bacteriophage t4 polynucleotide kinase  (Thermo Fisher)


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    Structured Review

    Thermo Fisher bacteriophage t4 polynucleotide kinase
    Bacteriophage T4 Polynucleotide Kinase, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/bacteriophage t4 polynucleotide kinase/product/Thermo Fisher
    Average 99 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    bacteriophage t4 polynucleotide kinase - by Bioz Stars, 2020-04
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    Centrifugation:

    Article Title: The three major types of CRISPR-Cas systems function independently in CRISPR RNA biogenesis in Streptococcus thermophilus
    Article Snippet: The cells were harvested by centrifugation at 10,000 × g for 10 minutes and lysed by bead-beating with a Mini-Beadbeater (Biospec Products). .. The 3′ ligated RNAs were gel purified away from free adaptor and treated with T4 polynucleotide kinase (Ambion) before 5′ ligation to an additional adaptor with T4 ssRNA Ligase 1 (NEB).

    Amplification:

    Article Title: The three major types of CRISPR-Cas systems function independently in CRISPR RNA biogenesis in Streptococcus thermophilus
    Article Snippet: The 3′ ligated RNAs were gel purified away from free adaptor and treated with T4 polynucleotide kinase (Ambion) before 5′ ligation to an additional adaptor with T4 ssRNA Ligase 1 (NEB). .. The 5′ and 3′ ligated RNAs were reverse transcribed with SuperScript II reverse transcriptase (Invitrogen), digested with RNase H (Promega), and PCR amplified with Crimson Taq DNA polymerase (NEB).

    DNA Synthesis:

    Article Title: Characterization of oligodeoxyribonucleotide synthesis on glass plates
    Article Snippet: DNA synthesis reagents were purchased from Glen Research (Sterling, VA). .. T4 polynucleotide kinase was purchased from Gibco.

    Mass Spectrometry:

    Article Title: Detection of Ligation Products of DNA Linkers with 5?-OH Ends by Denaturing PAGE Silver Stain
    Article Snippet: .. A quality inspection report of T4 DNA ligase from Fermentas showed that T4 PNK could not be detected in their T4 DNA ligase ( ); (iii) PNK could not be detected in T4 DNA ligase (Fermentas) by using mass spectrometry (MS) analysis ( and ); (iv) PNK is abundant in mammalian cells but absent in E. coli cells . .. Therefore, the endogenous PNK should be absent in the host E. coli cells that carry plasmids enabling T4 or E. coli DNA ligase high expression; (v) The ligation of linkers A–B and E–F could not be significantly inhibited by (NH4 )2 SO4 , a strong inhibitor of T4 PNK ( , and ); and (vi) T4 PNK requires ATP for activity.

    Positive Control:

    Article Title: Detection of Ligation Products of DNA Linkers with 5?-OH Ends by Denaturing PAGE Silver Stain
    Article Snippet: .. 25 µl of the positive control mixture contained 1 x PNK buffer A (50 mM Tris-HCl, pH 7.6 at 25°C, 10 mM MgCl2 , 5 mM DTT, 0.1 mM spermidine and 0.1 mM EDTA), 2 µM of oligo 11, 0.6 µCi/µl of [γ-32P] ATP, and 0.6 U/µl of T4 PNK (Fermentas). .. Added 25 µl of 1 x TE and 5 µl of 10% SDS to the phosphorylation products generated by T4 DNA ligase and the negative controls, and then, extracted twice with an equal volume of phenol/chloroform (1∶1), and once with chloroform/isopentyl alcohol (24∶1).

    Synthesized:

    Article Title: Four Methods of Preparing mRNA 5? End Libraries Using the Illumina Sequencing Platform
    Article Snippet: The products were then treated with T4 Polynucleotide Kinase to add mono-phosphate to non-capped mRNA to ready it for ligation; a reaction mixture consisting of 1 µl of T4 Polynucleotide Kinase (Fermentas, # EK0032), 2 µl of RNA Ligase Reaction Buffer (New England Biolabs), 0.5 µl of RNaseOUT (Invitrogen, #10777-019), 1 µl of 100 mM ATP solution (Fermentas, #R0441), and 15.5 µl of alkaline phosphatase-treated RNA was incubated for 30 minutes at 37°C. .. Next, 20 µl of T4 Polynucleotide Kinase-treated RNA were incubated with 2.5 µl of nuclease-free water, 1 µl of RNA Ligase Reaction Buffer (New England Biolabs), 4.5 µl of PEG8000 (New England Biolabs), 1 µl of STOP oligo { , STOP1 (50 µM): iGiCiG, STOP2 (50 µM): iCiGiC, STOP Mix (50 µM): mixture of STOP1 and STOP2, synthesized by Metabion, Germany}, and 1 µl of T4 RNA Ligase (New England Biolabs, M0204S) for 16 hours at 16°C to ligate STOP oligos to the non-capped mRNA.

