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  • 99
    New England Biolabs t4 dna ligase
    T4 Dna Ligase, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 99/100, based on 50072 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher t4 dna ligase
    T4 Dna Ligase, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 25057 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    New England Biolabs t4 polynucleotide kinase
    T4 Polynucleotide Kinase, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 99/100, based on 29397 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher t4 pnk
    T4 Pnk, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 812 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    New England Biolabs t4 dna polymerase
    T4 Dna Polymerase, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 99/100, based on 9070 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    94
    Promega t4 dna ligase
    ATP-fueled transient colloid assembly (TCA), and dynamic DNA surface polymerization (TSP). a Schematic illustration of DNA wrapping-induced TCA. b Time-dependent CLSM of TCA. Scale bars = 10 μm. c Schematic representation of TSP. d Time-dependent CLSM of transient DNA growth on colloidal surface. Scale bar = 5 μm; insert = 10 μm. e Time-dependent fluorescence intensity of the fluorescent shell for TCA with programmable lifetimes. Lines are guides to the eye. Error bars are standard deviations of 50 colloids. f Singlet fraction of the colloids for TCA ( d 1 b = 38 bp) and TSP. Points correspond to analysis at random places. g Thickness of the fluorescent shells for TSP and TCA at 1 h (i: TSP, ii: TCA, iii: extracted spectra of fluorescence intensities from i and ii). Scale bar, 1 μm. h Secondary DNA tile integration and surface function reconfiguration during a running TSP. The second tile with ATTO 647 was added 1 h after the Cy3 tile. CLSM before ( i ) and after ( j ) adding the ATTO 647 tile confirm the integration and reconfiguration. Scale bar = 2 μm. k Time dependent fluorescence intensities on the colloid surface. Lines are guides to the eye. Error bars are standard deviations of 50 colloids. Conditions for a : 10.0 μM dsDNA-Cy3, 12.0 μM ssDNA, 0.92 WU μL −1 <t>T4</t> DNA ligase, 1.0 U μL −1 BsaI, ca. 0.167 mg mL −1 docking strand modified colloids (ca. 1.17–1.67 × 10 8 beads mL −1 ), and 50 μM ATP, 37 °C. Conditions for c : same condition as a but using dsDNA functionalized colloids bearing sticky end a’ on the docking strand to initiate a surface tethered main chain of the DySS SfNAP. All CLSM measurements were conducted consecutively with the same microscopy settings (see Supplementary Table 1 for sequences).
    T4 Dna Ligase, supplied by Promega, used in various techniques. Bioz Stars score: 94/100, based on 11518 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    TaKaRa dna ligases
    ATP-fueled transient colloid assembly (TCA), and dynamic DNA surface polymerization (TSP). a Schematic illustration of DNA wrapping-induced TCA. b Time-dependent CLSM of TCA. Scale bars = 10 μm. c Schematic representation of TSP. d Time-dependent CLSM of transient DNA growth on colloidal surface. Scale bar = 5 μm; insert = 10 μm. e Time-dependent fluorescence intensity of the fluorescent shell for TCA with programmable lifetimes. Lines are guides to the eye. Error bars are standard deviations of 50 colloids. f Singlet fraction of the colloids for TCA ( d 1 b = 38 bp) and TSP. Points correspond to analysis at random places. g Thickness of the fluorescent shells for TSP and TCA at 1 h (i: TSP, ii: TCA, iii: extracted spectra of fluorescence intensities from i and ii). Scale bar, 1 μm. h Secondary DNA tile integration and surface function reconfiguration during a running TSP. The second tile with ATTO 647 was added 1 h after the Cy3 tile. CLSM before ( i ) and after ( j ) adding the ATTO 647 tile confirm the integration and reconfiguration. Scale bar = 2 μm. k Time dependent fluorescence intensities on the colloid surface. Lines are guides to the eye. Error bars are standard deviations of 50 colloids. Conditions for a : 10.0 μM dsDNA-Cy3, 12.0 μM ssDNA, 0.92 WU μL −1 <t>T4</t> DNA ligase, 1.0 U μL −1 BsaI, ca. 0.167 mg mL −1 docking strand modified colloids (ca. 1.17–1.67 × 10 8 beads mL −1 ), and 50 μM ATP, 37 °C. Conditions for c : same condition as a but using dsDNA functionalized colloids bearing sticky end a’ on the docking strand to initiate a surface tethered main chain of the DySS SfNAP. All CLSM measurements were conducted consecutively with the same microscopy settings (see Supplementary Table 1 for sequences).
    Dna Ligases, supplied by TaKaRa, used in various techniques. Bioz Stars score: 99/100, based on 38 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    93
    Promega t4 polynucleotide kinase
    Direct labelling of RNA on the 5′ termini of replication intermediates with vaccinia virus guanylyltransferase, an enzyme responsible for messenger RNA capping. ( A ) The 5′ ends of RNA primers attached to P. abyssi replication intermediates were capped with radioactive GTP and samples were fractionated on a sequencing gel (lane 2). Lane 3 contains the same reaction as lane 2, except that total DNA was treated with the restriction endonuclease Eco RI before analysis, confirming that nascent strands were efficiently denatured from template strands. Lane 1, size marker (10-base ladder; Gibco-BRL) labelled with [γ- 32 P]ATP using <t>T4</t> polynucleotide kinase. ( B ) Schematic diagram of the archaeal Okazaki fragment. The smallest replication intermediate detected (10 nt) is in accordance with the minimum length of RNA primers detected in vitro ).
    T4 Polynucleotide Kinase, supplied by Promega, used in various techniques. Bioz Stars score: 93/100, based on 5420 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    New England Biolabs t4 rna ligase
    Direct labelling of RNA on the 5′ termini of replication intermediates with vaccinia virus guanylyltransferase, an enzyme responsible for messenger RNA capping. ( A ) The 5′ ends of RNA primers attached to P. abyssi replication intermediates were capped with radioactive GTP and samples were fractionated on a sequencing gel (lane 2). Lane 3 contains the same reaction as lane 2, except that total DNA was treated with the restriction endonuclease Eco RI before analysis, confirming that nascent strands were efficiently denatured from template strands. Lane 1, size marker (10-base ladder; Gibco-BRL) labelled with [γ- 32 P]ATP using <t>T4</t> polynucleotide kinase. ( B ) Schematic diagram of the archaeal Okazaki fragment. The smallest replication intermediate detected (10 nt) is in accordance with the minimum length of RNA primers detected in vitro ).
    T4 Rna Ligase, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 99/100, based on 3908 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher t4 ligase
    Direct labelling of RNA on the 5′ termini of replication intermediates with vaccinia virus guanylyltransferase, an enzyme responsible for messenger RNA capping. ( A ) The 5′ ends of RNA primers attached to P. abyssi replication intermediates were capped with radioactive GTP and samples were fractionated on a sequencing gel (lane 2). Lane 3 contains the same reaction as lane 2, except that total DNA was treated with the restriction endonuclease Eco RI before analysis, confirming that nascent strands were efficiently denatured from template strands. Lane 1, size marker (10-base ladder; Gibco-BRL) labelled with [γ- 32 P]ATP using <t>T4</t> polynucleotide kinase. ( B ) Schematic diagram of the archaeal Okazaki fragment. The smallest replication intermediate detected (10 nt) is in accordance with the minimum length of RNA primers detected in vitro ).
