t4 dna polymerase Roche Search Results


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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 49148 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/t4 dna ligase/product/New England Biolabs
    Average 99 stars, based on 49148 article reviews
    Price from $9.99 to $1999.99
    t4 dna ligase - by Bioz Stars, 2020-08
    99/100 stars
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    92
    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: 92/100, based on 906 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Average 92 stars, based on 906 article reviews
    Price from $9.99 to $1999.99
    t4 ligase - by Bioz Stars, 2020-08
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    92
    Roche t4 dna polymerase
    Preparation of vector and inserts for LIC. (A) A schematic of the vector pEKD1024 showing the major steps to generate single stranded DNA ends for LIC using <t>T4</t> DNA polymerase and dGTP. Restriction enzyme sites are shaded and the nucleotide where the T4 DNA polymerase stops is indicated by white text over a dark background. (B, C, D) Schematics of various types of inserts that can be cloned into the vector pEKD1024. (B) A PCR amplified insert and the product after treatment with T4 DNA polymerase in the presence of dCTP. (C, D) Inserts formed with oligonucleotides. (C) Complete overlap of inserted sequences. (D) Partially randomized sequence with 13 nucleotide overlap.
    T4 Dna Polymerase, supplied by Roche, used in various techniques. Bioz Stars score: 92/100, based on 481 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Average 92 stars, based on 481 article reviews
    Price from $9.99 to $1999.99
    t4 dna polymerase - by Bioz Stars, 2020-08
    92/100 stars
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    92
    Roche t4 rna polymerase
    Preparation of vector and inserts for LIC. (A) A schematic of the vector pEKD1024 showing the major steps to generate single stranded DNA ends for LIC using <t>T4</t> DNA polymerase and dGTP. Restriction enzyme sites are shaded and the nucleotide where the T4 DNA polymerase stops is indicated by white text over a dark background. (B, C, D) Schematics of various types of inserts that can be cloned into the vector pEKD1024. (B) A PCR amplified insert and the product after treatment with T4 DNA polymerase in the presence of dCTP. (C, D) Inserts formed with oligonucleotides. (C) Complete overlap of inserted sequences. (D) Partially randomized sequence with 13 nucleotide overlap.
    T4 Rna Polymerase, supplied by Roche, used in various techniques. Bioz Stars score: 92/100, based on 9 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Average 92 stars, based on 9 article reviews
    Price from $9.99 to $1999.99
    t4 rna polymerase - by Bioz Stars, 2020-08
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    92
    Roche t4 ligase buffer
    Preparation of vector and inserts for LIC. (A) A schematic of the vector pEKD1024 showing the major steps to generate single stranded DNA ends for LIC using <t>T4</t> DNA polymerase and dGTP. Restriction enzyme sites are shaded and the nucleotide where the T4 DNA polymerase stops is indicated by white text over a dark background. (B, C, D) Schematics of various types of inserts that can be cloned into the vector pEKD1024. (B) A PCR amplified insert and the product after treatment with T4 DNA polymerase in the presence of dCTP. (C, D) Inserts formed with oligonucleotides. (C) Complete overlap of inserted sequences. (D) Partially randomized sequence with 13 nucleotide overlap.
    T4 Ligase Buffer, supplied by Roche, used in various techniques. Bioz Stars score: 92/100, based on 33 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Average 92 stars, based on 33 article reviews
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    t4 ligase buffer - by Bioz Stars, 2020-08
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    94
    Roche t4 dna ligase
    Construction of DNA control plasmid designed for the 4 CDC Category A DNA agents (Smallpox virus [seq1], Bacillus anthracis [seq2], Francisella tularensis [seq3], and Yersinia pestis [seq4]). Assembling of the smallpox virus and B. anthracis sequences is presented as an example. Successive steps are indicated by framed numbers. 1, PCR amplification of the two matrix sequences by primers consisting of the stabilization and the restriction site sequences (italics). PCR reactions were carried out in a volume of 50 μl that included 10 mM Tris-HCl [pH 9.0], 1.5 mM MgCl2, 50 mM KCl, 0.1% Triton X-100, 200 μM each dNTP, 0.4 μM of each oligonucleotide primer, 0.4 μM of the single stranded DNA, and 1.5 U of Taq DNA polymerase (Invitrogen, Cergy-Pontoise, France). The thermocycler (Biometra, Göttingen, Germany) profile was 5 min at 95°C, followed by 35 cycles of 30 sec at 95°C, 30 sec at 55°C, and 1 min at 72°C, and terminated by a final extension for 7 min at 72°C. PCR products were electrophorezed in 3% TAE-agarose gel containing ethidium bromide and visualized under UV transillumination. Column purification of the PCR products. PCR products of the expected size were column-purified by using the QIAquick PCR Purification Kit (Qiagen, Hilden, Germany) according to the manufacturer's instructions, and eluted in 50 μl of RNase free distillated water. When two bands or more were observed by gel analysis, the band of expected size was excised from the gel and purified by glass milk extraction with the GenClean III Kit (Q-Bio-Gene, Carlsbad CA, USA). 2, assemblage was conducted by pair, seq1 with seq2 (resulting in seq1-2). Equal volumes (10 μl) of purified seq1- and seq2-dsDNA were incubated at 37°C in the presence of Sac I . Sac I site is located at the 3' and 5' ends of seq1 and seq2, respectively. 