    Article Title: Characterization of oligodeoxyribonucleotide synthesis on glass plates
    Article Snippet: T4 polynucleotide kinase was purchased from Gibco. .. Fluorescein phosphoramidite was synthesized from 9-amino-4,5-dicarboxyfluorescein according to a modified literature procedure ( , ).

    Construct:

    Article Title: Four Methods of Preparing mRNA 5? End Libraries Using the Illumina Sequencing Platform
    Article Snippet: Preparation of cDNA libraries using the CapSMART method Libraries were constructed using both ExactSTART Eukyaryotic mRNA 5′- & 3′- RACE (epicentre) and SMART cDNA library Construction Kits (Clontech), with modified SMART oligonucleotides and STOP oligos ( , ). .. The products were then treated with T4 Polynucleotide Kinase to add mono-phosphate to non-capped mRNA to ready it for ligation; a reaction mixture consisting of 1 µl of T4 Polynucleotide Kinase (Fermentas, # EK0032), 2 µl of RNA Ligase Reaction Buffer (New England Biolabs), 0.5 µl of RNaseOUT (Invitrogen, #10777-019), 1 µl of 100 mM ATP solution (Fermentas, #R0441), and 15.5 µl of alkaline phosphatase-treated RNA was incubated for 30 minutes at 37°C.

    Incubation:

    Article Title: DNA-guided DNA interference by a prokaryotic Argonaute
    Article Snippet: Proteinase K (Ambion) and CaCl2 (final concentration, 5 mM) were added to purified proteins and samples were incubated for 1 h at 37 °C. .. Purified nucleic acids were [γ-32 P]ATP labelled with T4 PNK (Fermentas) in exchange- or forward-labelling reactions and thereafter separated from free [γ-32 P] ATP using a Sephadex G-25 column (GE).

    Article Title: Sequence and Generation of Mature Ribosomal RNA Transcripts in Dictyostelium discoideum
    Article Snippet: .. After precipitation, the RNA was incubated with 20 units of T4 polynucleotide kinase (Fermentas) and 20 units of RiboLock RNase Inhibitor (Fermentas) in 30 μl of 50 m m Tris-HCl (pH 7.6), 10 m m MgCl2 , 5 m m DTT, 100 μ m spermidine, 1 m m ATP for 30 min at 37 °C. .. After protein extraction with phenol and chloroform, RNA was precipitated with ethanol.

    Article Title: Four Methods of Preparing mRNA 5? End Libraries Using the Illumina Sequencing Platform
    Article Snippet: .. The products were then treated with T4 Polynucleotide Kinase to add mono-phosphate to non-capped mRNA to ready it for ligation; a reaction mixture consisting of 1 µl of T4 Polynucleotide Kinase (Fermentas, # EK0032), 2 µl of RNA Ligase Reaction Buffer (New England Biolabs), 0.5 µl of RNaseOUT (Invitrogen, #10777-019), 1 µl of 100 mM ATP solution (Fermentas, #R0441), and 15.5 µl of alkaline phosphatase-treated RNA was incubated for 30 minutes at 37°C. .. Next, 20 µl of T4 Polynucleotide Kinase-treated RNA were incubated with 2.5 µl of nuclease-free water, 1 µl of RNA Ligase Reaction Buffer (New England Biolabs), 4.5 µl of PEG8000 (New England Biolabs), 1 µl of STOP oligo { , STOP1 (50 µM): iGiCiG, STOP2 (50 µM): iCiGiC, STOP Mix (50 µM): mixture of STOP1 and STOP2, synthesized by Metabion, Germany}, and 1 µl of T4 RNA Ligase (New England Biolabs, M0204S) for 16 hours at 16°C to ligate STOP oligos to the non-capped mRNA.

