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Promega t4 dna polymerase
Same efficiency of the extension of DNA and RNA primers on hetero-homopolymeric hybrid and heteropolymeric DNA templates by the p180ΔN-core. For control of the full extension of the primers, we used reactions with <t>T4</t> DNA polymerase, which robustly
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Images

1) Product Images from "The C-terminal Domain of the DNA Polymerase Catalytic Subunit Regulates the Primase and Polymerase Activities of the Human DNA Polymerase α-Primase Complex *"

Article Title: The C-terminal Domain of the DNA Polymerase Catalytic Subunit Regulates the Primase and Polymerase Activities of the Human DNA Polymerase α-Primase Complex *

Journal: The Journal of Biological Chemistry

doi: 10.1074/jbc.M114.570333

Same efficiency of the extension of DNA and RNA primers on hetero-homopolymeric hybrid and heteropolymeric DNA templates by the p180ΔN-core. For control of the full extension of the primers, we used reactions with T4 DNA polymerase, which robustly
Figure Legend Snippet: Same efficiency of the extension of DNA and RNA primers on hetero-homopolymeric hybrid and heteropolymeric DNA templates by the p180ΔN-core. For control of the full extension of the primers, we used reactions with T4 DNA polymerase, which robustly

Techniques Used:

2) Product Images from "The Reovirus S4 Gene 3? Nontranslated Region Contains a Translational Operator Sequence †"

Article Title: The Reovirus S4 Gene 3? Nontranslated Region Contains a Translational Operator Sequence †

Journal: Journal of Virology

doi: 10.1128/JVI.75.14.6517-6526.2001

Diagram of templates used for transcription of full-length and deletion-mutant s4 mRNAs. (A) The relevant region of plasmid pGEM3z/f± used for cloning a full-length cDNA corresponding to the S4 gene of reovirus strain T3D. Indicated are a T7 RNA polymerase promoter (T7), which was cloned adjacent to the 5′-terminal guanosine of the S4 gene cDNA, the ς3 open reading frame (ORF), and a Pst I site, which was engineered adjacent to the 3′ terminus of the S4 gene cDNA. Digestion of plasmid constructs with Pst I followed by incubation with T4 DNA polymerase resulted in templates for transcription that initiate with the 5′ guanosine and terminate with the 3′ cytosine of the S4 gene. Capped s4 mRNAs and s4 3′Δ mRNAs were transcribed in vitro and used in translation studies. Nucleotide lengths are indicated on the right. The s4 3′Δ mRNA terminates with UAAC , corresponding to the stop codon of the transcript and the 3′-terminal cytosine of the full-length S4 gene. (B) Schematic of S4 gene 3′NTR deletion constructs generated from cDNA clones by PCR mutagenesis. The viral sequences deleted in each of the mutant S4 gene cDNAs are indicated on the left; the length of each mRNA transcript is indicated on the right.
Figure Legend Snippet: Diagram of templates used for transcription of full-length and deletion-mutant s4 mRNAs. (A) The relevant region of plasmid pGEM3z/f± used for cloning a full-length cDNA corresponding to the S4 gene of reovirus strain T3D. Indicated are a T7 RNA polymerase promoter (T7), which was cloned adjacent to the 5′-terminal guanosine of the S4 gene cDNA, the ς3 open reading frame (ORF), and a Pst I site, which was engineered adjacent to the 3′ terminus of the S4 gene cDNA. Digestion of plasmid constructs with Pst I followed by incubation with T4 DNA polymerase resulted in templates for transcription that initiate with the 5′ guanosine and terminate with the 3′ cytosine of the S4 gene. Capped s4 mRNAs and s4 3′Δ mRNAs were transcribed in vitro and used in translation studies. Nucleotide lengths are indicated on the right. The s4 3′Δ mRNA terminates with UAAC , corresponding to the stop codon of the transcript and the 3′-terminal cytosine of the full-length S4 gene. (B) Schematic of S4 gene 3′NTR deletion constructs generated from cDNA clones by PCR mutagenesis. The viral sequences deleted in each of the mutant S4 gene cDNAs are indicated on the left; the length of each mRNA transcript is indicated on the right.

Techniques Used: Mutagenesis, Plasmid Preparation, Clone Assay, Construct, Incubation, In Vitro, Generated, Polymerase Chain Reaction

3) Product Images from "Simple and Efficient Method for Heterologous Expression of Clostridial Proteins"

Article Title: Simple and Efficient Method for Heterologous Expression of Clostridial Proteins

Journal: Applied and Environmental Microbiology

doi:

Construction of plasmids encoding tRNAs. The ileX , argU , and leuW sequences encode tRNAs that recognize the ATA, AGA, and CTA codons, respectively. The Ap r , Tet r , and Cm r sequences encode genes for antibiotic resistance. The T7 and Sp6 sequences encode promoters from bacteriophages T7 and sp6, respectively. The shaded areas represent sequences of subB-E tRNA operons other than leuW . DNA pol Taq, Taq DNA polymerase; DNA polT4, T4 DNA polymerase.
Figure Legend Snippet: Construction of plasmids encoding tRNAs. The ileX , argU , and leuW sequences encode tRNAs that recognize the ATA, AGA, and CTA codons, respectively. The Ap r , Tet r , and Cm r sequences encode genes for antibiotic resistance. The T7 and Sp6 sequences encode promoters from bacteriophages T7 and sp6, respectively. The shaded areas represent sequences of subB-E tRNA operons other than leuW . DNA pol Taq, Taq DNA polymerase; DNA polT4, T4 DNA polymerase.

Techniques Used:

Related Articles

Clone Assay:

Article Title: Identification and Characterization of a Promoter Cassette Conferring Adipocyte-Specific Gene Expression
Article Snippet: A similar strategy was chosen for the 5-kb adiponectin promoter cloning with a 5′-end primer located at approximately 5 kb from the 3′ end of the first exon. .. To generate the adiponectin promoter-driven Cre recombinase transgenic construct, the Cre expression cassette from pBS185 was opened by Acc I digestion, which was then blunted by T4 DNA polymerase (Promega, Madison, WI).

Article Title: Purification and Biochemical Characterisation of Rabbit Calicivirus RNA-Dependent RNA Polymerases and Identification of Non-Nucleoside Inhibitors
Article Snippet: Resulting amplicons were cloned into the pET-26b(+) expression vector (Novagen, Darmstadt, Germany) to generate recombinant polymerases with a C-terminal His6 -tag using the following strategy. .. The pET-26b(+) expression vector was linearised using the Nde I restriction enzyme (New England BioLabs), 5′ overhangs were filled in using T4 DNA polymerase (Promega, Madison, WI, USA) to generate a blunt end and was further digested using the Xho I restriction enzyme.

Article Title: Macrophage and Galleria mellonella infection models reflect the virulence of naturally occurring isolates of B. pseudomallei, B. thailandensis and B. oklahomensis
Article Snippet: The PCR product was cloned into pBHR1-RFP via its Sac I/Spe I sites, resulting in plasmid pBHR1-groS-RFP (KmR , CmR ). .. The 4.1-kb fragment was treated with T4 DNA polymerase (Promega) according to the manufacturer's recommendations and re-ligated overnight at 15°C resulting in plasmid pBHR4 (CmR ).

Article Title: Human mutations affect the epigenetic/bookmarking function of HNF1B
Article Snippet: Blunt ends were created with the Klenow polymerase. pcDNA4/TO was digested by HindIII and ApaI, and blunt ends were created by using the T4 DNA polymerase (Promega). .. After ligation and transformation in DH5alpha cells (Life technologies), clones were tested by PCR.

Article Title: Mammalian splicing factor SF1 interacts with SURP domains of U2 snRNP-associated proteins
Article Snippet: Paragraph title: Cloning procedures ... The 3′ overhangs were blunt-ended with T4 DNA polymerase (Promega) followed by religation.