3, the reaction product was column purified using the protocol aforementioned to discard the 15-nt DNA fragments corresponding to the 5' and 3' ends to avoid their re-ligation to their respective complementary sequences at step 5. 4, Overnight incubation at 4°C in the presence of <t>T4</t> DNA ligase. Ten μl of the reaction was incubated with T4 DNA ligase (Roche, Basel, Switzerland) according to the manufaturer's instructions. 5, PCR amplification by using the external primers (italics) was performed according to the protocol described at step 1. Then column purification using the protocol detailed at step 2 of the resulting PCR product. At this step the seq1-2 PCR product may be cloned into PGEM-T for storage. The same procedure was performed for seq3 and seq4. Ultimately, seq1-2 and seq3-4 were assembled by using the same protocol (sections 1–9). The final product cloned into PGEM-T plasmid includes seq1-2-3-4 flanked by the two Sseq and restriction sites.
    T4 Dna Ligase, supplied by Roche, used in various techniques. Bioz Stars score: 94/100, based on 5002 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Average 94 stars, based on 5002 article reviews
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    t4 dna ligase - by Bioz Stars, 2020-08
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    91
    Roche 2x t4 ligase buffer
    Construction of DNA control plasmid designed for the 4 CDC Category A DNA agents (Smallpox virus [seq1], Bacillus anthracis [seq2], Francisella tularensis [seq3], and Yersinia pestis [seq4]). Assembling of the smallpox virus and B. anthracis sequences is presented as an example. Successive steps are indicated by framed numbers. 1, PCR amplification of the two matrix sequences by primers consisting of the stabilization and the restriction site sequences (italics). PCR reactions were carried out in a volume of 50 μl that included 10 mM Tris-HCl [pH 9.0], 1.5 mM MgCl2, 50 mM KCl, 0.1% Triton X-100, 200 μM each dNTP, 0.4 μM of each oligonucleotide primer, 0.4 μM of the single stranded DNA, and 1.5 U of Taq DNA polymerase (Invitrogen, Cergy-Pontoise, France). The thermocycler (Biometra, Göttingen, Germany) profile was 5 min at 95°C, followed by 35 cycles of 30 sec at 95°C, 30 sec at 55°C, and 1 min at 72°C, and terminated by a final extension for 7 min at 72°C. PCR products were electrophorezed in 3% TAE-agarose gel containing ethidium bromide and visualized under UV transillumination. Column purification of the PCR products. PCR products of the expected size were column-purified by using the QIAquick PCR Purification Kit (Qiagen, Hilden, Germany) according to the manufacturer's instructions, and eluted in 50 μl of RNase free distillated water. When two bands or more were observed by gel analysis, the band of expected size was excised from the gel and purified by glass milk extraction with the GenClean III Kit (Q-Bio-Gene, Carlsbad CA, USA). 2, assemblage was conducted by pair, seq1 with seq2 (resulting in seq1-2). Equal volumes (10 μl) of purified seq1- and seq2-dsDNA were incubated at 37°C in the presence of Sac I . Sac I site is located at the 3' and 5' ends of seq1 and seq2, respectively. 3, the reaction product was column purified using the protocol aforementioned to discard the 15-nt DNA fragments corresponding to the 5' and 3' ends to avoid their re-ligation to their respective complementary sequences at step 5. 4, Overnight incubation at 4°C in the presence of <t>T4</t> DNA ligase. Ten μl of the reaction was incubated with T4 DNA ligase (Roche, Basel, Switzerland) according to the manufaturer's instructions. 5, PCR amplification by using the external primers (italics) was performed according to the protocol described at step 1. Then column purification using the protocol detailed at step 2 of the resulting PCR product. At this step the seq1-2 PCR product may be cloned into PGEM-T for storage. The same procedure was performed for seq3 and seq4. Ultimately, seq1-2 and seq3-4 were assembled by using the same protocol (sections 1–9). The final product cloned into PGEM-T plasmid includes seq1-2-3-4 flanked by the two Sseq and restriction sites.
    2x T4 Ligase Buffer, supplied by Roche, used in various techniques. Bioz Stars score: 91/100, based on 8 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Average 91 stars, based on 8 article reviews
    Price from $9.99 to $1999.99
    2x t4 ligase buffer - by Bioz Stars, 2020-08
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    94
    Roche t4 polynucleotide kinase
    Construction of DNA control plasmid designed for the 4 CDC Category A DNA agents (Smallpox virus [seq1], Bacillus anthracis [seq2], Francisella tularensis [seq3], and Yersinia pestis [seq4]). Assembling of the smallpox virus and B. anthracis sequences is presented as an example. Successive steps are indicated by framed numbers. 1, PCR amplification of the two matrix sequences by primers consisting of the stabilization and the restriction site sequences (italics). PCR reactions were carried out in a volume of 50 μl that included 10 mM Tris-HCl [pH 9.0], 1.5 mM MgCl2, 50 mM KCl, 0.1% Triton X-100, 200 μM each dNTP, 0.4 μM of each oligonucleotide primer, 0.4 μM of the single stranded DNA, and 1.5 U of Taq DNA polymerase (Invitrogen, Cergy-Pontoise, France). The thermocycler (Biometra, Göttingen, Germany) profile was 5 min at 95°C, followed by 35 cycles of 30 sec at 95°C, 30 sec at 55°C, and 1 min at 72°C, and terminated by a final extension for 7 min at 72°C. PCR products were electrophorezed in 3% TAE-agarose gel containing ethidium bromide and visualized under UV transillumination. Column purification of the PCR products. PCR products of the expected size were column-purified by using the QIAquick PCR Purification Kit (Qiagen, Hilden, Germany) according to the manufacturer's instructions, and eluted in 50 μl of RNase free distillated water. When two bands or more were observed by gel analysis, the band of expected size was excised from the gel and purified by glass milk extraction with the GenClean III Kit (Q-Bio-Gene, Carlsbad CA, USA). 