    Article Title: Road blocks on paleogenomes--polymerase extension profiling reveals the frequency of blocking lesions in ancient DNA
    Article Snippet: .. PEP assays and sequencing For blunt end repair, ∼15 ng of PCR product pool, 2 ng of fragmented horse DNA, 4 ng of UV-irradiated horse DNA, 10 µl ancient DNA extract or a water sample were incubated for 15 min at 12°C and 15 min at 25°C in a 40 µl reaction containing in final concentrations 1× Tango buffer, 0.1 U/µl T4 DNA polymerase, 0.5 U/µl T4 polynucleotide kinase (all Fermentas), 1 mM ATP and 0.1 mM dNTP. .. Reactions were purified using the MinElute PCR Purification kit.

    Article Title: Sequence and Generation of Mature Ribosomal RNA Transcripts in Dictyostelium discoideum
    Article Snippet: As probes, DNA oligonucleotides were 5′ end-labeled by incubating 10 pmol of primer with 10 units of T4 polynucleotide kinase (Fermentas) in 50 m m Tris-HCl (pH 7.6), 10 m m MgCl2 , 5 m m DTT, 100 μ m spermidine, and 0.37 MBq of [γ-32 P]ATP. .. The complete reaction volume was placed on a Sephadex G-50 column (Fluka) and centrifuged at 1000 rpm for 3 min. After prehybridization of the membrane with Church buffer containing 250 m m (Na+ and H+ ) phosphate buffer (pH 7.0), 1 m m EDTA, 1% (w/v) BSA, and 7% (w/v) SDS for at least 1 h at 42 °C, the purified oligonucleotide probe was added and incubated at 42 °C overnight.

    Activity Assay:

    Article Title: Detection of Ligation Products of DNA Linkers with 5?-OH Ends by Denaturing PAGE Silver Stain
    Article Snippet: A quality inspection report of T4 DNA ligase from Fermentas showed that T4 PNK could not be detected in their T4 DNA ligase ( ); (iii) PNK could not be detected in T4 DNA ligase (Fermentas) by using mass spectrometry (MS) analysis ( and ); (iv) PNK is abundant in mammalian cells but absent in E. coli cells . .. Therefore, the endogenous PNK should be absent in the host E. coli cells that carry plasmids enabling T4 or E. coli DNA ligase high expression; (v) The ligation of linkers A–B and E–F could not be significantly inhibited by (NH4 )2 SO4 , a strong inhibitor of T4 PNK ( , and ); and (vi) T4 PNK requires ATP for activity.

    Expressing:

    Article Title: The three major types of CRISPR-Cas systems function independently in CRISPR RNA biogenesis in Streptococcus thermophilus
    Article Snippet: RNA libraries used to examine crRNA expression were produced as described previously ( ) except that no size-selection was done prior to RNA manipulation. .. The 3′ ligated RNAs were gel purified away from free adaptor and treated with T4 polynucleotide kinase (Ambion) before 5′ ligation to an additional adaptor with T4 ssRNA Ligase 1 (NEB).

    Article Title: Detection of Ligation Products of DNA Linkers with 5?-OH Ends by Denaturing PAGE Silver Stain
    Article Snippet: A quality inspection report of T4 DNA ligase from Fermentas showed that T4 PNK could not be detected in their T4 DNA ligase ( ); (iii) PNK could not be detected in T4 DNA ligase (Fermentas) by using mass spectrometry (MS) analysis ( and ); (iv) PNK is abundant in mammalian cells but absent in E. coli cells . .. Therefore, the endogenous PNK should be absent in the host E. coli cells that carry plasmids enabling T4 or E. coli DNA ligase high expression; (v) The ligation of linkers A–B and E–F could not be significantly inhibited by (NH4 )2 SO4 , a strong inhibitor of T4 PNK ( , and ); and (vi) T4 PNK requires ATP for activity.

    Modification:

    Article Title: Four Methods of Preparing mRNA 5? End Libraries Using the Illumina Sequencing Platform
    Article Snippet: Preparation of cDNA libraries using the CapSMART method Libraries were constructed using both ExactSTART Eukyaryotic mRNA 5′- & 3′- RACE (epicentre) and SMART cDNA library Construction Kits (Clontech), with modified SMART oligonucleotides and STOP oligos ( , ). .. The products were then treated with T4 Polynucleotide Kinase to add mono-phosphate to non-capped mRNA to ready it for ligation; a reaction mixture consisting of 1 µl of T4 Polynucleotide Kinase (Fermentas, # EK0032), 2 µl of RNA Ligase Reaction Buffer (New England Biolabs), 0.5 µl of RNaseOUT (Invitrogen, #10777-019), 1 µl of 100 mM ATP solution (Fermentas, #R0441), and 15.5 µl of alkaline phosphatase-treated RNA was incubated for 30 minutes at 37°C.