Article Title: Cloning, Expression, Sequence Analysis and Homology Modeling of the Prolyl Endoprotease from Eurygaster integriceps Puton
Article Snippet: Paragraph title: 2.4. Cloning of the Sunn Pest PEP Construct into the Expression Vector ... Similarly, the amplified LIC-PEP DNA was also treated with T4 DNA polymerase (Promega Corp., Madison, WI, USA) in the presence of only dCTPs at the same concentration as the dGTP above.

Article Title: Structural and biochemical insights into the V/I505T mutation found in the EIAV gp45 vaccine strain
Article Snippet: Both the NHR and CHR were amplified by polymerase chain reaction (PCR) using the gene encoding full-length template of EIAV gp140 (gp90 + gp45) and further cloned by employing the ligation-independent cloning (LIC) technique. .. Subsequently, the cleaved plasmids and purified PCR products were digested with a T4 DNA polymerase (Promega) in the presence of dGTP or dCTP, respectively.

Amplification:

Article Title: Size-selective separation and overall-amplification of cell-free fetal DNA fragments using PCR-based enrichment
Article Snippet: .. Overall amplification of maternal plasma total cell-free DNA 100 ng of cfDNA was blunt ended by T4 DNA Polymerase (Promega). .. Nuclease-free water was used as a negative control to replace cell-free DNA.

Article Title: Identification and Characterization of a Promoter Cassette Conferring Adipocyte-Specific Gene Expression
Article Snippet: To generate the 5.4-kb adiponectin promoter, 200 bp from the 5′ end of the first intron were linked to 200 bp from the 3′ end of the first intron by bridging PCR amplification. .. To generate the adiponectin promoter-driven Cre recombinase transgenic construct, the Cre expression cassette from pBS185 was opened by Acc I digestion, which was then blunted by T4 DNA polymerase (Promega, Madison, WI).

Article Title: Purification and Biochemical Characterisation of Rabbit Calicivirus RNA-Dependent RNA Polymerases and Identification of Non-Nucleoside Inhibitors
Article Snippet: The cDNAs encoding viral RdRps were amplified using Q5 High-Fidelity DNA polymerase (New England BioLabs, Ipswich, MA, USA) and gene-specific primers (GeneWorks; for sequence information see ), according to the instructions of the manufacturer. .. The pET-26b(+) expression vector was linearised using the Nde I restriction enzyme (New England BioLabs), 5′ overhangs were filled in using T4 DNA polymerase (Promega, Madison, WI, USA) to generate a blunt end and was further digested using the Xho I restriction enzyme.

Article Title: Macrophage and Galleria mellonella infection models reflect the virulence of naturally occurring isolates of B. pseudomallei, B. thailandensis and B. oklahomensis
Article Snippet: Finally, a 443-bp fragment spanning the upstream region of the groES gene on chromosome I of B. pseudomallei strain K96243 (BPSL2698) was PCR amplified using primers groESprom-fw (5'-CTT GAGCTC GAACGTCGATTCGGACGCAT-3') and groESprom-rv (5'-GCGG ACTAGT ATTCACTCCTCTCTTTGATT-3'), which included Sac I and Spe I restriction sites, respectively. .. The 4.1-kb fragment was treated with T4 DNA polymerase (Promega) according to the manufacturer's recommendations and re-ligated overnight at 15°C resulting in plasmid pBHR4 (CmR ).

Article Title: Human mutations affect the epigenetic/bookmarking function of HNF1B
Article Snippet: The obtained amplicon was ligated with the pEGFP-N1 plasmid (Roche) after digestion by Asp718 and NheI. .. Blunt ends were created with the Klenow polymerase. pcDNA4/TO was digested by HindIII and ApaI, and blunt ends were created by using the T4 DNA polymerase (Promega).

Article Title: Chromatin interaction maps reveal genetic regulation for quantitative traits in maize
Article Snippet: ChIP DNA on beads was used for end-repair and A-tailing using T4 DNA polymerase (Promega, cat. no. M421F) and Klenow enzyme (NEB, cat. no. M0212L), the ChIP DNA ends were proximity-ligated by the single biotinylated bridge-linker, Forward strand: 5′-[5Phos]CGCGATATC/iBIOdT/TATCTGACT-3′, Reverse strand: 5′-[5Phos]GTCAGATAAGATATCGCGT-3′, with the 3′ nucleotide T over-hanging on both strands. .. DNA fragments contained the bridge-linker at ligation junctions were captured by Streptavidin beads, and used as templates for PCR amplification.

Article Title: Cloning, Expression, Sequence Analysis and Homology Modeling of the Prolyl Endoprotease from Eurygaster integriceps Puton
Article Snippet: .. Similarly, the amplified LIC-PEP DNA was also treated with T4 DNA polymerase (Promega Corp., Madison, WI, USA) in the presence of only dCTPs at the same concentration as the dGTP above. .. For each reaction, 2 µL of vector were combined with 4 µL of LIC-PEP.

Article Title: Quantitative Analyses Reveal Novel Roles for N-Glycosylation in a Major Enteric Bacterial Pathogen
Article Snippet: Briefly, pUC18 was digested using PsiI in the pglB gene, and linearized plasmid was then blunt ended using T4 DNA polymerase (Promega) according to the manufacturer’s protocol. .. The antibiotic resistance gene aphA was amplified and cut with StuI and blunt end ligated to the linearized plasmid.

Article Title: Structural and biochemical insights into the V/I505T mutation found in the EIAV gp45 vaccine strain
Article Snippet: Both the NHR and CHR were amplified by polymerase chain reaction (PCR) using the gene encoding full-length template of EIAV gp140 (gp90 + gp45) and further cloned by employing the ligation-independent cloning (LIC) technique. .. Subsequently, the cleaved plasmids and purified PCR products were digested with a T4 DNA polymerase (Promega) in the presence of dGTP or dCTP, respectively.

DNA Ligation:

Article Title: Simple and Efficient Method for Heterologous Expression of Clostridial Proteins
Article Snippet: Restriction enzymes Acc 65I, Bam HI, Bgl II, Eco 52I, Eco ICRI, Eco RI, Hin dIII, Nco I, Nde I, Sac I, Sal GI, Stu I, and Xho I, as well as T4 DNA polymerase, were produced by Promega. .. A rapid DNA ligation kit and an Expand high-fidelity PCR system were supplied by Boehringer Mannheim.

Synthesized:

Article Title: Transcription factor p63 bookmarks and regulates dynamic enhancers during epidermal differentiation
Article Snippet: The second strand was synthesized by adding 20 μl 5× second-strand buffer (Invitrogen), 4 μl 5× first-strand buffer (Invitrogen), 2 μl DTT (0.1 M Invitrogen), 1 μl random hexamers (5 mg/ml Roche), dUTP mix (12.5 mM Invitrogen), 1 μl RNase H (8 U/ml Ambion), 1 μl E. coli DNA polymerase I (10 U/μl Invitrogen), and 1 μl E. coli DNA ligase (10 U/μl NEB) to the purified sample (100 μl total volume). .. After 2 h at 16°C, 1 μl of T4 DNA polymerase (10 U/μl Promega) was added and the reaction was incubated at 16°C for 10 min.

ChIA Pet Assay:

Article Title: Chromatin loops associated with active genes and heterochromatin shape rice genome architecture for transcriptional regulation
Article Snippet: Paragraph title: Long-read ChIA-PET library construction and sequencing ... ChIP DNA was used for end-repair and A-tailing using T4 DNA polymerase (Promega, cat. no. M421F) and Klenow enzyme (NEB, cat. no. M0212L).