    T4 Ligase, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 5002 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    New England Biolabs polynucleotide kinase
    Direct labelling of RNA on the 5′ termini of replication intermediates with vaccinia virus guanylyltransferase, an enzyme responsible for messenger RNA capping. ( A ) The 5′ ends of RNA primers attached to P. abyssi replication intermediates were capped with radioactive GTP and samples were fractionated on a sequencing gel (lane 2). Lane 3 contains the same reaction as lane 2, except that total DNA was treated with the restriction endonuclease Eco RI before analysis, confirming that nascent strands were efficiently denatured from template strands. Lane 1, size marker (10-base ladder; Gibco-BRL) labelled with [γ- 32 P]ATP using <t>T4</t> polynucleotide kinase. ( B ) Schematic diagram of the archaeal Okazaki fragment. The smallest replication intermediate detected (10 nt) is in accordance with the minimum length of RNA primers detected in vitro ).
    Polynucleotide Kinase, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 99/100, based on 3121 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    New England Biolabs t4 dna ligase buffer
    Direct labelling of RNA on the 5′ termini of replication intermediates with vaccinia virus guanylyltransferase, an enzyme responsible for messenger RNA capping. ( A ) The 5′ ends of RNA primers attached to P. abyssi replication intermediates were capped with radioactive GTP and samples were fractionated on a sequencing gel (lane 2). Lane 3 contains the same reaction as lane 2, except that total DNA was treated with the restriction endonuclease Eco RI before analysis, confirming that nascent strands were efficiently denatured from template strands. Lane 1, size marker (10-base ladder; Gibco-BRL) labelled with [γ- 32 P]ATP using <t>T4</t> polynucleotide kinase. ( B ) Schematic diagram of the archaeal Okazaki fragment. The smallest replication intermediate detected (10 nt) is in accordance with the minimum length of RNA primers detected in vitro ).
    T4 Dna Ligase Buffer, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 99/100, based on 2917 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    New England Biolabs t4 rna ligase 2
    Direct labelling of RNA on the 5′ termini of replication intermediates with vaccinia virus guanylyltransferase, an enzyme responsible for messenger RNA capping. ( A ) The 5′ ends of RNA primers attached to P. abyssi replication intermediates were capped with radioactive GTP and samples were fractionated on a sequencing gel (lane 2). Lane 3 contains the same reaction as lane 2, except that total DNA was treated with the restriction endonuclease Eco RI before analysis, confirming that nascent strands were efficiently denatured from template strands. Lane 1, size marker (10-base ladder; Gibco-BRL) labelled with [γ- 32 P]ATP using <t>T4</t> polynucleotide kinase. ( B ) Schematic diagram of the archaeal Okazaki fragment. The smallest replication intermediate detected (10 nt) is in accordance with the minimum length of RNA primers detected in vitro ).
    T4 Rna Ligase 2, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 99/100, based on 2023 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher t4 dna polymerase
    Direct labelling of RNA on the 5′ termini of replication intermediates with vaccinia virus guanylyltransferase, an enzyme responsible for messenger RNA capping. ( A ) The 5′ ends of RNA primers attached to P. abyssi replication intermediates were capped with radioactive GTP and samples were fractionated on a sequencing gel (lane 2). Lane 3 contains the same reaction as lane 2, except that total DNA was treated with the restriction endonuclease Eco RI before analysis, confirming that nascent strands were efficiently denatured from template strands. Lane 1, size marker (10-base ladder; Gibco-BRL) labelled with [γ- 32 P]ATP using <t>T4</t> polynucleotide kinase. ( B ) Schematic diagram of the archaeal Okazaki fragment. The smallest replication intermediate detected (10 nt) is in accordance with the minimum length of RNA primers detected in vitro ).
    T4 Dna Polymerase, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 2670 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    93
    Promega t4 ligase
    Time course of BER. 35-μl reactions were performed as described under “Experimental Procedures” with UL2, APE, and UL30 in the presence of either <t>T4</t> ligase (0.001 units/μl) ( A ), ligase I ( B ), or ligase IIIα-XRCC1 ( C ). 5-μl aliquots were removed at the times indicated. Activity is expressed as the fraction of maximum for the nicked ( N ) (●) and ligation ( L ) (○) products. The insets in each panel show the relevant gel images.
    T4 Ligase, supplied by Promega, used in various techniques. Bioz Stars score: 93/100, based on 2304 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Boehringer Mannheim t4 dna ligase
    <t>T4</t> DNA-ligase activity in the presence of SA. T4 DNA-ligase was treated or not with increasing SA concentrations (15 min at 25°C) and then incubated at 37°C for 1 min with the oligo substrate. ( A ) The oligo(dT) 16 multimers were separated in polyacrylamide/urea gels: T4 DNA-ligase without SA (lane 1) or incubated with increasing SA concentrations of 2.5(2), 5(3), 10(4), and 20(5) μM. ( B ) The activity was quantitated using an InstantImager (Packard). ( C ) Inhibition of enzyme-adenylate formation by SA. T4 DNA-ligase was incubated or not with increasing SA concentrations (15 min at 25°C) before the addition of [α- 32 P]ATP. The enzyme adenylate complexes were separated by electrophoresis and detected by autoradiography.
    T4 Dna Ligase, supplied by Boehringer Mannheim, used in various techniques. Bioz Stars score: 92/100, based on 908 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    TaKaRa t4 rna ligase
    <t>T4</t> DNA-ligase activity in the presence of SA. T4 DNA-ligase was treated or not with increasing SA concentrations (15 min at 25°C) and then incubated at 37°C for 1 min with the oligo substrate. ( A ) The oligo(dT) 16 multimers were separated in polyacrylamide/urea gels: T4 DNA-ligase without SA (lane 1) or incubated with increasing SA concentrations of 2.5(2), 5(3), 10(4), and 20(5) μM. ( B ) The activity was quantitated using an InstantImager (Packard). ( C ) Inhibition of enzyme-adenylate formation by SA. T4 DNA-ligase was incubated or not with increasing SA concentrations (15 min at 25°C) before the addition of [α- 32 P]ATP. The enzyme adenylate complexes were separated by electrophoresis and detected by autoradiography.
    T4 Rna Ligase, supplied by TaKaRa, used in various techniques. Bioz Stars score: 98/100, based on 1171 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    95
    TaKaRa t4 dna polymerase
    <t>T4</t> DNA-ligase activity in the presence of SA. T4 DNA-ligase was treated or not with increasing SA concentrations (15 min at 25°C) and then incubated at 37°C for 1 min with the oligo substrate. ( A ) The oligo(dT) 16 multimers were separated in polyacrylamide/urea gels: T4 DNA-ligase without SA (lane 1) or incubated with increasing SA concentrations of 2.5(2), 5(3), 10(4), and 20(5) μM. ( B ) The activity was quantitated using an InstantImager (Packard). ( C ) Inhibition of enzyme-adenylate formation by SA. T4 DNA-ligase was incubated or not with increasing SA concentrations (15 min at 25°C) before the addition of [α- 32 P]ATP. The enzyme adenylate complexes were separated by electrophoresis and detected by autoradiography.
    T4 Dna Polymerase, supplied by TaKaRa, used in various techniques. Bioz Stars score: 95/100, based on 864 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Image Search Results