    Article Title: Characterization of oligodeoxyribonucleotide synthesis on glass plates
    Article Snippet: T4 polynucleotide kinase was purchased from Gibco. .. Fluorescein phosphoramidite was synthesized from 9-amino-4,5-dicarboxyfluorescein according to a modified literature procedure ( , ).

    Kinase Assay:

    Article Title: Detection of Ligation Products of DNA Linkers with 5?-OH Ends by Denaturing PAGE Silver Stain
    Article Snippet: Paragraph title: Kinase Assay for T4 DNA Ligase ... 25 µl of the positive control mixture contained 1 x PNK buffer A (50 mM Tris-HCl, pH 7.6 at 25°C, 10 mM MgCl2 , 5 mM DTT, 0.1 mM spermidine and 0.1 mM EDTA), 2 µM of oligo 11, 0.6 µCi/µl of [γ-32P] ATP, and 0.6 U/µl of T4 PNK (Fermentas).

    Ligation:

    Article Title: Detection of Ligation Products of DNA Linkers with 5?-OH Ends by Denaturing PAGE Silver Stain
    Article Snippet: Kinase Assay for T4 DNA Ligase To explore the ligation mechanism of DNA linkers with 5′-OH ends, oligo 11 of linker F were phosphorylated by using [γ-32P] ATP and T4 DNA ligase. .. 25 µl of the positive control mixture contained 1 x PNK buffer A (50 mM Tris-HCl, pH 7.6 at 25°C, 10 mM MgCl2 , 5 mM DTT, 0.1 mM spermidine and 0.1 mM EDTA), 2 µM of oligo 11, 0.6 µCi/µl of [γ-32P] ATP, and 0.6 U/µl of T4 PNK (Fermentas).

    Article Title: The three major types of CRISPR-Cas systems function independently in CRISPR RNA biogenesis in Streptococcus thermophilus
    Article Snippet: .. The 3′ ligated RNAs were gel purified away from free adaptor and treated with T4 polynucleotide kinase (Ambion) before 5′ ligation to an additional adaptor with T4 ssRNA Ligase 1 (NEB). .. The 5′ and 3′ ligated RNAs were reverse transcribed with SuperScript II reverse transcriptase (Invitrogen), digested with RNase H (Promega), and PCR amplified with Crimson Taq DNA polymerase (NEB).

    Article Title: Four Methods of Preparing mRNA 5? End Libraries Using the Illumina Sequencing Platform
    Article Snippet: .. The products were then treated with T4 Polynucleotide Kinase to add mono-phosphate to non-capped mRNA to ready it for ligation; a reaction mixture consisting of 1 µl of T4 Polynucleotide Kinase (Fermentas, # EK0032), 2 µl of RNA Ligase Reaction Buffer (New England Biolabs), 0.5 µl of RNaseOUT (Invitrogen, #10777-019), 1 µl of 100 mM ATP solution (Fermentas, #R0441), and 15.5 µl of alkaline phosphatase-treated RNA was incubated for 30 minutes at 37°C. .. Next, 20 µl of T4 Polynucleotide Kinase-treated RNA were incubated with 2.5 µl of nuclease-free water, 1 µl of RNA Ligase Reaction Buffer (New England Biolabs), 4.5 µl of PEG8000 (New England Biolabs), 1 µl of STOP oligo { , STOP1 (50 µM): iGiCiG, STOP2 (50 µM): iCiGiC, STOP Mix (50 µM): mixture of STOP1 and STOP2, synthesized by Metabion, Germany}, and 1 µl of T4 RNA Ligase (New England Biolabs, M0204S) for 16 hours at 16°C to ligate STOP oligos to the non-capped mRNA.

    Article Title: Detection of Ligation Products of DNA Linkers with 5?-OH Ends by Denaturing PAGE Silver Stain
    Article Snippet: A quality inspection report of T4 DNA ligase from Fermentas showed that T4 PNK could not be detected in their T4 DNA ligase ( ); (iii) PNK could not be detected in T4 DNA ligase (Fermentas) by using mass spectrometry (MS) analysis ( and ); (iv) PNK is abundant in mammalian cells but absent in E. coli cells . .. Therefore, the endogenous PNK should be absent in the host E. coli cells that carry plasmids enabling T4 or E. coli DNA ligase high expression; (v) The ligation of linkers A–B and E–F could not be significantly inhibited by (NH4 )2 SO4 , a strong inhibitor of T4 PNK ( , and ); and (vi) T4 PNK requires ATP for activity.