Article Title: Chromatin interaction maps reveal genetic regulation for quantitative traits in maize
Article Snippet: Paragraph title: Plant material and ChIA-PET libraries preparation ... ChIP DNA on beads was used for end-repair and A-tailing using T4 DNA polymerase (Promega, cat. no. M421F) and Klenow enzyme (NEB, cat. no. M0212L), the ChIP DNA ends were proximity-ligated by the single biotinylated bridge-linker, Forward strand: 5′-[5Phos]CGCGATATC/iBIOdT/TATCTGACT-3′, Reverse strand: 5′-[5Phos]GTCAGATAAGATATCGCGT-3′, with the 3′ nucleotide T over-hanging on both strands.

Construct:

Article Title: Identification and Characterization of a Promoter Cassette Conferring Adipocyte-Specific Gene Expression
Article Snippet: .. To generate the adiponectin promoter-driven Cre recombinase transgenic construct, the Cre expression cassette from pBS185 was opened by Acc I digestion, which was then blunted by T4 DNA polymerase (Promega, Madison, WI). .. The Cre open reading frame fragment was then released by Xho I digestion, purified, and ligated to the 5.4-kb lac Z vector, which was linearized by Cla I digestion, blunted by T4 DNA polymerase, and opened by Xho I digestion.

Article Title: Cloning, Expression, Sequence Analysis and Homology Modeling of the Prolyl Endoprotease from Eurygaster integriceps Puton
Article Snippet: Paragraph title: 2.4. Cloning of the Sunn Pest PEP Construct into the Expression Vector ... Similarly, the amplified LIC-PEP DNA was also treated with T4 DNA polymerase (Promega Corp., Madison, WI, USA) in the presence of only dCTPs at the same concentration as the dGTP above.

Article Title: Quantitative Analyses Reveal Novel Roles for N-Glycosylation in a Major Enteric Bacterial Pathogen
Article Snippet: Briefly, pUC18 was digested using PsiI in the pglB gene, and linearized plasmid was then blunt ended using T4 DNA polymerase (Promega) according to the manufacturer’s protocol. .. The construct was then electroporated in C. jejuni 11168H, and colonies were selected on CBA supplemented with kanamycin (30 μg/ml).

Article Title: Structural and biochemical insights into the V/I505T mutation found in the EIAV gp45 vaccine strain
Article Snippet: Plasmids and molecular cloning For crystallography structure analysis, the EIAV gp45 was constructed by overlapping the NHR and CHR regions with a GGSGG linker. .. Subsequently, the cleaved plasmids and purified PCR products were digested with a T4 DNA polymerase (Promega) in the presence of dGTP or dCTP, respectively.

Primer Extension Assay:

Article Title: The C-terminal Domain of the DNA Polymerase Catalytic Subunit Regulates the Primase and Polymerase Activities of the Human DNA Polymerase α-Primase Complex *
Article Snippet: Paragraph title: Primer Extension Assay ... T4 DNA polymerase (Promega Corporation; stock concentration 25 n m ) was used as a control for the primer extensions on hetero-DNA and RNA.

Incubation:

Article Title: Transcription factor p63 bookmarks and regulates dynamic enhancers during epidermal differentiation
Article Snippet: .. After 2 h at 16°C, 1 μl of T4 DNA polymerase (10 U/μl Promega) was added and the reaction was incubated at 16°C for 10 min. .. The ds-cDNA was purified using the MinElute Reaction Cleanup Kit (Qiagen), according to the manufacturer's protocol.

Article Title: Chromatin interaction maps reveal genetic regulation for quantitative traits in maize
Article Snippet: Then, mixed fragmented chromatin extract with antibody-loaded beads and incubated it at 4 °C for overnight with rotation. .. ChIP DNA on beads was used for end-repair and A-tailing using T4 DNA polymerase (Promega, cat. no. M421F) and Klenow enzyme (NEB, cat. no. M0212L), the ChIP DNA ends were proximity-ligated by the single biotinylated bridge-linker, Forward strand: 5′-[5Phos]CGCGATATC/iBIOdT/TATCTGACT-3′, Reverse strand: 5′-[5Phos]GTCAGATAAGATATCGCGT-3′, with the 3′ nucleotide T over-hanging on both strands.

Article Title: The Reovirus S4 Gene 3? Nontranslated Region Contains a Translational Operator Sequence †
Article Snippet: .. Plasmids were linearized by digestion with 1 U of Pst I per μl, and 3′ overhang sequences were removed by incubation with 1 to 2 U of T4 DNA polymerase (Promega) per μl at 11°C for 20 min in the presence of 1 mM Tris-HCl (pH 7.9), 1 mM MgCl2 , 5 mM NaCl, 0.1 mM dithiothreitol (DTT), 10 μM deoxynucleoside triphosphate, and 50 μg of bovine serum albumin per in a final volume of 20 μl. .. Linearized plasmids were gel purified and resuspended in nuclease-free water.

Article Title: Cloning, Expression, Sequence Analysis and Homology Modeling of the Prolyl Endoprotease from Eurygaster integriceps Puton
Article Snippet: Similarly, the amplified LIC-PEP DNA was also treated with T4 DNA polymerase (Promega Corp., Madison, WI, USA) in the presence of only dCTPs at the same concentration as the dGTP above. .. Following incubation, 2 µL of 25 mM EDTA was added to each reaction tube and incubated at room temperature for 5 min. Two microliters of each LIC reaction were transformed into the BL21(DE3)-competent E. coli cells (NEB, Ipswich, UK).

Expressing:

Article Title: Identification and Characterization of a Promoter Cassette Conferring Adipocyte-Specific Gene Expression
Article Snippet: .. To generate the adiponectin promoter-driven Cre recombinase transgenic construct, the Cre expression cassette from pBS185 was opened by Acc I digestion, which was then blunted by T4 DNA polymerase (Promega, Madison, WI). .. The Cre open reading frame fragment was then released by Xho I digestion, purified, and ligated to the 5.4-kb lac Z vector, which was linearized by Cla I digestion, blunted by T4 DNA polymerase, and opened by Xho I digestion.

Article Title: Purification and Biochemical Characterisation of Rabbit Calicivirus RNA-Dependent RNA Polymerases and Identification of Non-Nucleoside Inhibitors
Article Snippet: .. The pET-26b(+) expression vector was linearised using the Nde I restriction enzyme (New England BioLabs), 5′ overhangs were filled in using T4 DNA polymerase (Promega, Madison, WI, USA) to generate a blunt end and was further digested using the Xho I restriction enzyme. .. All inserts were ligated into linearised vectors using the T4 DNA ligase (New England BioLabs).

Article Title: Mammalian splicing factor SF1 interacts with SURP domains of U2 snRNP-associated proteins
Article Snippet: For transient expression of N-terminal GFP fusion proteins in HeLa cells, SF1 sequences were cloned into the Gateway vector pDEST53 (Life Technologies). .. The 3′ overhangs were blunt-ended with T4 DNA polymerase (Promega) followed by religation.

Article Title: Cloning, Expression, Sequence Analysis and Homology Modeling of the Prolyl Endoprotease from Eurygaster integriceps Puton
Article Snippet: Paragraph title: 2.4. Cloning of the Sunn Pest PEP Construct into the Expression Vector ... Similarly, the amplified LIC-PEP DNA was also treated with T4 DNA polymerase (Promega Corp., Madison, WI, USA) in the presence of only dCTPs at the same concentration as the dGTP above.

Transformation Assay:

Article Title: Human mutations affect the epigenetic/bookmarking function of HNF1B
Article Snippet: Blunt ends were created with the Klenow polymerase. pcDNA4/TO was digested by HindIII and ApaI, and blunt ends were created by using the T4 DNA polymerase (Promega). .. After ligation and transformation in DH5alpha cells (Life technologies), clones were tested by PCR.