    ATP-fueled transient colloid assembly (TCA), and dynamic DNA surface polymerization (TSP). a Schematic illustration of DNA wrapping-induced TCA. b Time-dependent CLSM of TCA. Scale bars = 10 μm. c Schematic representation of TSP. d Time-dependent CLSM of transient DNA growth on colloidal surface. Scale bar = 5 μm; insert = 10 μm. e Time-dependent fluorescence intensity of the fluorescent shell for TCA with programmable lifetimes. Lines are guides to the eye. Error bars are standard deviations of 50 colloids. f Singlet fraction of the colloids for TCA ( d 1 b = 38 bp) and TSP. Points correspond to analysis at random places. g Thickness of the fluorescent shells for TSP and TCA at 1 h (i: TSP, ii: TCA, iii: extracted spectra of fluorescence intensities from i and ii). Scale bar, 1 μm. h Secondary DNA tile integration and surface function reconfiguration during a running TSP. The second tile with ATTO 647 was added 1 h after the Cy3 tile. CLSM before ( i ) and after ( j ) adding the ATTO 647 tile confirm the integration and reconfiguration. Scale bar = 2 μm. k Time dependent fluorescence intensities on the colloid surface. Lines are guides to the eye. Error bars are standard deviations of 50 colloids. Conditions for a : 10.0 μM dsDNA-Cy3, 12.0 μM ssDNA, 0.92 WU μL −1 T4 DNA ligase, 1.0 U μL −1 BsaI, ca. 0.167 mg mL −1 docking strand modified colloids (ca. 1.17–1.67 × 10 8 beads mL −1 ), and 50 μM ATP, 37 °C. Conditions for c : same condition as a but using dsDNA functionalized colloids bearing sticky end a’ on the docking strand to initiate a surface tethered main chain of the DySS SfNAP. All CLSM measurements were conducted consecutively with the same microscopy settings (see Supplementary Table 1 for sequences).