2, assemblage was conducted by pair, seq1 with seq2 (resulting in seq1-2). Equal volumes (10 μl) of purified seq1- and seq2-dsDNA were incubated at 37°C in the presence of Sac I . Sac I site is located at the 3' and 5' ends of seq1 and seq2, respectively. 3, the reaction product was column purified using the protocol aforementioned to discard the 15-nt DNA fragments corresponding to the 5' and 3' ends to avoid their re-ligation to their respective complementary sequences at step 5. 4, Overnight incubation at 4°C in the presence of <t>T4</t> DNA ligase. Ten μl of the reaction was incubated with T4 DNA ligase (Roche, Basel, Switzerland) according to the manufaturer's instructions. 5, PCR amplification by using the external primers (italics) was performed according to the protocol described at step 1. Then column purification using the protocol detailed at step 2 of the resulting PCR product. At this step the seq1-2 PCR product may be cloned into PGEM-T for storage. The same procedure was performed for seq3 and seq4. Ultimately, seq1-2 and seq3-4 were assembled by using the same protocol (sections 1–9). The final product cloned into PGEM-T plasmid includes seq1-2-3-4 flanked by the two Sseq and restriction sites.
    T4 Polynucleotide Kinase, supplied by Roche, used in various techniques. Bioz Stars score: 94/100, based on 1666 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Average 94 stars, based on 1666 article reviews
    Price from $9.99 to $1999.99
    t4 polynucleotide kinase - by Bioz Stars, 2020-08
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    85
    Roche enzyme t4 ligase
    Construction of DNA control plasmid designed for the 4 CDC Category A DNA agents (Smallpox virus [seq1], Bacillus anthracis [seq2], Francisella tularensis [seq3], and Yersinia pestis [seq4]). Assembling of the smallpox virus and B. anthracis sequences is presented as an example. Successive steps are indicated by framed numbers. 1, PCR amplification of the two matrix sequences by primers consisting of the stabilization and the restriction site sequences (italics). PCR reactions were carried out in a volume of 50 μl that included 10 mM Tris-HCl [pH 9.0], 1.5 mM MgCl2, 50 mM KCl, 0.1% Triton X-100, 200 μM each dNTP, 0.4 μM of each oligonucleotide primer, 0.4 μM of the single stranded DNA, and 1.5 U of Taq DNA polymerase (Invitrogen, Cergy-Pontoise, France). The thermocycler (Biometra, Göttingen, Germany) profile was 5 min at 95°C, followed by 35 cycles of 30 sec at 95°C, 30 sec at 55°C, and 1 min at 72°C, and terminated by a final extension for 7 min at 72°C. PCR products were electrophorezed in 3% TAE-agarose gel containing ethidium bromide and visualized under UV transillumination. Column purification of the PCR products. PCR products of the expected size were column-purified by using the QIAquick PCR Purification Kit (Qiagen, Hilden, Germany) according to the manufacturer's instructions, and eluted in 50 μl of RNase free distillated water. When two bands or more were observed by gel analysis, the band of expected size was excised from the gel and purified by glass milk extraction with the GenClean III Kit (Q-Bio-Gene, Carlsbad CA, USA). 2, assemblage was conducted by pair, seq1 with seq2 (resulting in seq1-2). Equal volumes (10 μl) of purified seq1- and seq2-dsDNA were incubated at 37°C in the presence of Sac I . Sac I site is located at the 3' and 5' ends of seq1 and seq2, respectively. 3, the reaction product was column purified using the protocol aforementioned to discard the 15-nt DNA fragments corresponding to the 5' and 3' ends to avoid their re-ligation to their respective complementary sequences at step 5. 4, Overnight incubation at 4°C in the presence of <t>T4</t> DNA ligase. Ten μl of the reaction was incubated with T4 DNA ligase (Roche, Basel, Switzerland) according to the manufaturer's instructions. 5, PCR amplification by using the external primers (italics) was performed according to the protocol described at step 1. Then column purification using the protocol detailed at step 2 of the resulting PCR product. At this step the seq1-2 PCR product may be cloned into PGEM-T for storage. The same procedure was performed for seq3 and seq4. Ultimately, seq1-2 and seq3-4 were assembled by using the same protocol (sections 1–9). The final product cloned into PGEM-T plasmid includes seq1-2-3-4 flanked by the two Sseq and restriction sites.
    Enzyme T4 Ligase, supplied by Roche, used in various techniques. Bioz Stars score: 85/100, based on 5 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/enzyme t4 ligase/product/Roche
    Average 85 stars, based on 5 article reviews
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    enzyme t4 ligase - by Bioz Stars, 2020-08
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    88
    Roche dpni fragments
    Construction of DNA control plasmid designed for the 4 CDC Category A DNA agents (Smallpox virus [seq1], Bacillus anthracis [seq2], Francisella tularensis [seq3], and Yersinia pestis [seq4]). Assembling of the smallpox virus and B. anthracis sequences is presented as an example. Successive steps are indicated by framed numbers. 