Structured Review

Roche t4 ligase
pol μ ribonucleotide incorporation with mixed nucleotide pools. (A) Alkali cleavage assay for incorporation of ribonucleotides. A single-nucleotide-gapped DNA substrate with template C was 5′ 32 P labeled as indicated (star). Alkali treatment of products containing incorporated ribonucleotides results in product cleavage, as well as transfer of the 32 P from the 20-nt downstream strand to the primer strand, generating a novel 26-nt species (species II). (B) DNA substrate (5 nM) as in panel A was present in all reaction mixtures. Where indicated, 0.05 U of <t>T4</t> ligase (+) and 0.5 nM pol μ (+) or 5 pM pol β (β) were added. Following a 1-min polymerization-ligation reaction, samples were treated with alkali as noted (+). A 25 μM concentration of each dNTP (d) or rNTP (r) was present as noted. Reaction mixtures 8 to 10 contained a mixture of both dNTPs and rNTPs (d + r), with 25 μM each dNTP and 25 μM (lane 8), 125 μM (lane 9), or 500 μM (lane 10) each rNTP. I, species I; II, species II.
T4 Ligase, supplied by Roche, used in various techniques. Bioz Stars score: 94/100, based on 11 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/result/t4 ligase/product/Roche
Average 94 stars, based on 11 article reviews
Price from $9.99 to $1999.99
t4 ligase - by Bioz Stars, 2020-05
94/100 stars

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1) Product Images from "Polymerase Mu Is a DNA-Directed DNA/RNA Polymerase"

Article Title: Polymerase Mu Is a DNA-Directed DNA/RNA Polymerase

Journal: Molecular and Cellular Biology

doi: 10.1128/MCB.23.7.2309-2315.2003

pol μ ribonucleotide incorporation with mixed nucleotide pools. (A) Alkali cleavage assay for incorporation of ribonucleotides. A single-nucleotide-gapped DNA substrate with template C was 5′ 32 P labeled as indicated (star). Alkali treatment of products containing incorporated ribonucleotides results in product cleavage, as well as transfer of the 32 P from the 20-nt downstream strand to the primer strand, generating a novel 26-nt species (species II). (B) DNA substrate (5 nM) as in panel A was present in all reaction mixtures. Where indicated, 0.05 U of T4 ligase (+) and 0.5 nM pol μ (+) or 5 pM pol β (β) were added. Following a 1-min polymerization-ligation reaction, samples were treated with alkali as noted (+). A 25 μM concentration of each dNTP (d) or rNTP (r) was present as noted. Reaction mixtures 8 to 10 contained a mixture of both dNTPs and rNTPs (d + r), with 25 μM each dNTP and 25 μM (lane 8), 125 μM (lane 9), or 500 μM (lane 10) each rNTP. I, species I; II, species II.
Figure Legend Snippet: pol μ ribonucleotide incorporation with mixed nucleotide pools. (A) Alkali cleavage assay for incorporation of ribonucleotides. A single-nucleotide-gapped DNA substrate with template C was 5′ 32 P labeled as indicated (star). Alkali treatment of products containing incorporated ribonucleotides results in product cleavage, as well as transfer of the 32 P from the 20-nt downstream strand to the primer strand, generating a novel 26-nt species (species II). (B) DNA substrate (5 nM) as in panel A was present in all reaction mixtures. Where indicated, 0.05 U of T4 ligase (+) and 0.5 nM pol μ (+) or 5 pM pol β (β) were added. Following a 1-min polymerization-ligation reaction, samples were treated with alkali as noted (+). A 25 μM concentration of each dNTP (d) or rNTP (r) was present as noted. Reaction mixtures 8 to 10 contained a mixture of both dNTPs and rNTPs (d + r), with 25 μM each dNTP and 25 μM (lane 8), 125 μM (lane 9), or 500 μM (lane 10) each rNTP. I, species I; II, species II.