Article Title: Cloning, Expression, Sequence Analysis and Homology Modeling of the Prolyl Endoprotease from Eurygaster integriceps Puton
Article Snippet: Similarly, the amplified LIC-PEP DNA was also treated with T4 DNA polymerase (Promega Corp., Madison, WI, USA) in the presence of only dCTPs at the same concentration as the dGTP above. .. Following incubation, 2 µL of 25 mM EDTA was added to each reaction tube and incubated at room temperature for 5 min. Two microliters of each LIC reaction were transformed into the BL21(DE3)-competent E. coli cells (NEB, Ipswich, UK).

Article Title: Structural and biochemical insights into the V/I505T mutation found in the EIAV gp45 vaccine strain
Article Snippet: Subsequently, the cleaved plasmids and purified PCR products were digested with a T4 DNA polymerase (Promega) in the presence of dGTP or dCTP, respectively. .. The annealed mixture containing pET30-TEV/LIC-gp45 was transformed into E.coli DH5α competent cells for plasmid propagation.

Derivative Assay:

Article Title: Structural and biochemical insights into the V/I505T mutation found in the EIAV gp45 vaccine strain
Article Snippet: Subsequently, the cleaved plasmids and purified PCR products were digested with a T4 DNA polymerase (Promega) in the presence of dGTP or dCTP, respectively. .. Using this method, a 6× His-tag and a TEV cleavage site derived from the LIC vector were fused upstream of gp45.

Ligation:

Article Title: Chromatin loops associated with active genes and heterochromatin shape rice genome architecture for transcriptional regulation
Article Snippet: ChIP DNA was used for end-repair and A-tailing using T4 DNA polymerase (Promega, cat. no. M421F) and Klenow enzyme (NEB, cat. no. M0212L). .. Proximity ligation of ChIP DNA was performed using biotinylated bridge-linker (forward strand: 5′-[5Phos]CGCGATATC/iBIOdT/TATCTGACT-3′, reverse strand: 5′-[5Phos]GTCAGATAAGATATCGCGT-3′).

Article Title: Human mutations affect the epigenetic/bookmarking function of HNF1B
Article Snippet: Blunt ends were created with the Klenow polymerase. pcDNA4/TO was digested by HindIII and ApaI, and blunt ends were created by using the T4 DNA polymerase (Promega). .. After ligation and transformation in DH5alpha cells (Life technologies), clones were tested by PCR.

Article Title: Chromatin interaction maps reveal genetic regulation for quantitative traits in maize
Article Snippet: ChIP DNA on beads was used for end-repair and A-tailing using T4 DNA polymerase (Promega, cat. no. M421F) and Klenow enzyme (NEB, cat. no. M0212L), the ChIP DNA ends were proximity-ligated by the single biotinylated bridge-linker, Forward strand: 5′-[5Phos]CGCGATATC/iBIOdT/TATCTGACT-3′, Reverse strand: 5′-[5Phos]GTCAGATAAGATATCGCGT-3′, with the 3′ nucleotide T over-hanging on both strands. .. Proximity ligation DNA was reverse cross-linked and fragmented before adding sequencing adaptors simultaneously by using Tn5 transposase (VAHTS; cat. no. TD501).

Article Title: Cloning, Expression, Sequence Analysis and Homology Modeling of the Prolyl Endoprotease from Eurygaster integriceps Puton
Article Snippet: In addition, it has a Kanamycin resistance gene and allows for ligation-independent cloning (LIC) and IPTG induction. .. Similarly, the amplified LIC-PEP DNA was also treated with T4 DNA polymerase (Promega Corp., Madison, WI, USA) in the presence of only dCTPs at the same concentration as the dGTP above.

Article Title: Structural and biochemical insights into the V/I505T mutation found in the EIAV gp45 vaccine strain
Article Snippet: Both the NHR and CHR were amplified by polymerase chain reaction (PCR) using the gene encoding full-length template of EIAV gp140 (gp90 + gp45) and further cloned by employing the ligation-independent cloning (LIC) technique. .. Subsequently, the cleaved plasmids and purified PCR products were digested with a T4 DNA polymerase (Promega) in the presence of dGTP or dCTP, respectively.

Infection:

Article Title: Purification and Biochemical Characterisation of Rabbit Calicivirus RNA-Dependent RNA Polymerases and Identification of Non-Nucleoside Inhibitors
Article Snippet: Plasmids RHDV RNA was purified from a commercial RHDV suspension (Czech strain V351, GenBank accession number KF594473.1, Elizabeth Macarthur Agricultural Institute, Menangle, Australia); RCV RNA was purified from the homogenised small intestine of a rabbit infected with the non-pathogenic calicivirus RCV-A1 (GenBank accession number EU871528.1) [ ] using the RNeasy Mini Kit (Qiagen, Hilden, Germany). .. The pET-26b(+) expression vector was linearised using the Nde I restriction enzyme (New England BioLabs), 5′ overhangs were filled in using T4 DNA polymerase (Promega, Madison, WI, USA) to generate a blunt end and was further digested using the Xho I restriction enzyme.

Generated:

Article Title: Mammalian splicing factor SF1 interacts with SURP domains of U2 snRNP-associated proteins
Article Snippet: Templates for in vitro transcription of 3′ splice site pre-mRNAs were generated by HindIII and BstEII digestion of a pBluescript plasmid encoding the region spanning exons 1 and 2 of AdML pre-mRNA ( ). .. The 3′ overhangs were blunt-ended with T4 DNA polymerase (Promega) followed by religation.

Article Title: Quantitative Analyses Reveal Novel Roles for N-Glycosylation in a Major Enteric Bacterial Pathogen
Article Snippet: Construction of C. jejuni pglB ::aphA was achieved by nonpolar insertion of the aphA gene into a unique restriction site present in the pglB gene of the pUC18 plasmid generated from a random genomic library used in sequencing C. jejuni 11168 ( ). .. Briefly, pUC18 was digested using PsiI in the pglB gene, and linearized plasmid was then blunt ended using T4 DNA polymerase (Promega) according to the manufacturer’s protocol.

Sequencing:

Article Title: Chromatin loops associated with active genes and heterochromatin shape rice genome architecture for transcriptional regulation
Article Snippet: Paragraph title: Long-read ChIA-PET library construction and sequencing ... ChIP DNA was used for end-repair and A-tailing using T4 DNA polymerase (Promega, cat. no. M421F) and Klenow enzyme (NEB, cat. no. M0212L).

Article Title: Purification and Biochemical Characterisation of Rabbit Calicivirus RNA-Dependent RNA Polymerases and Identification of Non-Nucleoside Inhibitors
Article Snippet: The cDNAs encoding viral RdRps were amplified using Q5 High-Fidelity DNA polymerase (New England BioLabs, Ipswich, MA, USA) and gene-specific primers (GeneWorks; for sequence information see ), according to the instructions of the manufacturer. .. The pET-26b(+) expression vector was linearised using the Nde I restriction enzyme (New England BioLabs), 5′ overhangs were filled in using T4 DNA polymerase (Promega, Madison, WI, USA) to generate a blunt end and was further digested using the Xho I restriction enzyme.

Article Title: Chromatin interaction maps reveal genetic regulation for quantitative traits in maize
Article Snippet: ChIP DNA on beads was used for end-repair and A-tailing using T4 DNA polymerase (Promega, cat. no. M421F) and Klenow enzyme (NEB, cat. no. M0212L), the ChIP DNA ends were proximity-ligated by the single biotinylated bridge-linker, Forward strand: 5′-[5Phos]CGCGATATC/iBIOdT/TATCTGACT-3′, Reverse strand: 5′-[5Phos]GTCAGATAAGATATCGCGT-3′, with the 3′ nucleotide T over-hanging on both strands. .. Proximity ligation DNA was reverse cross-linked and fragmented before adding sequencing adaptors simultaneously by using Tn5 transposase (VAHTS; cat. no. TD501).