    Journal: Nature Communications

    Article Title: ATP-powered molecular recognition to engineer transient multivalency and self-sorting 4D hierarchical systems

    doi: 10.1038/s41467-020-17479-9

    Figure Lengend Snippet: ATP-fueled transient colloid assembly (TCA), and dynamic DNA surface polymerization (TSP). a Schematic illustration of DNA wrapping-induced TCA. b Time-dependent CLSM of TCA. Scale bars = 10 μm. c Schematic representation of TSP. d Time-dependent CLSM of transient DNA growth on colloidal surface. Scale bar = 5 μm; insert = 10 μm. e Time-dependent fluorescence intensity of the fluorescent shell for TCA with programmable lifetimes. Lines are guides to the eye. Error bars are standard deviations of 50 colloids. f Singlet fraction of the colloids for TCA ( d 1 b = 38 bp) and TSP. Points correspond to analysis at random places. g Thickness of the fluorescent shells for TSP and TCA at 1 h (i: TSP, ii: TCA, iii: extracted spectra of fluorescence intensities from i and ii). Scale bar, 1 μm. h Secondary DNA tile integration and surface function reconfiguration during a running TSP. The second tile with ATTO 647 was added 1 h after the Cy3 tile. CLSM before ( i ) and after ( j ) adding the ATTO 647 tile confirm the integration and reconfiguration. Scale bar = 2 μm. k Time dependent fluorescence intensities on the colloid surface. Lines are guides to the eye. Error bars are standard deviations of 50 colloids. Conditions for a : 10.0 μM dsDNA-Cy3, 12.0 μM ssDNA, 0.92 WU μL −1 T4 DNA ligase, 1.0 U μL −1 BsaI, ca. 0.167 mg mL −1 docking strand modified colloids (ca. 1.17–1.67 × 10 8 beads mL −1 ), and 50 μM ATP, 37 °C. Conditions for c : same condition as a but using dsDNA functionalized colloids bearing sticky end a’ on the docking strand to initiate a surface tethered main chain of the DySS SfNAP. All CLSM measurements were conducted consecutively with the same microscopy settings (see Supplementary Table 1 for sequences).

    Article Snippet: The transient polymerization of such dsDNA tiles was carried out at 25 and 37 °C in 1x NEB CutSmart buffer (50 mM potassium acetate, 20 mM tris-acetate, 10 mM magnesium acetate, 100 μg mL−1 BSA) with 0.05 mM M1, 0.46 WU μL−1 T4 DNA ligase (HC, 20 WU μL−1 , Promega), and 2.5 U μL−1 BsaI (20 U μL−1 , NEB ).

    Techniques: Confocal Laser Scanning Microscopy, Fluorescence, Modification, Microscopy

    ATP-fueled transient self-sorting (TSeSo) on a multicomponent systems level. a , b Schemes representing two different self-sorting multicomponent colloidal systems. c CLSM of two ATP-fueled colloidal assemblies, TSeSo-A, operating in the same systems in parallel. Scale bar = 5 μm; insert = 10 μm. d CLSM of TSeSo-B, whereby one colloid species is driven into assembly (TCA), while the other species is encapsulated (TSP). e Time-dependent AGE of the SfNAPs formed in TSeSo-A. f Time-dependent singlet fraction of both self-sorting systems. Points correspond to analysis at random places. g Fluorescence intensity of the colloids with time for both self-sorting systems. Lines are guides to the eye. Error bars are standard deviations of 50 colloids. Conditions for a : 10.0 μM dsDNA 1 -Cy3, 12.0 μM ssDNA 1 , 10.0 μM dsDNA 2 -ATTO 647, 12.0 μM ssDNA 2 , 0.92 WU μL −1 T4 DNA ligase, 1.0 U μL −1 BsaI, ca. 0.1 mg mL −1 DoS 1 modified colloid, 0.1 mg mL −1 DoS 2 modified colloid, 83.3 μM ATP, 37 °C. Conditions for b : The same condition as a except that the DoS 1 is changed to sticky-end a’ terminated dsDNA. All CLSM measurements were conducted consecutively with the same microscopy settings (see Supplementary Table 1 for sequences).

    Journal: Nature Communications

    Article Title: ATP-powered molecular recognition to engineer transient multivalency and self-sorting 4D hierarchical systems

    doi: 10.1038/s41467-020-17479-9

    Figure Lengend Snippet: ATP-fueled transient self-sorting (TSeSo) on a multicomponent systems level. a , b Schemes representing two different self-sorting multicomponent colloidal systems. c CLSM of two ATP-fueled colloidal assemblies, TSeSo-A, operating in the same systems in parallel. Scale bar = 5 μm; insert = 10 μm. d CLSM of TSeSo-B, whereby one colloid species is driven into assembly (TCA), while the other species is encapsulated (TSP). e Time-dependent AGE of the SfNAPs formed in TSeSo-A. f Time-dependent singlet fraction of both self-sorting systems. Points correspond to analysis at random places. g Fluorescence intensity of the colloids with time for both self-sorting systems. Lines are guides to the eye. Error bars are standard deviations of 50 colloids. Conditions for a : 10.0 μM dsDNA 1 -Cy3, 12.0 μM ssDNA 1 , 10.0 μM dsDNA 2 -ATTO 647, 12.0 μM ssDNA 2 , 0.92 WU μL −1 T4 DNA ligase, 1.0 U μL −1 BsaI, ca. 0.1 mg mL −1 DoS 1 modified colloid, 0.1 mg mL −1 DoS 2 modified colloid, 83.3 μM ATP, 37 °C. Conditions for b : The same condition as a except that the DoS 1 is changed to sticky-end a’ terminated dsDNA. All CLSM measurements were conducted consecutively with the same microscopy settings (see Supplementary Table 1 for sequences).