1, PCR amplification of the two matrix sequences by primers consisting of the stabilization and the restriction site sequences (italics). PCR reactions were carried out in a volume of 50 μl that included 10 mM Tris-HCl [pH 9.0], 1.5 mM MgCl2, 50 mM KCl, 0.1% Triton X-100, 200 μM each dNTP, 0.4 μM of each oligonucleotide primer, 0.4 μM of the single stranded DNA, and 1.5 U of Taq DNA polymerase (Invitrogen, Cergy-Pontoise, France). The thermocycler (Biometra, Göttingen, Germany) profile was 5 min at 95°C, followed by 35 cycles of 30 sec at 95°C, 30 sec at 55°C, and 1 min at 72°C, and terminated by a final extension for 7 min at 72°C. PCR products were electrophorezed in 3% TAE-agarose gel containing ethidium bromide and visualized under UV transillumination. Column purification of the PCR products. PCR products of the expected size were column-purified by using the QIAquick PCR Purification Kit (Qiagen, Hilden, Germany) according to the manufacturer's instructions, and eluted in 50 μl of RNase free distillated water. When two bands or more were observed by gel analysis, the band of expected size was excised from the gel and purified by glass milk extraction with the GenClean III Kit (Q-Bio-Gene, Carlsbad CA, USA). 2, assemblage was conducted by pair, seq1 with seq2 (resulting in seq1-2). Equal volumes (10 μl) of purified seq1- and seq2-dsDNA were incubated at 37°C in the presence of Sac I . Sac I site is located at the 3' and 5' ends of seq1 and seq2, respectively. 3, the reaction product was column purified using the protocol aforementioned to discard the 15-nt DNA fragments corresponding to the 5' and 3' ends to avoid their re-ligation to their respective complementary sequences at step 5. 4, Overnight incubation at 4°C in the presence of <t>T4</t> DNA ligase. Ten μl of the reaction was incubated with T4 DNA ligase (Roche, Basel, Switzerland) according to the manufaturer's instructions. 5, PCR amplification by using the external primers (italics) was performed according to the protocol described at step 1. Then column purification using the protocol detailed at step 2 of the resulting PCR product. At this step the seq1-2 PCR product may be cloned into PGEM-T for storage. The same procedure was performed for seq3 and seq4. Ultimately, seq1-2 and seq3-4 were assembled by using the same protocol (sections 1–9). The final product cloned into PGEM-T plasmid includes seq1-2-3-4 flanked by the two Sseq and restriction sites.
    Dpni Fragments, supplied by Roche, used in various techniques. Bioz Stars score: 88/100, based on 21 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Average 88 stars, based on 21 article reviews
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    91
    Roche on column dnase i
    Construction of DNA control plasmid designed for the 4 CDC Category A DNA agents (Smallpox virus [seq1], Bacillus anthracis [seq2], Francisella tularensis [seq3], and Yersinia pestis [seq4]). Assembling of the smallpox virus and B. anthracis sequences is presented as an example. Successive steps are indicated by framed numbers. 1, PCR amplification of the two matrix sequences by primers consisting of the stabilization and the restriction site sequences (italics). PCR reactions were carried out in a volume of 50 μl that included 10 mM Tris-HCl [pH 9.0], 1.5 mM MgCl2, 50 mM KCl, 0.1% Triton X-100, 200 μM each dNTP, 0.4 μM of each oligonucleotide primer, 0.4 μM of the single stranded DNA, and 1.5 U of Taq DNA polymerase (Invitrogen, Cergy-Pontoise, France). The thermocycler (Biometra, Göttingen, Germany) profile was 5 min at 95°C, followed by 35 cycles of 30 sec at 95°C, 30 sec at 55°C, and 1 min at 72°C, and terminated by a final extension for 7 min at 72°C. PCR products were electrophorezed in 3% TAE-agarose gel containing ethidium bromide and visualized under UV transillumination. Column purification of the PCR products. PCR products of the expected size were column-purified by using the QIAquick PCR Purification Kit (Qiagen, Hilden, Germany) according to the manufacturer's instructions, and eluted in 50 μl of RNase free distillated water. When two bands or more were observed by gel analysis, the band of expected size was excised from the gel and purified by glass milk extraction with the GenClean III Kit (Q-Bio-Gene, Carlsbad CA, USA). 2, assemblage was conducted by pair, seq1 with seq2 (resulting in seq1-2). Equal volumes (10 μl) of purified seq1- and seq2-dsDNA were incubated at 37°C in the presence of Sac I . Sac I site is located at the 3' and 5' ends of seq1 and seq2, respectively. 3, the reaction product was column purified using the protocol aforementioned to discard the 15-nt DNA fragments corresponding to the 5' and 3' ends to avoid their re-ligation to their respective complementary sequences at step 5. 4, Overnight incubation at 4°C in the presence of <t>T4</t> DNA ligase. Ten μl of the reaction was incubated with T4 DNA ligase (Roche, Basel, Switzerland) according to the manufaturer's instructions. 5, PCR amplification by using the external primers (italics) was performed according to the protocol described at step 1. Then column purification using the protocol detailed at step 2 of the resulting PCR product. At this step the seq1-2 PCR product may be cloned into PGEM-T for storage. The same procedure was performed for seq3 and seq4. Ultimately, seq1-2 and seq3-4 were assembled by using the same protocol (sections 1–9). The final product cloned into PGEM-T plasmid includes seq1-2-3-4 flanked by the two Sseq and restriction sites.
    On Column Dnase I, supplied by Roche, used in various techniques. Bioz Stars score: 91/100, based on 9 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    94
    Roche dnase i
    DNase I-PCR Assay of the <t>DNase</t> I Sensitivity of a Transgenic Enhancer Sequence. (A) . (B) A diagram illustrating the design of a PCR primer that allows the transgenic L3 locus, but not the endogenous L3, to be specifically amplified. (C) DNase I-PCR shows differential DNase I sensitivity of chromatin in selected genomic regions. The DNase I sensitivity of the transgenic L3 locus was similar to that of the open chromatin control ACTIN7 .
    Dnase I, supplied by Roche, used in various techniques. Bioz Stars score: 94/100, based on 33815 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    85
    Roche 11004760001 t4 dna ligase