Article Title: Quantitative Analyses Reveal Novel Roles for N-Glycosylation in a Major Enteric Bacterial Pathogen
Article Snippet: Construction of C. jejuni pglB ::aphA was achieved by nonpolar insertion of the aphA gene into a unique restriction site present in the pglB gene of the pUC18 plasmid generated from a random genomic library used in sequencing C. jejuni 11168 ( ). .. Briefly, pUC18 was digested using PsiI in the pglB gene, and linearized plasmid was then blunt ended using T4 DNA polymerase (Promega) according to the manufacturer’s protocol.

Sonication:

Article Title: Chromatin interaction maps reveal genetic regulation for quantitative traits in maize
Article Snippet: The chromatin extract was fragmented into 1–3 kb by sonication using a Bioruptor (Diagenode) under HIGH intensity (30 cycles of 30 s ON and 50 s OFF). .. ChIP DNA on beads was used for end-repair and A-tailing using T4 DNA polymerase (Promega, cat. no. M421F) and Klenow enzyme (NEB, cat. no. M0212L), the ChIP DNA ends were proximity-ligated by the single biotinylated bridge-linker, Forward strand: 5′-[5Phos]CGCGATATC/iBIOdT/TATCTGACT-3′, Reverse strand: 5′-[5Phos]GTCAGATAAGATATCGCGT-3′, with the 3′ nucleotide T over-hanging on both strands.

Injection:

Article Title: Identification and Characterization of a Promoter Cassette Conferring Adipocyte-Specific Gene Expression
Article Snippet: To generate the adiponectin promoter-driven Cre recombinase transgenic construct, the Cre expression cassette from pBS185 was opened by Acc I digestion, which was then blunted by T4 DNA polymerase (Promega, Madison, WI). .. The resulting construct was linearized by Nae I digestion and purified for pronuclear injection.

Binding Assay:

Article Title: The C-terminal Domain of the DNA Polymerase Catalytic Subunit Regulates the Primase and Polymerase Activities of the Human DNA Polymerase α-Primase Complex *
Article Snippet: The low ratio of primase to template allowed us to approach single-hit conditions, whereas at the equal ratio the primase was able to synthesize longer products due to multiple binding events with substrate. .. T4 DNA polymerase (Promega Corporation; stock concentration 25 n m ) was used as a control for the primer extensions on hetero-DNA and RNA.

Crocin Bleaching Assay:

Article Title: Quantitative Analyses Reveal Novel Roles for N-Glycosylation in a Major Enteric Bacterial Pathogen
Article Snippet: Briefly, pUC18 was digested using PsiI in the pglB gene, and linearized plasmid was then blunt ended using T4 DNA polymerase (Promega) according to the manufacturer’s protocol. .. The construct was then electroporated in C. jejuni 11168H, and colonies were selected on CBA supplemented with kanamycin (30 μg/ml).

Molecular Cloning:

Article Title: Structural and biochemical insights into the V/I505T mutation found in the EIAV gp45 vaccine strain
Article Snippet: Paragraph title: Plasmids and molecular cloning ... Subsequently, the cleaved plasmids and purified PCR products were digested with a T4 DNA polymerase (Promega) in the presence of dGTP or dCTP, respectively.

RNA Sequencing Assay:

Article Title: Transcription factor p63 bookmarks and regulates dynamic enhancers during epidermal differentiation
Article Snippet: Paragraph title: RNA extraction and RNA-seq ... After 2 h at 16°C, 1 μl of T4 DNA polymerase (10 U/μl Promega) was added and the reaction was incubated at 16°C for 10 min.

Fluorescence:

Article Title: The C-terminal Domain of the DNA Polymerase Catalytic Subunit Regulates the Primase and Polymerase Activities of the Human DNA Polymerase α-Primase Complex *
Article Snippet: T4 DNA polymerase (Promega Corporation; stock concentration 25 n m ) was used as a control for the primer extensions on hetero-DNA and RNA. .. Visualization of the products used the Typhoon 9410 imager (emission of fluorescence at 645 nm).

Magnetic Beads:

Article Title: Chromatin loops associated with active genes and heterochromatin shape rice genome architecture for transcriptional regulation
Article Snippet: To coat antibodies to magnetic beads, take 60–100 μg antibody (RNAPII, BioLegend, 920102; H3K9me2, Abcam, ab1220 and H3K4me3, ABclonal, A2357) and mix with 800 μl suspended protein G (~1:100 dilution) magnetic beads at 4 °C for 8 h with rotation. .. ChIP DNA was used for end-repair and A-tailing using T4 DNA polymerase (Promega, cat. no. M421F) and Klenow enzyme (NEB, cat. no. M0212L).

Article Title: Chromatin interaction maps reveal genetic regulation for quantitative traits in maize
Article Snippet: To prepare antibody-loaded beads, 60–100 μL antibody (RNAPII, BioLegend cat. no. 920102 and H3K4me3, ABclonal cat. no. A2357) was mixed with 800 μL suspended protein G magnetic beads (~1:100 dilution) followed by incubation at 4 °C for 8 h with rotation. .. ChIP DNA on beads was used for end-repair and A-tailing using T4 DNA polymerase (Promega, cat. no. M421F) and Klenow enzyme (NEB, cat. no. M0212L), the ChIP DNA ends were proximity-ligated by the single biotinylated bridge-linker, Forward strand: 5′-[5Phos]CGCGATATC/iBIOdT/TATCTGACT-3′, Reverse strand: 5′-[5Phos]GTCAGATAAGATATCGCGT-3′, with the 3′ nucleotide T over-hanging on both strands.

Mutagenesis:

Article Title: Mammalian splicing factor SF1 interacts with SURP domains of U2 snRNP-associated proteins
Article Snippet: The 3′ overhangs were blunt-ended with T4 DNA polymerase (Promega) followed by religation. .. This plasmid was used to mutate the original AdML BPS (UACUUAU) to a consensus (UACUAAC) or weak (AAUUCAC) BPS with the GeneTailor Site-Directed Mutagenesis System (Life Technologies).

Article Title: Structural and biochemical insights into the V/I505T mutation found in the EIAV gp45 vaccine strain
Article Snippet: Subsequently, the cleaved plasmids and purified PCR products were digested with a T4 DNA polymerase (Promega) in the presence of dGTP or dCTP, respectively. .. The QuikChange site-directed mutagenesis kit (Stratagene) was used to generate point mutations.

Isolation:

Article Title: Macrophage and Galleria mellonella infection models reflect the virulence of naturally occurring isolates of B. pseudomallei, B. thailandensis and B. oklahomensis
Article Snippet: Firstly, unmethylated pBHR1 plasmid DNA isolated from a dcm - /dam - E. coli strain C2925 (New England Biolabs) was cut with Stu I/PpuM I, which resulted in a 1.2-kb fragment encompassing the kanamycin resistance cassette and a 4.1-kb plasmid backbone fragment. .. The 4.1-kb fragment was treated with T4 DNA polymerase (Promega) according to the manufacturer's recommendations and re-ligated overnight at 15°C resulting in plasmid pBHR4 (CmR ).

Negative Control:

Article Title: Size-selective separation and overall-amplification of cell-free fetal DNA fragments using PCR-based enrichment
Article Snippet: Overall amplification of maternal plasma total cell-free DNA 100 ng of cfDNA was blunt ended by T4 DNA Polymerase (Promega). .. Nuclease-free water was used as a negative control to replace cell-free DNA.

Purification:

Article Title: Size-selective separation and overall-amplification of cell-free fetal DNA fragments using PCR-based enrichment
Article Snippet: Overall amplification of maternal plasma total cell-free DNA 100 ng of cfDNA was blunt ended by T4 DNA Polymerase (Promega). .. Products were purified by AxyPrep PCRCleanup Kit (AXYGEN). dA Tailing Kit (Tiandz, China) was used for adding adenine bases to the 3′-ends.