    Northern Blot:

    Article Title: Sequence and Generation of Mature Ribosomal RNA Transcripts in Dictyostelium discoideum
    Article Snippet: Paragraph title: Northern Analysis ... As probes, DNA oligonucleotides were 5′ end-labeled by incubating 10 pmol of primer with 10 units of T4 polynucleotide kinase (Fermentas) in 50 m m Tris-HCl (pH 7.6), 10 m m MgCl2 , 5 m m DTT, 100 μ m spermidine, and 0.37 MBq of [γ-32 P]ATP.

    Oligonucleotide Labeling:

    Article Title: An In Vitro DNA Double-Strand Break Repair Assay Based on End-Joining of Defined Duplex Oligonucleotides
    Article Snippet: Paragraph title: 2.3. Oligonucleotide Labeling and Duplex Formation ... T4 polynucleotide kinase and 10× buffer (Fermentas; Hanover, MD).

    Generated:

    Article Title: Detection of Ligation Products of DNA Linkers with 5?-OH Ends by Denaturing PAGE Silver Stain
    Article Snippet: 25 µl of the positive control mixture contained 1 x PNK buffer A (50 mM Tris-HCl, pH 7.6 at 25°C, 10 mM MgCl2 , 5 mM DTT, 0.1 mM spermidine and 0.1 mM EDTA), 2 µM of oligo 11, 0.6 µCi/µl of [γ-32P] ATP, and 0.6 U/µl of T4 PNK (Fermentas). .. Added 25 µl of 1 x TE and 5 µl of 10% SDS to the phosphorylation products generated by T4 DNA ligase and the negative controls, and then, extracted twice with an equal volume of phenol/chloroform (1∶1), and once with chloroform/isopentyl alcohol (24∶1).

    Article Title: The three major types of CRISPR-Cas systems function independently in CRISPR RNA biogenesis in Streptococcus thermophilus
    Article Snippet: The csn1-1 ( cas9 ) and csn2-1 gene disruption strains were generated in a previous study ( ). .. The 3′ ligated RNAs were gel purified away from free adaptor and treated with T4 polynucleotide kinase (Ambion) before 5′ ligation to an additional adaptor with T4 ssRNA Ligase 1 (NEB).

    Article Title: Road blocks on paleogenomes--polymerase extension profiling reveals the frequency of blocking lesions in ancient DNA
    Article Snippet: PEP assays and sequencing For blunt end repair, ∼15 ng of PCR product pool, 2 ng of fragmented horse DNA, 4 ng of UV-irradiated horse DNA, 10 µl ancient DNA extract or a water sample were incubated for 15 min at 12°C and 15 min at 25°C in a 40 µl reaction containing in final concentrations 1× Tango buffer, 0.1 U/µl T4 DNA polymerase, 0.5 U/µl T4 polynucleotide kinase (all Fermentas), 1 mM ATP and 0.1 mM dNTP. .. An adapter (B-BL) was generated by annealing two partially complementary oligonucleotides (B-BL1 and B-BL2; see Supplementary Table S3 for the sequences) in a reaction containing 1× T4 ligase buffer (Fermentas) and 200 µM of each oligonucleotide.

    Polymerase Chain Reaction:

    Article Title: The three major types of CRISPR-Cas systems function independently in CRISPR RNA biogenesis in Streptococcus thermophilus
    Article Snippet: The 3′ ligated RNAs were gel purified away from free adaptor and treated with T4 polynucleotide kinase (Ambion) before 5′ ligation to an additional adaptor with T4 ssRNA Ligase 1 (NEB). .. The 5′ and 3′ ligated RNAs were reverse transcribed with SuperScript II reverse transcriptase (Invitrogen), digested with RNase H (Promega), and PCR amplified with Crimson Taq DNA polymerase (NEB).