    DNase I-PCR Assay of the <t>DNase</t> I Sensitivity of a Transgenic Enhancer Sequence. (A) . (B) A diagram illustrating the design of a PCR primer that allows the transgenic L3 locus, but not the endogenous L3, to be specifically amplified. (C) DNase I-PCR shows differential DNase I sensitivity of chromatin in selected genomic regions. The DNase I sensitivity of the transgenic L3 locus was similar to that of the open chromatin control ACTIN7 .
    11004760001 T4 Dna Ligase, supplied by Roche, used in various techniques. Bioz Stars score: 85/100, based on 5 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    91
    Roche dnase i recombinant
    DNase I-PCR Assay of the <t>DNase</t> I Sensitivity of a Transgenic Enhancer Sequence. (A) . (B) A diagram illustrating the design of a PCR primer that allows the transgenic L3 locus, but not the endogenous L3, to be specifically amplified. (C) DNase I-PCR shows differential DNase I sensitivity of chromatin in selected genomic regions. The DNase I sensitivity of the transgenic L3 locus was similar to that of the open chromatin control ACTIN7 .
    Dnase I Recombinant, supplied by Roche, used in various techniques. Bioz Stars score: 91/100, based on 600 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    90
    Roche t4 dna ligase enzyme
    DNase I-PCR Assay of the <t>DNase</t> I Sensitivity of a Transgenic Enhancer Sequence. (A) . (B) A diagram illustrating the design of a PCR primer that allows the transgenic L3 locus, but not the endogenous L3, to be specifically amplified. (C) DNase I-PCR shows differential DNase I sensitivity of chromatin in selected genomic regions. The DNase I sensitivity of the transgenic L3 locus was similar to that of the open chromatin control ACTIN7 .
    T4 Dna Ligase Enzyme, supplied by Roche, used in various techniques. Bioz Stars score: 90/100, based on 17 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    92
    Roche t4 dna ligase buffer
    DNase I-PCR Assay of the <t>DNase</t> I Sensitivity of a Transgenic Enhancer Sequence. (A) . (B) A diagram illustrating the design of a PCR primer that allows the transgenic L3 locus, but not the endogenous L3, to be specifically amplified. (C) DNase I-PCR shows differential DNase I sensitivity of chromatin in selected genomic regions. The DNase I sensitivity of the transgenic L3 locus was similar to that of the open chromatin control ACTIN7 .
    T4 Dna Ligase Buffer, supplied by Roche, used in various techniques. Bioz Stars score: 92/100, based on 26 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    90
    Roche thermo scientific dnase i
    DNase I-PCR Assay of the <t>DNase</t> I Sensitivity of a Transgenic Enhancer Sequence. (A) . (B) A diagram illustrating the design of a PCR primer that allows the transgenic L3 locus, but not the endogenous L3, to be specifically amplified. (C) DNase I-PCR shows differential DNase I sensitivity of chromatin in selected genomic regions. The DNase I sensitivity of the transgenic L3 locus was similar to that of the open chromatin control ACTIN7 .
    Thermo Scientific Dnase I, supplied by Roche, used in various techniques. Bioz Stars score: 90/100, based on 10 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    93
    Roche dnase i treatment
    DNase I-PCR Assay of the <t>DNase</t> I Sensitivity of a Transgenic Enhancer Sequence. (A) . (B) A diagram illustrating the design of a PCR primer that allows the transgenic L3 locus, but not the endogenous L3, to be specifically amplified. (C) DNase I-PCR shows differential DNase I sensitivity of chromatin in selected genomic regions. The DNase I sensitivity of the transgenic L3 locus was similar to that of the open chromatin control ACTIN7 .
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    Analysis of transcript levels in 7 h germlings. A. parasiticus SU-1, AFS10, and Δ veA were grown on YES solid media and spores were collected at 5 days. Germlings were grown from fresh spores in GMS liquid medium for 7 h, frozen in liquid nitrogen, and stored at −80 °C. RNA was extracted from germlings using grinding and sonication as described in Methods. RT-PCR was performed on total RNA treated with <t>RNAse-free</t> <t>DNAse</t> I with primers specific for each gene ( Table 3 ). PCR products were separated on a 1% agarose gel by electrophoresis. Citrate synthase, a constitutively expressed gene, was used as a positive control. gDNA, genomic DNA.
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    Analysis of transcript levels in 7 h germlings. A. parasiticus SU-1, AFS10, and Δ veA were grown on YES solid media and spores were collected at 5 days. Germlings were grown from fresh spores in GMS liquid medium for 7 h, frozen in liquid nitrogen, and stored at −80 °C. RNA was extracted from germlings using grinding and sonication as described in Methods. RT-PCR was performed on total RNA treated with <t>RNAse-free</t> <t>DNAse</t> I with primers specific for each gene ( Table 3 ). PCR products were separated on a 1% agarose gel by electrophoresis. Citrate synthase, a constitutively expressed gene, was used as a positive control. gDNA, genomic DNA.
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    Analysis of transcript levels in 7 h germlings. A. parasiticus SU-1, AFS10, and Δ veA were grown on YES solid media and spores were collected at 5 days. Germlings were grown from fresh spores in GMS liquid medium for 7 h, frozen in liquid nitrogen, and stored at −80 °C. RNA was extracted from germlings using grinding and sonication as described in Methods. RT-PCR was performed on total RNA treated with <t>RNAse-free</t> <t>DNAse</t> I with primers specific for each gene ( Table 3 ). PCR products were separated on a 1% agarose gel by electrophoresis. Citrate synthase, a constitutively expressed gene, was used as a positive control. gDNA, genomic DNA.
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    Image Search Results