    Article Snippet: The transient polymerization of such dsDNA tiles was carried out at 25 and 37 °C in 1x NEB CutSmart buffer (50 mM potassium acetate, 20 mM tris-acetate, 10 mM magnesium acetate, 100 μg mL−1 BSA) with 0.05 mM M1, 0.46 WU μL−1 T4 DNA ligase (HC, 20 WU μL−1 , Promega), and 2.5 U μL−1 BsaI (20 U μL−1 , NEB ).

    Techniques: Confocal Laser Scanning Microscopy, Fluorescence, Modification, Microscopy

    ATP-fueled transient DySS DNA polymerization. a , b General scheme for ATP-powered transient DySS polymerization and control over lifetimes and adaptive DySS properties. The system enters into a DySS, state B, from its monomer state, state A, upon fueling with ATP, and returns to state A once the ATP is consumed. c Time-dependent AGE (2 wt. %, 90 V, 2 h) for transient DNA polymerizations with programmable lifetimes by fueling with varied ATP concentrations (0.1, 0.2, 0.6, and 1.0 mM). d Gray scale profiles from AGE at 1.0 mM ATP quantifying the transient shift of molecular weight, which is used to calculate the mass-weighted average chain length ( \documentclass[12pt]{minimal} \usepackage{amsmath} \usepackage{wasysym} \usepackage{amsfonts} \usepackage{amssymb} \usepackage{amsbsy} \usepackage{mathrsfs} \usepackage{upgreek} \setlength{\oddsidemargin}{-69pt} \begin{document}$$\overline {{\mathrm{bp}}}$$\end{document} bp ¯ w ) for each kinetic aliquot. e The \documentclass[12pt]{minimal} \usepackage{amsmath} \usepackage{wasysym} \usepackage{amsfonts} \usepackage{amssymb} \usepackage{amsbsy} \usepackage{mathrsfs} \usepackage{upgreek} \setlength{\oddsidemargin}{-69pt} \begin{document}$$\overline {{\mathrm{bp}}}$$\end{document} bp ¯ w development with time by varying the ATP concentration from 0.1 to 1.0 mM. Lines are guides to the eye. Multiple ATP injections are shown in Fig. 3 . f Lifetimes are controlled by the ATP concentration. Error bars are standard deviations of duplicate measurements. Conditions: 37 °C, 0.05 mM M1, 0.46 WU μL −1 T4 DNA ligase, 2.5 U μL −1 BsaI, and varying amounts of ATP.

    Journal: Nature Communications

    Article Title: ATP-powered molecular recognition to engineer transient multivalency and self-sorting 4D hierarchical systems

    doi: 10.1038/s41467-020-17479-9

    Figure Lengend Snippet: ATP-fueled transient DySS DNA polymerization. a , b General scheme for ATP-powered transient DySS polymerization and control over lifetimes and adaptive DySS properties. The system enters into a DySS, state B, from its monomer state, state A, upon fueling with ATP, and returns to state A once the ATP is consumed. c Time-dependent AGE (2 wt. %, 90 V, 2 h) for transient DNA polymerizations with programmable lifetimes by fueling with varied ATP concentrations (0.1, 0.2, 0.6, and 1.0 mM). d Gray scale profiles from AGE at 1.0 mM ATP quantifying the transient shift of molecular weight, which is used to calculate the mass-weighted average chain length ( \documentclass[12pt]{minimal} \usepackage{amsmath} \usepackage{wasysym} \usepackage{amsfonts} \usepackage{amssymb} \usepackage{amsbsy} \usepackage{mathrsfs} \usepackage{upgreek} \setlength{\oddsidemargin}{-69pt} \begin{document}$$\overline {{\mathrm{bp}}}$$\end{document} bp ¯ w ) for each kinetic aliquot. e The \documentclass[12pt]{minimal} \usepackage{amsmath} \usepackage{wasysym} \usepackage{amsfonts} \usepackage{amssymb} \usepackage{amsbsy} \usepackage{mathrsfs} \usepackage{upgreek} \setlength{\oddsidemargin}{-69pt} \begin{document}$$\overline {{\mathrm{bp}}}$$\end{document} bp ¯ w development with time by varying the ATP concentration from 0.1 to 1.0 mM. Lines are guides to the eye. Multiple ATP injections are shown in Fig. 3 . f Lifetimes are controlled by the ATP concentration. Error bars are standard deviations of duplicate measurements. Conditions: 37 °C, 0.05 mM M1, 0.46 WU μL −1 T4 DNA ligase, 2.5 U μL −1 BsaI, and varying amounts of ATP.

    Article Snippet: The transient polymerization of such dsDNA tiles was carried out at 25 and 37 °C in 1x NEB CutSmart buffer (50 mM potassium acetate, 20 mM tris-acetate, 10 mM magnesium acetate, 100 μg mL−1 BSA) with 0.05 mM M1, 0.46 WU μL−1 T4 DNA ligase (HC, 20 WU μL−1 , Promega), and 2.5 U μL−1 BsaI (20 U μL−1 , NEB ).