Article Title: Identification and Characterization of a Promoter Cassette Conferring Adipocyte-Specific Gene Expression
Article Snippet: To generate the adiponectin promoter-driven Cre recombinase transgenic construct, the Cre expression cassette from pBS185 was opened by Acc I digestion, which was then blunted by T4 DNA polymerase (Promega, Madison, WI). .. The Cre open reading frame fragment was then released by Xho I digestion, purified, and ligated to the 5.4-kb lac Z vector, which was linearized by Cla I digestion, blunted by T4 DNA polymerase, and opened by Xho I digestion.

Article Title: Purification and Biochemical Characterisation of Rabbit Calicivirus RNA-Dependent RNA Polymerases and Identification of Non-Nucleoside Inhibitors
Article Snippet: Plasmids RHDV RNA was purified from a commercial RHDV suspension (Czech strain V351, GenBank accession number KF594473.1, Elizabeth Macarthur Agricultural Institute, Menangle, Australia); RCV RNA was purified from the homogenised small intestine of a rabbit infected with the non-pathogenic calicivirus RCV-A1 (GenBank accession number EU871528.1) [ ] using the RNeasy Mini Kit (Qiagen, Hilden, Germany). .. The pET-26b(+) expression vector was linearised using the Nde I restriction enzyme (New England BioLabs), 5′ overhangs were filled in using T4 DNA polymerase (Promega, Madison, WI, USA) to generate a blunt end and was further digested using the Xho I restriction enzyme.

Article Title: Transcription factor p63 bookmarks and regulates dynamic enhancers during epidermal differentiation
Article Snippet: The second strand was synthesized by adding 20 μl 5× second-strand buffer (Invitrogen), 4 μl 5× first-strand buffer (Invitrogen), 2 μl DTT (0.1 M Invitrogen), 1 μl random hexamers (5 mg/ml Roche), dUTP mix (12.5 mM Invitrogen), 1 μl RNase H (8 U/ml Ambion), 1 μl E. coli DNA polymerase I (10 U/μl Invitrogen), and 1 μl E. coli DNA ligase (10 U/μl NEB) to the purified sample (100 μl total volume). .. After 2 h at 16°C, 1 μl of T4 DNA polymerase (10 U/μl Promega) was added and the reaction was incubated at 16°C for 10 min.

Article Title: The Reovirus S4 Gene 3? Nontranslated Region Contains a Translational Operator Sequence †
Article Snippet: Plasmids were linearized by digestion with 1 U of Pst I per μl, and 3′ overhang sequences were removed by incubation with 1 to 2 U of T4 DNA polymerase (Promega) per μl at 11°C for 20 min in the presence of 1 mM Tris-HCl (pH 7.9), 1 mM MgCl2 , 5 mM NaCl, 0.1 mM dithiothreitol (DTT), 10 μM deoxynucleoside triphosphate, and 50 μg of bovine serum albumin per in a final volume of 20 μl. .. Linearized plasmids were gel purified and resuspended in nuclease-free water.

Article Title: Structural and biochemical insights into the V/I505T mutation found in the EIAV gp45 vaccine strain
Article Snippet: .. Subsequently, the cleaved plasmids and purified PCR products were digested with a T4 DNA polymerase (Promega) in the presence of dGTP or dCTP, respectively. .. The annealed mixture containing pET30-TEV/LIC-gp45 was transformed into E.coli DH5α competent cells for plasmid propagation.

Polymerase Chain Reaction:

Article Title: Identification and Characterization of a Promoter Cassette Conferring Adipocyte-Specific Gene Expression
Article Snippet: To generate the 5.4-kb adiponectin promoter, 200 bp from the 5′ end of the first intron were linked to 200 bp from the 3′ end of the first intron by bridging PCR amplification. .. To generate the adiponectin promoter-driven Cre recombinase transgenic construct, the Cre expression cassette from pBS185 was opened by Acc I digestion, which was then blunted by T4 DNA polymerase (Promega, Madison, WI).

Article Title: Purification and Biochemical Characterisation of Rabbit Calicivirus RNA-Dependent RNA Polymerases and Identification of Non-Nucleoside Inhibitors
Article Snippet: PCR products were digested using the Xho I restriction enzyme (New England BioLabs). .. The pET-26b(+) expression vector was linearised using the Nde I restriction enzyme (New England BioLabs), 5′ overhangs were filled in using T4 DNA polymerase (Promega, Madison, WI, USA) to generate a blunt end and was further digested using the Xho I restriction enzyme.

Article Title: Macrophage and Galleria mellonella infection models reflect the virulence of naturally occurring isolates of B. pseudomallei, B. thailandensis and B. oklahomensis
Article Snippet: The PCR product was cloned into pBHR1-RFP via its Sac I/Spe I sites, resulting in plasmid pBHR1-groS-RFP (KmR , CmR ). .. The 4.1-kb fragment was treated with T4 DNA polymerase (Promega) according to the manufacturer's recommendations and re-ligated overnight at 15°C resulting in plasmid pBHR4 (CmR ).

Article Title: Human mutations affect the epigenetic/bookmarking function of HNF1B
Article Snippet: For the construction of ΔH3 (house name 10p7), a PCR was carried out on the 6p15 plasmid, containing the human cDNA of HNF1B isoform A ( ) with the primers 10i1 and 10i4 (amplification from codon 1 to 291). .. Blunt ends were created with the Klenow polymerase. pcDNA4/TO was digested by HindIII and ApaI, and blunt ends were created by using the T4 DNA polymerase (Promega).

Article Title: Chromatin interaction maps reveal genetic regulation for quantitative traits in maize
Article Snippet: ChIP DNA on beads was used for end-repair and A-tailing using T4 DNA polymerase (Promega, cat. no. M421F) and Klenow enzyme (NEB, cat. no. M0212L), the ChIP DNA ends were proximity-ligated by the single biotinylated bridge-linker, Forward strand: 5′-[5Phos]CGCGATATC/iBIOdT/TATCTGACT-3′, Reverse strand: 5′-[5Phos]GTCAGATAAGATATCGCGT-3′, with the 3′ nucleotide T over-hanging on both strands. .. DNA fragments contained the bridge-linker at ligation junctions were captured by Streptavidin beads, and used as templates for PCR amplification.

Article Title: Cloning, Expression, Sequence Analysis and Homology Modeling of the Prolyl Endoprotease from Eurygaster integriceps Puton
Article Snippet: The complete PEP gene was amplified by PCR with the LIC-modified PEP Start (Lic PEP start) and PEP Stop (Lic PEP stop) primers ( ) using the PEP clone from as the template; the amplicon was designated as LIC-PEP. .. Similarly, the amplified LIC-PEP DNA was also treated with T4 DNA polymerase (Promega Corp., Madison, WI, USA) in the presence of only dCTPs at the same concentration as the dGTP above.

Article Title: Structural and biochemical insights into the V/I505T mutation found in the EIAV gp45 vaccine strain
Article Snippet: .. Subsequently, the cleaved plasmids and purified PCR products were digested with a T4 DNA polymerase (Promega) in the presence of dGTP or dCTP, respectively. .. The annealed mixture containing pET30-TEV/LIC-gp45 was transformed into E.coli DH5α competent cells for plasmid propagation.

Article Title: Simple and Efficient Method for Heterologous Expression of Clostridial Proteins
Article Snippet: Restriction enzymes Acc 65I, Bam HI, Bgl II, Eco 52I, Eco ICRI, Eco RI, Hin dIII, Nco I, Nde I, Sac I, Sal GI, Stu I, and Xho I, as well as T4 DNA polymerase, were produced by Promega. .. A rapid DNA ligation kit and an Expand high-fidelity PCR system were supplied by Boehringer Mannheim.