    Article Title: Road blocks on paleogenomes--polymerase extension profiling reveals the frequency of blocking lesions in ancient DNA
    Article Snippet: .. PEP assays and sequencing For blunt end repair, ∼15 ng of PCR product pool, 2 ng of fragmented horse DNA, 4 ng of UV-irradiated horse DNA, 10 µl ancient DNA extract or a water sample were incubated for 15 min at 12°C and 15 min at 25°C in a 40 µl reaction containing in final concentrations 1× Tango buffer, 0.1 U/µl T4 DNA polymerase, 0.5 U/µl T4 polynucleotide kinase (all Fermentas), 1 mM ATP and 0.1 mM dNTP. .. Reactions were purified using the MinElute PCR Purification kit.

    Copurification:

    Article Title: DNA-guided DNA interference by a prokaryotic Argonaute
    Article Snippet: Paragraph title: Guide co-purification and sequencing ... Purified nucleic acids were [γ-32 P]ATP labelled with T4 PNK (Fermentas) in exchange- or forward-labelling reactions and thereafter separated from free [γ-32 P] ATP using a Sephadex G-25 column (GE).

    Radioactivity:

    Article Title: DNA-guided DNA interference by a prokaryotic Argonaute
    Article Snippet: Purified nucleic acids were [γ-32 P]ATP labelled with T4 PNK (Fermentas) in exchange- or forward-labelling reactions and thereafter separated from free [γ-32 P] ATP using a Sephadex G-25 column (GE). .. Radioactivity was captured from gels using phosphor screens.

    RNA Sequencing Assay:

    Article Title: The three major types of CRISPR-Cas systems function independently in CRISPR RNA biogenesis in Streptococcus thermophilus
    Article Snippet: The 3′ ligated RNAs were gel purified away from free adaptor and treated with T4 polynucleotide kinase (Ambion) before 5′ ligation to an additional adaptor with T4 ssRNA Ligase 1 (NEB). .. For assessing cas gene mRNA expression profiles associated with each of the four CRISPR-Cas loci, including possible readthrough of Cas gene expression into each CRISPR locus , strand-specific RNA-seq libraries were prepared using prerelease Directional RNA-Seq Library Kits (Illumina).

    Isolation:

    Article Title: The three major types of CRISPR-Cas systems function independently in CRISPR RNA biogenesis in Streptococcus thermophilus
    Article Snippet: Paragraph title: RNA isolation and RNA library preparation ... The 3′ ligated RNAs were gel purified away from free adaptor and treated with T4 polynucleotide kinase (Ambion) before 5′ ligation to an additional adaptor with T4 ssRNA Ligase 1 (NEB).

    Labeling:

    Article Title: Detection of Ligation Products of DNA Linkers with 5?-OH Ends by Denaturing PAGE Silver Stain
    Article Snippet: The kinase assay for E. coli DNA ligase was not performed because NAD+ labeled with 32P was not available for us. .. 25 µl of the positive control mixture contained 1 x PNK buffer A (50 mM Tris-HCl, pH 7.6 at 25°C, 10 mM MgCl2 , 5 mM DTT, 0.1 mM spermidine and 0.1 mM EDTA), 2 µM of oligo 11, 0.6 µCi/µl of [γ-32P] ATP, and 0.6 U/µl of T4 PNK (Fermentas).

    Article Title: Characterization of oligodeoxyribonucleotide synthesis on glass plates
    Article Snippet: .. A portion of the sample (3 µl) was labeled with [γ-32 P]ATP (5 µCi, 3000 Ci/mmol) using T4 polynucleotide kinase (1 U) and the conditions recommended by the manufacturer (Gibco). ..

    Purification:

    Article Title: DNA-guided DNA interference by a prokaryotic Argonaute
    Article Snippet: .. Purified nucleic acids were [γ-32 P]ATP labelled with T4 PNK (Fermentas) in exchange- or forward-labelling reactions and thereafter separated from free [γ-32 P] ATP using a Sephadex G-25 column (GE). .. Labelled nucleic acids were incubated with nucleases (DNase-free RNase A (Fermentas),RQ1 RNase-free DNaseI (Promega) or P1 nuclease (Sigma)) for 1 h at 37 °C.

    Article Title: Cleavage of poly(A) tails on the 3?-end of RNA by ribonuclease E of Escherichia coli
    Article Snippet: Four to five pmoles of RNA that had been either chemically synthesised or transcribed in vitro and then dephosphorylated was radiolabelled at the 5′-end by incubating with 23 pmol (160 µCi) [γ-32 P]ATP (ICN) and 10 U T4 polynucleotide kinase (MBI Fermentas) as described previously ( ). .. The reactions were quenched with urea loading buffer [7 M urea, 0.1% (w/v) bromophenol blue and xylene cyanol] and the 5′-labelled RNAs were gel purified ( ).