    Techniques: Molecular Weight, Concentration Assay

    ATP-fueled transient sequence-defined functionalized nucleic acid oligomers and polymers (SfNAPs). a – e Combinatorial organization of transient and programmable sequence-defined oligomers composed of varying inputs with orthogonally complementary ligation sites. Phi indicates disassembly after ATP consumption. c Time-dependent AGE with d extracted gray scale profiles for the transient oligomerization of the tetramer and e the distribution profiles of all oligomers in the DySS. f – j ATP-fueled DySS transient SfNAP with transient FRET signaling function. g Logic pathways for the ligation of the inputs to achieve FRET DNA polymers. h Time-dependent AGE (2 wt.%, 90 V, 2 h) of the transient SfNAP fueled with 0.12 mM ATP. i The \documentclass[12pt]{minimal} \usepackage{amsmath} \usepackage{wasysym} \usepackage{amsfonts} \usepackage{amssymb} \usepackage{amsbsy} \usepackage{mathrsfs} \usepackage{upgreek} \setlength{\oddsidemargin}{-69pt} \begin{document}$$\overline {{\mathrm{bp}}}$$\end{document} bp ¯ w development with time at different ATP concentrations. Lines are guides to the eye. j Transient FRET signaling at different ATP concentrations ( λ ex = 530 nm), and repeated fuel addition for 0.05 mM ATP; insert: FRET lifetime control by changing the ATP concentration. Shaded areas and error bars correspond to standard deviations from duplicate measurements. Conditions for a : 0.05 mM dsDNA tiles in total; 0.2 mM ATP, 0.92 WU μL −1 T4 DNA ligase, 1.0 U μL −1 BsaI, 37 °C. Conditions for f – j : 8.0 μM dsDNA-A, 8.0 μM dsDNA-C, 8.0 μM ssDNA-B, 0.92 WU μL −1 T4 DNA ligase, 1.0 U μL −1 BsaI, varied concentration of ATP, 37 °C.

    Journal: Nature Communications

    Article Title: ATP-powered molecular recognition to engineer transient multivalency and self-sorting 4D hierarchical systems

    doi: 10.1038/s41467-020-17479-9

    Figure Lengend Snippet: ATP-fueled transient sequence-defined functionalized nucleic acid oligomers and polymers (SfNAPs). a – e Combinatorial organization of transient and programmable sequence-defined oligomers composed of varying inputs with orthogonally complementary ligation sites. Phi indicates disassembly after ATP consumption. c Time-dependent AGE with d extracted gray scale profiles for the transient oligomerization of the tetramer and e the distribution profiles of all oligomers in the DySS. f – j ATP-fueled DySS transient SfNAP with transient FRET signaling function. g Logic pathways for the ligation of the inputs to achieve FRET DNA polymers. h Time-dependent AGE (2 wt.%, 90 V, 2 h) of the transient SfNAP fueled with 0.12 mM ATP. i The \documentclass[12pt]{minimal} \usepackage{amsmath} \usepackage{wasysym} \usepackage{amsfonts} \usepackage{amssymb} \usepackage{amsbsy} \usepackage{mathrsfs} \usepackage{upgreek} \setlength{\oddsidemargin}{-69pt} \begin{document}$$\overline {{\mathrm{bp}}}$$\end{document} bp ¯ w development with time at different ATP concentrations. Lines are guides to the eye. j Transient FRET signaling at different ATP concentrations ( λ ex = 530 nm), and repeated fuel addition for 0.05 mM ATP; insert: FRET lifetime control by changing the ATP concentration. Shaded areas and error bars correspond to standard deviations from duplicate measurements. Conditions for a : 0.05 mM dsDNA tiles in total; 0.2 mM ATP, 0.92 WU μL −1 T4 DNA ligase, 1.0 U μL −1 BsaI, 37 °C. Conditions for f – j : 8.0 μM dsDNA-A, 8.0 μM dsDNA-C, 8.0 μM ssDNA-B, 0.92 WU μL −1 T4 DNA ligase, 1.0 U μL −1 BsaI, varied concentration of ATP, 37 °C.

    Article Snippet: The transient polymerization of such dsDNA tiles was carried out at 25 and 37 °C in 1x NEB CutSmart buffer (50 mM potassium acetate, 20 mM tris-acetate, 10 mM magnesium acetate, 100 μg mL−1 BSA) with 0.05 mM M1, 0.46 WU μL−1 T4 DNA ligase (HC, 20 WU μL−1 , Promega), and 2.5 U μL−1 BsaI (20 U μL−1 , NEB ).

    Techniques: Sequencing, Ligation, Concentration Assay

    Direct labelling of RNA on the 5′ termini of replication intermediates with vaccinia virus guanylyltransferase, an enzyme responsible for messenger RNA capping. ( A ) The 5′ ends of RNA primers attached to P. abyssi replication intermediates were capped with radioactive GTP and samples were fractionated on a sequencing gel (lane 2). Lane 3 contains the same reaction as lane 2, except that total DNA was treated with the restriction endonuclease Eco RI before analysis, confirming that nascent strands were efficiently denatured from template strands. Lane 1, size marker (10-base ladder; Gibco-BRL) labelled with [γ- 32 P]ATP using T4 polynucleotide kinase. ( B ) Schematic diagram of the archaeal Okazaki fragment. The smallest replication intermediate detected (10 nt) is in accordance with the minimum length of RNA primers detected in vitro ).

    Journal: EMBO Reports

    Article Title: Identification of short 'eukaryotic' Okazaki fragments synthesized from a prokaryotic replication origin

    doi: 10.1038/sj.embor.embor732

    Figure Lengend Snippet: Direct labelling of RNA on the 5′ termini of replication intermediates with vaccinia virus guanylyltransferase, an enzyme responsible for messenger RNA capping. ( A ) The 5′ ends of RNA primers attached to P. abyssi replication intermediates were capped with radioactive GTP and samples were fractionated on a sequencing gel (lane 2). Lane 3 contains the same reaction as lane 2, except that total DNA was treated with the restriction endonuclease Eco RI before analysis, confirming that nascent strands were efficiently denatured from template strands. Lane 1, size marker (10-base ladder; Gibco-BRL) labelled with [γ- 32 P]ATP using T4 polynucleotide kinase. ( B ) Schematic diagram of the archaeal Okazaki fragment. The smallest replication intermediate detected (10 nt) is in accordance with the minimum length of RNA primers detected in vitro ).