Positron Emission Tomography:

Article Title: Purification and Biochemical Characterisation of Rabbit Calicivirus RNA-Dependent RNA Polymerases and Identification of Non-Nucleoside Inhibitors
Article Snippet: .. The pET-26b(+) expression vector was linearised using the Nde I restriction enzyme (New England BioLabs), 5′ overhangs were filled in using T4 DNA polymerase (Promega, Madison, WI, USA) to generate a blunt end and was further digested using the Xho I restriction enzyme. .. All inserts were ligated into linearised vectors using the T4 DNA ligase (New England BioLabs).

Lysis:

Article Title: Chromatin loops associated with active genes and heterochromatin shape rice genome architecture for transcriptional regulation
Article Snippet: The supernatant was discarded and the beads were sequentially washed with 5 mL 0.1% SDS FA cell lysis buffer (0.05 M HEPES-KOH, 0.15 M NaCl, 0.001 M EDTA, 1% Triton X-100, 0.1% sodium deoxycholate, and 0.1% SDS) for three times, 5 mL High salt ChIP buffer (0.05 M HEPES-KOH, 0.35 M NaCl, 0.001 M EDTA, 1% Triton X-100, 0.1% sodium deoxycholate, and 0.1% SDS) twice, 5 mL ChIP wash buffer (0.01 M Tris–HCl, 0.25 M LiCl, 0.001 M EDTA, 2.5% NP-40, and 0.5% sodium deoxycholate) once and 5 mL 1 × TE buffer (pH 8.0, Ambion, cat. no. AM9849) twice. .. ChIP DNA was used for end-repair and A-tailing using T4 DNA polymerase (Promega, cat. no. M421F) and Klenow enzyme (NEB, cat. no. M0212L).

Article Title: Chromatin interaction maps reveal genetic regulation for quantitative traits in maize
Article Snippet: The supernatant was discarded and the beads were washed using 5 mL 0.1% SDS FA cell lysis buffer (0.05 M, HEPES-KOH, 0.15 M NaCl, 0.001 M EDTA, 1% Triton X-100, 0.1% sodium deoxycholate, and 0.1% SDS) for three times, then followed washing with 5 mL High salt ChIP buffer (0.05 M, HEPES-KOH, 0.35 M NaCl, 0.001 M EDTA, 1% Triton X-100, 0.1% sodium deoxycholate, and 0.1% SDS) twice; the beads were washed with 5 mL ChIP wash buffer (0.01 M Tris–HCl, 0.25 M LiCl, 0.001 M EDTA, 2.5% NP-40, and 0.5% sodium deoxycholate) once and finally followed washing with 5 mL 1× TE buffer (pH 8.0, Ambion, cat. no. AM9849) twice. .. ChIP DNA on beads was used for end-repair and A-tailing using T4 DNA polymerase (Promega, cat. no. M421F) and Klenow enzyme (NEB, cat. no. M0212L), the ChIP DNA ends were proximity-ligated by the single biotinylated bridge-linker, Forward strand: 5′-[5Phos]CGCGATATC/iBIOdT/TATCTGACT-3′, Reverse strand: 5′-[5Phos]GTCAGATAAGATATCGCGT-3′, with the 3′ nucleotide T over-hanging on both strands.

Concentration Assay:

Article Title: The C-terminal Domain of the DNA Polymerase Catalytic Subunit Regulates the Primase and Polymerase Activities of the Human DNA Polymerase α-Primase Complex *
Article Snippet: .. T4 DNA polymerase (Promega Corporation; stock concentration 25 n m ) was used as a control for the primer extensions on hetero-DNA and RNA. ..

Article Title: Cloning, Expression, Sequence Analysis and Homology Modeling of the Prolyl Endoprotease from Eurygaster integriceps Puton
Article Snippet: .. Similarly, the amplified LIC-PEP DNA was also treated with T4 DNA polymerase (Promega Corp., Madison, WI, USA) in the presence of only dCTPs at the same concentration as the dGTP above. .. For each reaction, 2 µL of vector were combined with 4 µL of LIC-PEP.

Chromatin Immunoprecipitation:

Article Title: Chromatin loops associated with active genes and heterochromatin shape rice genome architecture for transcriptional regulation
Article Snippet: .. ChIP DNA was used for end-repair and A-tailing using T4 DNA polymerase (Promega, cat. no. M421F) and Klenow enzyme (NEB, cat. no. M0212L). .. Proximity ligation of ChIP DNA was performed using biotinylated bridge-linker (forward strand: 5′-[5Phos]CGCGATATC/iBIOdT/TATCTGACT-3′, reverse strand: 5′-[5Phos]GTCAGATAAGATATCGCGT-3′).

Article Title: Chromatin interaction maps reveal genetic regulation for quantitative traits in maize
Article Snippet: .. ChIP DNA on beads was used for end-repair and A-tailing using T4 DNA polymerase (Promega, cat. no. M421F) and Klenow enzyme (NEB, cat. no. M0212L), the ChIP DNA ends were proximity-ligated by the single biotinylated bridge-linker, Forward strand: 5′-[5Phos]CGCGATATC/iBIOdT/TATCTGACT-3′, Reverse strand: 5′-[5Phos]GTCAGATAAGATATCGCGT-3′, with the 3′ nucleotide T over-hanging on both strands. .. Proximity ligation DNA was reverse cross-linked and fragmented before adding sequencing adaptors simultaneously by using Tn5 transposase (VAHTS; cat. no. TD501).

Plasmid Preparation:

Article Title: Identification and Characterization of a Promoter Cassette Conferring Adipocyte-Specific Gene Expression
Article Snippet: Paragraph title: Plasmid construction and transgenic mouse generation ... To generate the adiponectin promoter-driven Cre recombinase transgenic construct, the Cre expression cassette from pBS185 was opened by Acc I digestion, which was then blunted by T4 DNA polymerase (Promega, Madison, WI).

Article Title: Purification and Biochemical Characterisation of Rabbit Calicivirus RNA-Dependent RNA Polymerases and Identification of Non-Nucleoside Inhibitors
Article Snippet: .. The pET-26b(+) expression vector was linearised using the Nde I restriction enzyme (New England BioLabs), 5′ overhangs were filled in using T4 DNA polymerase (Promega, Madison, WI, USA) to generate a blunt end and was further digested using the Xho I restriction enzyme. .. All inserts were ligated into linearised vectors using the T4 DNA ligase (New England BioLabs).

Article Title: Macrophage and Galleria mellonella infection models reflect the virulence of naturally occurring isolates of B. pseudomallei, B. thailandensis and B. oklahomensis
Article Snippet: .. The 4.1-kb fragment was treated with T4 DNA polymerase (Promega) according to the manufacturer's recommendations and re-ligated overnight at 15°C resulting in plasmid pBHR4 (CmR ). .. Finally, a 1-kb fragment representing the cat gene of plasmid pBHR4 was replaced by a 3.2-kb fragment of plasmid pBHR1-groS-RFP, which encompassed the RFP gene linked to the groES promoter, the rrnB terminator and the cat gene, via BstB I restriction as described for the construction of pBHR-MCS1 & 2.

Article Title: Human mutations affect the epigenetic/bookmarking function of HNF1B
Article Snippet: The obtained amplicon was ligated with the pEGFP-N1 plasmid (Roche) after digestion by Asp718 and NheI. .. Blunt ends were created with the Klenow polymerase. pcDNA4/TO was digested by HindIII and ApaI, and blunt ends were created by using the T4 DNA polymerase (Promega).