    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.

    Journal: Molecular and Cellular Biology

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

    doi: 10.1128/MCB.23.7.2309-2315.2003

    Figure Lengend 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.

    Article Snippet: One unit of T4 ligase (Roche) was equivalent to ∼350 μg of X4-LIV.

    Techniques: Cleavage Assay, Labeling, Ligation, Concentration Assay

    Preparation of vector and inserts for LIC. (A) A schematic of the vector pEKD1024 showing the major steps to generate single stranded DNA ends for LIC using T4 DNA polymerase and dGTP. Restriction enzyme sites are shaded and the nucleotide where the T4 DNA polymerase stops is indicated by white text over a dark background. (B, C, D) Schematics of various types of inserts that can be cloned into the vector pEKD1024. (B) A PCR amplified insert and the product after treatment with T4 DNA polymerase in the presence of dCTP. (C, D) Inserts formed with oligonucleotides. (C) Complete overlap of inserted sequences. (D) Partially randomized sequence with 13 nucleotide overlap.

    Journal: Methods in enzymology

    Article Title: RNA-ID, a Powerful Tool for Identifying and Characterizing Regulatory Sequences

    doi: 10.1016/bs.mie.2016.02.003

    Figure Lengend Snippet: Preparation of vector and inserts for LIC. (A) A schematic of the vector pEKD1024 showing the major steps to generate single stranded DNA ends for LIC using T4 DNA polymerase and dGTP. Restriction enzyme sites are shaded and the nucleotide where the T4 DNA polymerase stops is indicated by white text over a dark background. (B, C, D) Schematics of various types of inserts that can be cloned into the vector pEKD1024. (B) A PCR amplified insert and the product after treatment with T4 DNA polymerase in the presence of dCTP. (C, D) Inserts formed with oligonucleotides. (C) Complete overlap of inserted sequences. (D) Partially randomized sequence with 13 nucleotide overlap.