Techniques Used: Cleavage Assay, Labeling, Ligation, Concentration Assay

2) Product Images from "Polymerase Mu Is a DNA-Directed DNA/RNA Polymerase"

Article Title: Polymerase Mu Is a DNA-Directed DNA/RNA Polymerase

Journal: Molecular and Cellular Biology

doi: 10.1128/MCB.23.7.2309-2315.2003

pol μ ribonucleotide incorporation with mixed nucleotide pools. (A) Alkali cleavage assay for incorporation of ribonucleotides. A single-nucleotide-gapped DNA substrate with template C was 5′ 32 P labeled as indicated (star). Alkali treatment of products containing incorporated ribonucleotides results in product cleavage, as well as transfer of the 32 P from the 20-nt downstream strand to the primer strand, generating a novel 26-nt species (species II). (B) DNA substrate (5 nM) as in panel A was present in all reaction mixtures. Where indicated, 0.05 U of T4 ligase (+) and 0.5 nM pol μ (+) or 5 pM pol β (β) were added. Following a 1-min polymerization-ligation reaction, samples were treated with alkali as noted (+). A 25 μM concentration of each dNTP (d) or rNTP (r) was present as noted. Reaction mixtures 8 to 10 contained a mixture of both dNTPs and rNTPs (d + r), with 25 μM each dNTP and 25 μM (lane 8), 125 μM (lane 9), or 500 μM (lane 10) each rNTP. I, species I; II, species II.
Figure Legend Snippet: pol μ ribonucleotide incorporation with mixed nucleotide pools. (A) Alkali cleavage assay for incorporation of ribonucleotides. A single-nucleotide-gapped DNA substrate with template C was 5′ 32 P labeled as indicated (star). Alkali treatment of products containing incorporated ribonucleotides results in product cleavage, as well as transfer of the 32 P from the 20-nt downstream strand to the primer strand, generating a novel 26-nt species (species II). (B) DNA substrate (5 nM) as in panel A was present in all reaction mixtures. Where indicated, 0.05 U of T4 ligase (+) and 0.5 nM pol μ (+) or 5 pM pol β (β) were added. Following a 1-min polymerization-ligation reaction, samples were treated with alkali as noted (+). A 25 μM concentration of each dNTP (d) or rNTP (r) was present as noted. Reaction mixtures 8 to 10 contained a mixture of both dNTPs and rNTPs (d + r), with 25 μM each dNTP and 25 μM (lane 8), 125 μM (lane 9), or 500 μM (lane 10) each rNTP. I, species I; II, species II.

Techniques Used: Cleavage Assay, Labeling, Ligation, Concentration Assay

Related Articles

Ligation:

Article Title: Assessing the severity of the small inframe deletion mutation in the ?-subunit of ?-hexosaminidase A found in the Turkish population by reproducing it in the more stable ?-subunit
Article Snippet: .. Triple ligation was performed overnight at 4°C by T4 ligase (Roche, Penzberg, Germany). .. Mutant pEFneoβ constructs were transformed into competent E. coli cells and clones were verified by DNA sequencing of the entire β-cDNA.

Article Title: Protein-DNA interaction mapping using genomic tiling path microarrays in Drosophila
Article Snippet: .. After inactivation of Dpn I at 80°C for 20 min, 4 μg of the Dpn I-digested genomic DNA was ligated to 40 pmol of a double-stranded unphosphorylated adaptor (top strand: 5′-CTAATACGACTCACTATAGGGCAGCGTGGTCGCGGCCGAGGA-3′, bottom strand: 5′-TCCTCGGCCG-3′) for 2 h at 16°C with 5 units of T4-Ligase (Roche Molecular Biochemicals) in a total volume of 20 μl of ligation buffer. .. To prevent amplification of DNA fragments containing unmethylated GATCs, 1 μg of the adaptor-ligated DNA was cut with 2 units of Dpn II (New England Biolabs) for1hat37°C in a total volume of 20 μl of Dpn II buffer.