    Article Snippet: All 5′-OH termini existing in denatured DNA were first 'masked' by treatment for 1 h with 8 units of T4 polynucleotide kinase (PNK; Promega) and non-radioactive ATP at 37 °C in the buffer supplied by the manufacturer.

    Techniques: Sequencing, Marker, In Vitro

    Unmasking assay showing short replication intermediates in Pyrococcus abyssi and Sulfolobus acidocaldarius . The 5′-OH groups of RNA-primed replication intermediates were selectively exposed by alkaline treatment. 'Unmasked' 5′-OH ends of replication intermediates were labelled with T4 polynucleotide kinase and [γ- 32 P]ATP, and run on an alkaline agarose gel. A control experiment without alkaline treatment is also shown for each experiment. The positions of size markers (100, 200 and 500 nucleotides (nt)) are shown on the left.

    Journal: EMBO Reports

    Article Title: Identification of short 'eukaryotic' Okazaki fragments synthesized from a prokaryotic replication origin

    doi: 10.1038/sj.embor.embor732

    Figure Lengend Snippet: Unmasking assay showing short replication intermediates in Pyrococcus abyssi and Sulfolobus acidocaldarius . The 5′-OH groups of RNA-primed replication intermediates were selectively exposed by alkaline treatment. 'Unmasked' 5′-OH ends of replication intermediates were labelled with T4 polynucleotide kinase and [γ- 32 P]ATP, and run on an alkaline agarose gel. A control experiment without alkaline treatment is also shown for each experiment. The positions of size markers (100, 200 and 500 nucleotides (nt)) are shown on the left.

    Article Snippet: All 5′-OH termini existing in denatured DNA were first 'masked' by treatment for 1 h with 8 units of T4 polynucleotide kinase (PNK; Promega) and non-radioactive ATP at 37 °C in the buffer supplied by the manufacturer.

    Techniques: Agarose Gel Electrophoresis

    Time course of BER. 35-μl reactions were performed as described under “Experimental Procedures” with UL2, APE, and UL30 in the presence of either T4 ligase (0.001 units/μl) ( A ), ligase I ( B ), or ligase IIIα-XRCC1 ( C ). 5-μl aliquots were removed at the times indicated. Activity is expressed as the fraction of maximum for the nicked ( N ) (●) and ligation ( L ) (○) products. The insets in each panel show the relevant gel images.

    Journal: The Journal of Biological Chemistry

    Article Title: Reconstitution of Uracil DNA Glycosylase-initiated Base Excision Repair in Herpes Simplex Virus-1 *

    doi: 10.1074/jbc.M109.010413

    Figure Lengend Snippet: Time course of BER. 35-μl reactions were performed as described under “Experimental Procedures” with UL2, APE, and UL30 in the presence of either T4 ligase (0.001 units/μl) ( A ), ligase I ( B ), or ligase IIIα-XRCC1 ( C ). 5-μl aliquots were removed at the times indicated. Activity is expressed as the fraction of maximum for the nicked ( N ) (●) and ligation ( L ) (○) products. The insets in each panel show the relevant gel images.

    Article Snippet: Unless otherwise stated, reactions (10 μl) contained 10 n m (molecules) DNA substrate in 20 m m HEPES-NaOH, pH 7.5, 100 μg/ml bovine serum albumin, 10% glycerol, 5 m m MgCl2 , 4 m m ATP, 4 μ m [α-32 P]dCTP (∼60 Ci/mmol), and the following proteins as indicated: UL2 (200 n m ), E. coli UDG (1 unit), APE (1.25 units), UL30 (100 n m ), UL42 (100 n m ), T4 ligase (Promega), ligase I (50 n m ), ligase IIIα-XRCC1 (50 n m ), Pol β (100 n m ), Pol δ (400 n m ), or exonuclease-deficient Klenow Pol (1 unit).

    Techniques: Activity Assay, Ligation

    Completion of BER; formation of ligation product. Reactions were performed as described under “Experimental Procedures” with the indicated proteins. Storage phosphorimage of reaction products obtained with UL2, APE, and UL30 ( lane 1 ) and identical reactions supplemented with T4 ligase (1.5 units) ( lane 2 ), ligase I ( lane 3 ), or ligase IIIα-XRCC1 ( lane 4 ) are shown. Lane 5 , DNA only; lane 6 , linear 100-mer. The positions of nicked product ( N ), ligated product ( L ), and of the 100-mer are as indicated. The values in italics below the lane numbers indicate the L/N ratios.

    Journal: The Journal of Biological Chemistry

    Article Title: Reconstitution of Uracil DNA Glycosylase-initiated Base Excision Repair in Herpes Simplex Virus-1 *

    doi: 10.1074/jbc.M109.010413

    Figure Lengend Snippet: Completion of BER; formation of ligation product. Reactions were performed as described under “Experimental Procedures” with the indicated proteins. Storage phosphorimage of reaction products obtained with UL2, APE, and UL30 ( lane 1 ) and identical reactions supplemented with T4 ligase (1.5 units) ( lane 2 ), ligase I ( lane 3 ), or ligase IIIα-XRCC1 ( lane 4 ) are shown. Lane 5 , DNA only; lane 6 , linear 100-mer. The positions of nicked product ( N ), ligated product ( L ), and of the 100-mer are as indicated. The values in italics below the lane numbers indicate the L/N ratios.