    Article Snippet: Incubate at 22 °C (room temperature) for 40 minutes, then inactivate the T4 DNA polymerase by incubating the reaction at 75 °C for 40 minutes.

    Techniques: Plasmid Preparation, Clone Assay, Polymerase Chain Reaction, Amplification, Sequencing

    Construction of DNA control plasmid designed for the 4 CDC Category A DNA agents (Smallpox virus [seq1], Bacillus anthracis [seq2], Francisella tularensis [seq3], and Yersinia pestis [seq4]). Assembling of the smallpox virus and B. anthracis sequences is presented as an example. Successive steps are indicated by framed numbers. 1, PCR amplification of the two matrix sequences by primers consisting of the stabilization and the restriction site sequences (italics). PCR reactions were carried out in a volume of 50 μl that included 10 mM Tris-HCl [pH 9.0], 1.5 mM MgCl2, 50 mM KCl, 0.1% Triton X-100, 200 μM each dNTP, 0.4 μM of each oligonucleotide primer, 0.4 μM of the single stranded DNA, and 1.5 U of Taq DNA polymerase (Invitrogen, Cergy-Pontoise, France). The thermocycler (Biometra, Göttingen, Germany) profile was 5 min at 95°C, followed by 35 cycles of 30 sec at 95°C, 30 sec at 55°C, and 1 min at 72°C, and terminated by a final extension for 7 min at 72°C. PCR products were electrophorezed in 3% TAE-agarose gel containing ethidium bromide and visualized under UV transillumination. Column purification of the PCR products. PCR products of the expected size were column-purified by using the QIAquick PCR Purification Kit (Qiagen, Hilden, Germany) according to the manufacturer's instructions, and eluted in 50 μl of RNase free distillated water. When two bands or more were observed by gel analysis, the band of expected size was excised from the gel and purified by glass milk extraction with the GenClean III Kit (Q-Bio-Gene, Carlsbad CA, USA). 2, assemblage was conducted by pair, seq1 with seq2 (resulting in seq1-2). Equal volumes (10 μl) of purified seq1- and seq2-dsDNA were incubated at 37°C in the presence of Sac I . Sac I site is located at the 3' and 5' ends of seq1 and seq2, respectively. 3, the reaction product was column purified using the protocol aforementioned to discard the 15-nt DNA fragments corresponding to the 5' and 3' ends to avoid their re-ligation to their respective complementary sequences at step 5. 4, Overnight incubation at 4°C in the presence of T4 DNA ligase. Ten μl of the reaction was incubated with T4 DNA ligase (Roche, Basel, Switzerland) according to the manufaturer's instructions. 5, PCR amplification by using the external primers (italics) was performed according to the protocol described at step 1. Then column purification using the protocol detailed at step 2 of the resulting PCR product. At this step the seq1-2 PCR product may be cloned into PGEM-T for storage. The same procedure was performed for seq3 and seq4. Ultimately, seq1-2 and seq3-4 were assembled by using the same protocol (sections 1–9). The final product cloned into PGEM-T plasmid includes seq1-2-3-4 flanked by the two Sseq and restriction sites.

    Journal: BMC Microbiology

    Article Title: Multi-pathogens sequence containing plasmids as positive controls for universal detection of potential agents of bioterrorism

    doi: 10.1186/1471-2180-4-21

    Figure Lengend Snippet: Construction of DNA control plasmid designed for the 4 CDC Category A DNA agents (Smallpox virus [seq1], Bacillus anthracis [seq2], Francisella tularensis [seq3], and Yersinia pestis [seq4]). Assembling of the smallpox virus and B. anthracis sequences is presented as an example. Successive steps are indicated by framed numbers. 1, PCR amplification of the two matrix sequences by primers consisting of the stabilization and the restriction site sequences (italics). PCR reactions were carried out in a volume of 50 μl that included 10 mM Tris-HCl [pH 9.0], 1.5 mM MgCl2, 50 mM KCl, 0.1% Triton X-100, 200 μM each dNTP, 0.4 μM of each oligonucleotide primer, 0.4 μM of the single stranded DNA, and 1.5 U of Taq DNA polymerase (Invitrogen, Cergy-Pontoise, France). The thermocycler (Biometra, Göttingen, Germany) profile was 5 min at 95°C, followed by 35 cycles of 30 sec at 95°C, 30 sec at 55°C, and 1 min at 72°C, and terminated by a final extension for 7 min at 72°C. PCR products were electrophorezed in 3% TAE-agarose gel containing ethidium bromide and visualized under UV transillumination. Column purification of the PCR products. PCR products of the expected size were column-purified by using the QIAquick PCR Purification Kit (Qiagen, Hilden, Germany) according to the manufacturer's instructions, and eluted in 50 μl of RNase free distillated water. When two bands or more were observed by gel analysis, the band of expected size was excised from the gel and purified by glass milk extraction with the GenClean III Kit (Q-Bio-Gene, Carlsbad CA, USA). 2, assemblage was conducted by pair, seq1 with seq2 (resulting in seq1-2). Equal volumes (10 μl) of purified seq1- and seq2-dsDNA were incubated at 37°C in the presence of Sac I . Sac I site is located at the 3' and 5' ends of seq1 and seq2, respectively. 3, the reaction product was column purified using the protocol aforementioned to discard the 15-nt DNA fragments corresponding to the 5' and 3' ends to avoid their re-ligation to their respective complementary sequences at step 5. 4, Overnight incubation at 4°C in the presence of T4 DNA ligase. Ten μl of the reaction was incubated with T4 DNA ligase (Roche, Basel, Switzerland) according to the manufaturer's instructions. 5, PCR amplification by using the external primers (italics) was performed according to the protocol described at step 1. Then column purification using the protocol detailed at step 2 of the resulting PCR product. At this step the seq1-2 PCR product may be cloned into PGEM-T for storage. The same procedure was performed for seq3 and seq4. Ultimately, seq1-2 and seq3-4 were assembled by using the same protocol (sections 1–9). The final product cloned into PGEM-T plasmid includes seq1-2-3-4 flanked by the two Sseq and restriction sites.

