucleosides can be utilized by T4 DNA ligase as universal template bases at ligation junctions

    doi:

    Figure Lengend Snippet: Optimization of ligation reactions. ( A and B ) ATP and MgCl 2 concentration-dependent ligations using (c + p c)/T2. The MgCl 2 concentration was first optimized under standard reaction conditions of 30 mM Tris–HCl (pH 7.5), 10 mM DTT, 1 mM ATP, 2.5 pmol acceptor molecule, 70 fmol donor molecule, 3.125 pmol template and 3 U T4 DNA ligase at 30°C for 18 h. An optimum concentration of 3 mM MgCl 2 was found, which was then used in the subsequent ATP-dependent assay. ( C ) DMSO-dependent ligation reaction using (c + p c)/C2 in 3 mM MgCl 2 and 10 µM ATP. ( D ) Time-dependent ligation reaction using (c + p c)/T2 in 3 mM MgCl 2 , 10 µM ATP and 20% DMSO. In each case, product is represented by squares and intermediate by circles.

    Article Snippet: Harada K. and Orgei,L.E. (1993) Unexpected substrate specificity of T4 DNA ligase revealed by in vitro selection.

    Techniques: Ligation, Concentration Assay