    Article Snippet: Unless otherwise stated, reactions (10 μl) contained 10 n m (molecules) DNA substrate in 20 m m HEPES-NaOH, pH 7.5, 100 μg/ml bovine serum albumin, 10% glycerol, 5 m m MgCl2 , 4 m m ATP, 4 μ m [α-32 P]dCTP (∼60 Ci/mmol), and the following proteins as indicated: UL2 (200 n m ), E. coli UDG (1 unit), APE (1.25 units), UL30 (100 n m ), UL42 (100 n m ), T4 ligase (Promega), ligase I (50 n m ), ligase IIIα-XRCC1 (50 n m ), Pol β (100 n m ), Pol δ (400 n m ), or exonuclease-deficient Klenow Pol (1 unit).

    Techniques: Ligation

    Product formation is dependent on UL2, APE, UL30, and ligase. Storage phosphorimages of reactions performed as described under “Experimental Procedures” with the indicated components. Lane 1 , UL2; lane 2 , APE; lane 3 ; UL30; lane 4 , T4 ligase (1.5 units); lane 5 , ligase I; lane 6 , ligase IIIα-XRCC1; lane 7 , UL2 and APE; lane 8 , APE and UL30; lane 9 , UL2 and UL30; lane 10 , UL2, APE and UL30; lane 11 , UL2, APE, UL30 and T4 ligase (1.5 units); lane 12 , UL2, APE, UL30, and ligase I; lane 13 , UL2, APE, UL30, and ligase IIIα-XRCC1; lane 14 , DNA only. The positions of nicked ( N ) and ligated ( L ) products are as indicated.

    Journal: The Journal of Biological Chemistry

    Article Title: Reconstitution of Uracil DNA Glycosylase-initiated Base Excision Repair in Herpes Simplex Virus-1 *

    doi: 10.1074/jbc.M109.010413

    Figure Lengend Snippet: Product formation is dependent on UL2, APE, UL30, and ligase. Storage phosphorimages of reactions performed as described under “Experimental Procedures” with the indicated components. Lane 1 , UL2; lane 2 , APE; lane 3 ; UL30; lane 4 , T4 ligase (1.5 units); lane 5 , ligase I; lane 6 , ligase IIIα-XRCC1; lane 7 , UL2 and APE; lane 8 , APE and UL30; lane 9 , UL2 and UL30; lane 10 , UL2, APE and UL30; lane 11 , UL2, APE, UL30 and T4 ligase (1.5 units); lane 12 , UL2, APE, UL30, and ligase I; lane 13 , UL2, APE, UL30, and ligase IIIα-XRCC1; lane 14 , DNA only. The positions of nicked ( N ) and ligated ( L ) products are as indicated.

    Article Snippet: Unless otherwise stated, reactions (10 μl) contained 10 n m (molecules) DNA substrate in 20 m m HEPES-NaOH, pH 7.5, 100 μg/ml bovine serum albumin, 10% glycerol, 5 m m MgCl2 , 4 m m ATP, 4 μ m [α-32 P]dCTP (∼60 Ci/mmol), and the following proteins as indicated: UL2 (200 n m ), E. coli UDG (1 unit), APE (1.25 units), UL30 (100 n m ), UL42 (100 n m ), T4 ligase (Promega), ligase I (50 n m ), ligase IIIα-XRCC1 (50 n m ), Pol β (100 n m ), Pol δ (400 n m ), or exonuclease-deficient Klenow Pol (1 unit).

    Techniques:

    T4 DNA-ligase activity in the presence of SA. T4 DNA-ligase was treated or not with increasing SA concentrations (15 min at 25°C) and then incubated at 37°C for 1 min with the oligo substrate. ( A ) The oligo(dT) 16 multimers were separated in polyacrylamide/urea gels: T4 DNA-ligase without SA (lane 1) or incubated with increasing SA concentrations of 2.5(2), 5(3), 10(4), and 20(5) μM. ( B ) The activity was quantitated using an InstantImager (Packard). ( C ) Inhibition of enzyme-adenylate formation by SA. T4 DNA-ligase was incubated or not with increasing SA concentrations (15 min at 25°C) before the addition of [α- 32 P]ATP. The enzyme adenylate complexes were separated by electrophoresis and detected by autoradiography.

    Journal: Proceedings of the National Academy of Sciences of the United States of America

    Article Title: Deficient DNA-ligase activity in the metabolic disease tyrosinemia type I

    doi:

    Figure Lengend Snippet: T4 DNA-ligase activity in the presence of SA. T4 DNA-ligase was treated or not with increasing SA concentrations (15 min at 25°C) and then incubated at 37°C for 1 min with the oligo substrate. ( A ) The oligo(dT) 16 multimers were separated in polyacrylamide/urea gels: T4 DNA-ligase without SA (lane 1) or incubated with increasing SA concentrations of 2.5(2), 5(3), 10(4), and 20(5) μM. ( B ) The activity was quantitated using an InstantImager (Packard). ( C ) Inhibition of enzyme-adenylate formation by SA. T4 DNA-ligase was incubated or not with increasing SA concentrations (15 min at 25°C) before the addition of [α- 32 P]ATP. The enzyme adenylate complexes were separated by electrophoresis and detected by autoradiography.

    Article Snippet: T4 DNA-ligase was from Boehringer Mannheim.

    Techniques: Activity Assay, Incubation, Inhibition, Electrophoresis, Autoradiography