    Article Snippet: Ten μl of the reaction was incubated with T4 DNA ligase (Roche, Basel, Switzerland) according to the manufaturer's instructions.

    Techniques: Plasmid Preparation, Polymerase Chain Reaction, Amplification, Size-exclusion Chromatography, Agarose Gel Electrophoresis, Purification, Incubation, Ligation, Clone Assay

    DNase I-PCR Assay of the DNase I Sensitivity of a Transgenic Enhancer Sequence. (A) . (B) A diagram illustrating the design of a PCR primer that allows the transgenic L3 locus, but not the endogenous L3, to be specifically amplified. (C) DNase I-PCR shows differential DNase I sensitivity of chromatin in selected genomic regions. The DNase I sensitivity of the transgenic L3 locus was similar to that of the open chromatin control ACTIN7 .

    Journal: The Plant Cell

    Article Title: Genome-Wide Prediction and Validation of Intergenic Enhancers in Arabidopsis Using Open Chromatin Signatures [OPEN]

    doi: 10.1105/tpc.15.00537

    Figure Lengend Snippet: DNase I-PCR Assay of the DNase I Sensitivity of a Transgenic Enhancer Sequence. (A) . (B) A diagram illustrating the design of a PCR primer that allows the transgenic L3 locus, but not the endogenous L3, to be specifically amplified. (C) DNase I-PCR shows differential DNase I sensitivity of chromatin in selected genomic regions. The DNase I sensitivity of the transgenic L3 locus was similar to that of the open chromatin control ACTIN7 .

    Article Snippet: A DNase I (RNase-free; 10 U/μL; Roche Applied Science; catalog number 04716728001) dilution series was prepared by step-wise dilution using digestion buffer.

    Techniques: Polymerase Chain Reaction, Transgenic Assay, Sequencing, Amplification

    Analysis of transcript levels in 7 h germlings. A. parasiticus SU-1, AFS10, and Δ veA were grown on YES solid media and spores were collected at 5 days. Germlings were grown from fresh spores in GMS liquid medium for 7 h, frozen in liquid nitrogen, and stored at −80 °C. RNA was extracted from germlings using grinding and sonication as described in Methods. RT-PCR was performed on total RNA treated with RNAse-free DNAse I with primers specific for each gene ( Table 3 ). PCR products were separated on a 1% agarose gel by electrophoresis. Citrate synthase, a constitutively expressed gene, was used as a positive control. gDNA, genomic DNA.

    Journal: Toxins

    Article Title: Aflatoxin Biosynthesis Is a Novel Source of Reactive Oxygen Species—A Potential Redox Signal to Initiate Resistance to Oxidative Stress?

    doi: 10.3390/toxins7051411

    Figure Lengend Snippet: Analysis of transcript levels in 7 h germlings. A. parasiticus SU-1, AFS10, and Δ veA were grown on YES solid media and spores were collected at 5 days. Germlings were grown from fresh spores in GMS liquid medium for 7 h, frozen in liquid nitrogen, and stored at −80 °C. RNA was extracted from germlings using grinding and sonication as described in Methods. RT-PCR was performed on total RNA treated with RNAse-free DNAse I with primers specific for each gene ( Table 3 ). PCR products were separated on a 1% agarose gel by electrophoresis. Citrate synthase, a constitutively expressed gene, was used as a positive control. gDNA, genomic DNA.

    Article Snippet: RT-PCR was performed on total RNA treated with RNAse-free DNAse I (Roche Diagnostics, Indianapolis, IN, USA) with primers specific for the coding region of each gene.

    Techniques: Sonication, Reverse Transcription Polymerase Chain Reaction, Polymerase Chain Reaction, Agarose Gel Electrophoresis, Electrophoresis, Positive Control