Article Title: Mammalian splicing factor SF1 interacts with SURP domains of U2 snRNP-associated proteins
Article Snippet: Templates for in vitro transcription of 3′ splice site pre-mRNAs were generated by HindIII and BstEII digestion of a pBluescript plasmid encoding the region spanning exons 1 and 2 of AdML pre-mRNA ( ). .. The 3′ overhangs were blunt-ended with T4 DNA polymerase (Promega) followed by religation.

Article Title: The Reovirus S4 Gene 3? Nontranslated Region Contains a Translational Operator Sequence †
Article Snippet: Plasmids were linearized by digestion with 1 U of Pst I per μl, and 3′ overhang sequences were removed by incubation with 1 to 2 U of T4 DNA polymerase (Promega) per μl at 11°C for 20 min in the presence of 1 mM Tris-HCl (pH 7.9), 1 mM MgCl2 , 5 mM NaCl, 0.1 mM dithiothreitol (DTT), 10 μM deoxynucleoside triphosphate, and 50 μg of bovine serum albumin per in a final volume of 20 μl. .. In vitro transcription of capped reovirus mRNAs was performed using linearized plasmid as template and the mMessage mMachine T7 RNA polymerase transcription system (Ambion, Austin, Tex.).

Article Title: Cloning, Expression, Sequence Analysis and Homology Modeling of the Prolyl Endoprotease from Eurygaster integriceps Puton
Article Snippet: Paragraph title: 2.4. Cloning of the Sunn Pest PEP Construct into the Expression Vector ... Similarly, the amplified LIC-PEP DNA was also treated with T4 DNA polymerase (Promega Corp., Madison, WI, USA) in the presence of only dCTPs at the same concentration as the dGTP above.

Article Title: Quantitative Analyses Reveal Novel Roles for N-Glycosylation in a Major Enteric Bacterial Pathogen
Article Snippet: .. Briefly, pUC18 was digested using PsiI in the pglB gene, and linearized plasmid was then blunt ended using T4 DNA polymerase (Promega) according to the manufacturer’s protocol. .. The antibiotic resistance gene aphA was amplified and cut with StuI and blunt end ligated to the linearized plasmid.

Article Title: Structural and biochemical insights into the V/I505T mutation found in the EIAV gp45 vaccine strain
Article Snippet: Subsequently, the cleaved plasmids and purified PCR products were digested with a T4 DNA polymerase (Promega) in the presence of dGTP or dCTP, respectively. .. The annealed mixture containing pET30-TEV/LIC-gp45 was transformed into E.coli DH5α competent cells for plasmid propagation.

RNA Extraction:

Article Title: Transcription factor p63 bookmarks and regulates dynamic enhancers during epidermal differentiation
Article Snippet: Paragraph title: RNA extraction and RNA-seq ... After 2 h at 16°C, 1 μl of T4 DNA polymerase (10 U/μl Promega) was added and the reaction was incubated at 16°C for 10 min.

Recombinant:

Article Title: Purification and Biochemical Characterisation of Rabbit Calicivirus RNA-Dependent RNA Polymerases and Identification of Non-Nucleoside Inhibitors
Article Snippet: Resulting amplicons were cloned into the pET-26b(+) expression vector (Novagen, Darmstadt, Germany) to generate recombinant polymerases with a C-terminal His6 -tag using the following strategy. .. The pET-26b(+) expression vector was linearised using the Nde I restriction enzyme (New England BioLabs), 5′ overhangs were filled in using T4 DNA polymerase (Promega, Madison, WI, USA) to generate a blunt end and was further digested using the Xho I restriction enzyme.

In Vitro:

Article Title: Mammalian splicing factor SF1 interacts with SURP domains of U2 snRNP-associated proteins
Article Snippet: Templates for in vitro transcription of 3′ splice site pre-mRNAs were generated by HindIII and BstEII digestion of a pBluescript plasmid encoding the region spanning exons 1 and 2 of AdML pre-mRNA ( ). .. The 3′ overhangs were blunt-ended with T4 DNA polymerase (Promega) followed by religation.

Article Title: The Reovirus S4 Gene 3? Nontranslated Region Contains a Translational Operator Sequence †
Article Snippet: Plasmids were linearized by digestion with 1 U of Pst I per μl, and 3′ overhang sequences were removed by incubation with 1 to 2 U of T4 DNA polymerase (Promega) per μl at 11°C for 20 min in the presence of 1 mM Tris-HCl (pH 7.9), 1 mM MgCl2 , 5 mM NaCl, 0.1 mM dithiothreitol (DTT), 10 μM deoxynucleoside triphosphate, and 50 μg of bovine serum albumin per in a final volume of 20 μl. .. In vitro transcription of capped reovirus mRNAs was performed using linearized plasmid as template and the mMessage mMachine T7 RNA polymerase transcription system (Ambion, Austin, Tex.).

Transgenic Assay:

Article Title: Identification and Characterization of a Promoter Cassette Conferring Adipocyte-Specific Gene Expression
Article Snippet: .. To generate the adiponectin promoter-driven Cre recombinase transgenic construct, the Cre expression cassette from pBS185 was opened by Acc I digestion, which was then blunted by T4 DNA polymerase (Promega, Madison, WI). .. The Cre open reading frame fragment was then released by Xho I digestion, purified, and ligated to the 5.4-kb lac Z vector, which was linearized by Cla I digestion, blunted by T4 DNA polymerase, and opened by Xho I digestion.

Ethanol Precipitation:

Article Title: Transcription factor p63 bookmarks and regulates dynamic enhancers during epidermal differentiation
Article Snippet: Each 50-μl fragmentation reaction was incubated at 95°C for 1.5 min on a thermal cycler and placed on ice for 10 min. Ethanol precipitation was used to purify the reactions. .. After 2 h at 16°C, 1 μl of T4 DNA polymerase (10 U/μl Promega) was added and the reaction was incubated at 16°C for 10 min.

Produced:

Article Title: Simple and Efficient Method for Heterologous Expression of Clostridial Proteins
Article Snippet: .. Restriction enzymes Acc 65I, Bam HI, Bgl II, Eco 52I, Eco ICRI, Eco RI, Hin dIII, Nco I, Nde I, Sac I, Sal GI, Stu I, and Xho I, as well as T4 DNA polymerase, were produced by Promega. .. A rapid DNA ligation kit and an Expand high-fidelity PCR system were supplied by Boehringer Mannheim.

Immunoprecipitation:

Article Title: Chromatin loops associated with active genes and heterochromatin shape rice genome architecture for transcriptional regulation
Article Snippet: Centrifuge the chromatin fragments at 2000 × g for 10 min at 4 °C and transfer the supernatant to a new tube for immunoprecipitation. .. ChIP DNA was used for end-repair and A-tailing using T4 DNA polymerase (Promega, cat. no. M421F) and Klenow enzyme (NEB, cat. no. M0212L).

FLAG-tag:

Article Title: Cloning, Expression, Sequence Analysis and Homology Modeling of the Prolyl Endoprotease from Eurygaster integriceps Puton
Article Snippet: The vector includes an inducible T7 promoter, an N terminal flag epitope and a 10X His tag. .. Similarly, the amplified LIC-PEP DNA was also treated with T4 DNA polymerase (Promega Corp., Madison, WI, USA) in the presence of only dCTPs at the same concentration as the dGTP above.

Gel Extraction:

Article Title: Structural and biochemical insights into the V/I505T mutation found in the EIAV gp45 vaccine strain
Article Snippet: The pET30-TEV/LIC was digested with SspI and extracted using a gel extraction kit (Axygen). .. Subsequently, the cleaved plasmids and purified PCR products were digested with a T4 DNA polymerase (Promega) in the presence of dGTP or dCTP, respectively.

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    Promega rq1 rnase free dnase i
    Rq1 Rnase Free Dnase I, supplied by Promega, used in various techniques. Bioz Stars score: 90/100, based on 71 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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