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    Roche t4 dna ligase
    A–C : Comparison of apoptotic ( A , B ) and necrotic ( C ) thymus stained by DAPI (blue fluorescence) and by in situ ligation using oligonucleotide probes with single dA overhangs (red fluorescence). A and C are dual-stained images, B is a single-stained (DAPI) image and is provided for the easy comparison of nuclear morphology with C . D and E : Comparison of apoptotic and necrotic thymus stained by TUNEL. D : Apoptotic thymus (green fluorescence, TUNEL staining; blue fluorescence, DAPI staining). E : Necrotic thymus (green fluorescence, TUNEL staining; blue fluorescence, DAPI staining). Insets show high-magnification images of apoptotic and necrotic nuclei labeled by TUNEL (side of the inset , 80 μm). Strong positive staining is present in both cases. However necrotic thymus is uniformly positive with the loss of the characteristic pattern of cell death seen in glucocorticoid-induced apoptosis. F and G : Comparison of apoptotic and necrotic thymus using in situ ligation with blunt-ended probes detecting double-strand DNA breaks bearing 5′ phosphates. F : Apoptotic thymus, blunt ends detection. G : Necrotic thymus, blunt ends detection. Note that in situ ligation does not detect double-strand DNA breaks bearing 5′ phosphates in necrotic thymus. H and I : DNA nicks relegation in necrotic thymus using <t>T4</t> DNA ligase. H : Necrotic thymus TUNEL-stained before T4 ligase pretreatment; I : the same thymus after T4 ligase pretreatment. DNA nicks do not contribute to the strong positive staining of necrotic thymus by TUNEL. No change in TUNEL signal intensity occurred after treatment of necrotic tissue with T4 DNA ligase to seal DNA nicks. Apoptosis was induced in the thymus by an intraperitoneal injection of 6 mg/kg of dexamethasone. Necrosis was induced in the thymus by freezing with liquid nitrogen. Scale bars: 200 μm ( C , E ); 300 μm ( G ); 150 μm ( I ).
    T4 Dna Ligase, supplied by Roche, used in various techniques. Bioz Stars score: 95/100, based on 202 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/t4 dna ligase/product/Roche
    Average 95 stars, based on 202 article reviews
    Price from $9.99 to $1999.99
    t4 dna ligase - by Bioz Stars, 2020-05
    95/100 stars
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    A–C : Comparison of apoptotic ( A , B ) and necrotic ( C ) thymus stained by DAPI (blue fluorescence) and by in situ ligation using oligonucleotide probes with single dA overhangs (red fluorescence). A and C are dual-stained images, B is a single-stained (DAPI) image and is provided for the easy comparison of nuclear morphology with C . D and E : Comparison of apoptotic and necrotic thymus stained by TUNEL. D : Apoptotic thymus (green fluorescence, TUNEL staining; blue fluorescence, DAPI staining). E : Necrotic thymus (green fluorescence, TUNEL staining; blue fluorescence, DAPI staining). Insets show high-magnification images of apoptotic and necrotic nuclei labeled by TUNEL (side of the inset , 80 μm). Strong positive staining is present in both cases. However necrotic thymus is uniformly positive with the loss of the characteristic pattern of cell death seen in glucocorticoid-induced apoptosis. F and G : Comparison of apoptotic and necrotic thymus using in situ ligation with blunt-ended probes detecting double-strand DNA breaks bearing 5′ phosphates. F : Apoptotic thymus, blunt ends detection. G : Necrotic thymus, blunt ends detection. Note that in situ ligation does not detect double-strand DNA breaks bearing 5′ phosphates in necrotic thymus. H and I : DNA nicks relegation in necrotic thymus using T4 DNA ligase. H : Necrotic thymus TUNEL-stained before T4 ligase pretreatment; I : the same thymus after T4 ligase pretreatment. DNA nicks do not contribute to the strong positive staining of necrotic thymus by TUNEL. No change in TUNEL signal intensity occurred after treatment of necrotic tissue with T4 DNA ligase to seal DNA nicks. Apoptosis was induced in the thymus by an intraperitoneal injection of 6 mg/kg of dexamethasone. Necrosis was induced in the thymus by freezing with liquid nitrogen. Scale bars: 200 μm ( C , E ); 300 μm ( G ); 150 μm ( I ).

    Journal: The American Journal of Pathology

    Article Title: Early Necrotic DNA Degradation

    doi:

    Figure Lengend Snippet: A–C : Comparison of apoptotic ( A , B ) and necrotic ( C ) thymus stained by DAPI (blue fluorescence) and by in situ ligation using oligonucleotide probes with single dA overhangs (red fluorescence). A and C are dual-stained images, B is a single-stained (DAPI) image and is provided for the easy comparison of nuclear morphology with C . D and E : Comparison of apoptotic and necrotic thymus stained by TUNEL. D : Apoptotic thymus (green fluorescence, TUNEL staining; blue fluorescence, DAPI staining). E : Necrotic thymus (green fluorescence, TUNEL staining; blue fluorescence, DAPI staining). Insets show high-magnification images of apoptotic and necrotic nuclei labeled by TUNEL (side of the inset , 80 μm). Strong positive staining is present in both cases. However necrotic thymus is uniformly positive with the loss of the characteristic pattern of cell death seen in glucocorticoid-induced apoptosis. F and G : Comparison of apoptotic and necrotic thymus using in situ ligation with blunt-ended probes detecting double-strand DNA breaks bearing 5′ phosphates. F : Apoptotic thymus, blunt ends detection. G : Necrotic thymus, blunt ends detection. Note that in situ ligation does not detect double-strand DNA breaks bearing 5′ phosphates in necrotic thymus. H and I : DNA nicks relegation in necrotic thymus using T4 DNA ligase. H : Necrotic thymus TUNEL-stained before T4 ligase pretreatment; I : the same thymus after T4 ligase pretreatment. DNA nicks do not contribute to the strong positive staining of necrotic thymus by TUNEL. No change in TUNEL signal intensity occurred after treatment of necrotic tissue with T4 DNA ligase to seal DNA nicks. Apoptosis was induced in the thymus by an intraperitoneal injection of 6 mg/kg of dexamethasone. Necrosis was induced in the thymus by freezing with liquid nitrogen. Scale bars: 200 μm ( C , E ); 300 μm ( G ); 150 μm ( I ).

    Article Snippet: In a mock control reaction solution, an equal volume of 50% glycerol in water was substituted for T4 DNA ligase.

    Techniques: Staining, Fluorescence, In Situ, Ligation, TUNEL Assay, Labeling, Injection

    BER analysis of mitochondrial extracts. BER analysis was carried out in duplicate using two independently prepared mitochondrial extracts from HeLa (A) and U2OS (B) cell lines. The increased level of the upper band after T4 DNA ligase treatment relative

    Journal: DNA repair

    Article Title: Overexpression of DNA ligase III in mitochondria protects cells against oxidative stress and improves mitochondrial DNA base excision repair

    doi: 10.1016/j.dnarep.2014.01.015

    Figure Lengend Snippet: BER analysis of mitochondrial extracts. BER analysis was carried out in duplicate using two independently prepared mitochondrial extracts from HeLa (A) and U2OS (B) cell lines. The increased level of the upper band after T4 DNA ligase treatment relative

    Article Snippet: T4 DNA ligase was inactivated at 70 °C for 10 min and DNA was digested with XbaI and HindIII restriction enzymes and separated in 12% denaturing polyacrylamide gel at 300 V for 1 h.

    Techniques:

    ISL probe and regular ISL labeling. ( a ) A regular ISL probe is a blunt-ended DNA fragment with 3′OH/5′OH at the ends. It can be tagged with either digoxigenin or fluorophores. The probe's actual sequence is unimportant for the regular ISL procedure. The probe can be used in the isothermal signal amplification procedure if it contains a single recognition site for a blunt-end restrictase (Sma I in this example). ( b ) In the regular ISL, probes are ligated to 5′PO 4 blunt-ended apoptotic DNA breaks in tissue sections. They cannot ligate to each other due to the absence of 5′PO 4 termini required by T4 DNA ligase. Therefore, the regular ISL reaction stops when all DNA probes are ligated to 5′PO 4 breaks in apoptotic DNA

    Journal: Methods in molecular biology (Clifton, N.J.)

    Article Title: Zebra Tail Amplification: Accelerated Detection of Apoptotic Blunt-Ended DNA Breaks by In Situ Ligation

    doi: 10.1007/978-1-4939-7187-9_15

    Figure Lengend Snippet: ISL probe and regular ISL labeling. ( a ) A regular ISL probe is a blunt-ended DNA fragment with 3′OH/5′OH at the ends. It can be tagged with either digoxigenin or fluorophores. The probe's actual sequence is unimportant for the regular ISL procedure. The probe can be used in the isothermal signal amplification procedure if it contains a single recognition site for a blunt-end restrictase (Sma I in this example). ( b ) In the regular ISL, probes are ligated to 5′PO 4 blunt-ended apoptotic DNA breaks in tissue sections. They cannot ligate to each other due to the absence of 5′PO 4 termini required by T4 DNA ligase. Therefore, the regular ISL reaction stops when all DNA probes are ligated to 5′PO 4 breaks in apoptotic DNA

    Article Snippet: In the mock reaction an equal volume of 50% glycerol in water is substituted for T4 DNA ligase.

    Techniques: Labeling, Sequencing, Amplification