    Article Title: The three major types of CRISPR-Cas systems function independently in CRISPR RNA biogenesis in Streptococcus thermophilus
    Article Snippet: .. The 3′ ligated RNAs were gel purified away from free adaptor and treated with T4 polynucleotide kinase (Ambion) before 5′ ligation to an additional adaptor with T4 ssRNA Ligase 1 (NEB). .. The 5′ and 3′ ligated RNAs were reverse transcribed with SuperScript II reverse transcriptase (Invitrogen), digested with RNase H (Promega), and PCR amplified with Crimson Taq DNA polymerase (NEB).

    Article Title: Four Methods of Preparing mRNA 5? End Libraries Using the Illumina Sequencing Platform
    Article Snippet: After the reaction, the products were purified using the RNeasy MinElute Cleanup kit (Qiagen), and eluted with 18 µl nuclease-free water. .. The products were then treated with T4 Polynucleotide Kinase to add mono-phosphate to non-capped mRNA to ready it for ligation; a reaction mixture consisting of 1 µl of T4 Polynucleotide Kinase (Fermentas, # EK0032), 2 µl of RNA Ligase Reaction Buffer (New England Biolabs), 0.5 µl of RNaseOUT (Invitrogen, #10777-019), 1 µl of 100 mM ATP solution (Fermentas, #R0441), and 15.5 µl of alkaline phosphatase-treated RNA was incubated for 30 minutes at 37°C.

    Article Title: Road blocks on paleogenomes--polymerase extension profiling reveals the frequency of blocking lesions in ancient DNA
    Article Snippet: PEP assays and sequencing For blunt end repair, ∼15 ng of PCR product pool, 2 ng of fragmented horse DNA, 4 ng of UV-irradiated horse DNA, 10 µl ancient DNA extract or a water sample were incubated for 15 min at 12°C and 15 min at 25°C in a 40 µl reaction containing in final concentrations 1× Tango buffer, 0.1 U/µl T4 DNA polymerase, 0.5 U/µl T4 polynucleotide kinase (all Fermentas), 1 mM ATP and 0.1 mM dNTP. .. Reactions were purified using the MinElute PCR Purification kit.

    Article Title: Sequence and Generation of Mature Ribosomal RNA Transcripts in Dictyostelium discoideum
    Article Snippet: As probes, DNA oligonucleotides were 5′ end-labeled by incubating 10 pmol of primer with 10 units of T4 polynucleotide kinase (Fermentas) in 50 m m Tris-HCl (pH 7.6), 10 m m MgCl2 , 5 m m DTT, 100 μ m spermidine, and 0.37 MBq of [γ-32 P]ATP. .. The complete reaction volume was placed on a Sephadex G-50 column (Fluka) and centrifuged at 1000 rpm for 3 min. After prehybridization of the membrane with Church buffer containing 250 m m (Na+ and H+ ) phosphate buffer (pH 7.0), 1 m m EDTA, 1% (w/v) BSA, and 7% (w/v) SDS for at least 1 h at 42 °C, the purified oligonucleotide probe was added and incubated at 42 °C overnight.

    Sequencing:

    Article Title: DNA-guided DNA interference by a prokaryotic Argonaute
    Article Snippet: Paragraph title: Guide co-purification and sequencing ... Purified nucleic acids were [γ-32 P]ATP labelled with T4 PNK (Fermentas) in exchange- or forward-labelling reactions and thereafter separated from free [γ-32 P] ATP using a Sephadex G-25 column (GE).

    Article Title: Road blocks on paleogenomes--polymerase extension profiling reveals the frequency of blocking lesions in ancient DNA
    Article Snippet: .. PEP assays and sequencing For blunt end repair, ∼15 ng of PCR product pool, 2 ng of fragmented horse DNA, 4 ng of UV-irradiated horse DNA, 10 µl ancient DNA extract or a water sample were incubated for 15 min at 12°C and 15 min at 25°C in a 40 µl reaction containing in final concentrations 1× Tango buffer, 0.1 U/µl T4 DNA polymerase, 0.5 U/µl T4 polynucleotide kinase (all Fermentas), 1 mM ATP and 0.1 mM dNTP. .. Reactions were purified using the MinElute PCR Purification kit.

    Protein Extraction:

    Article Title: Sequence and Generation of Mature Ribosomal RNA Transcripts in Dictyostelium discoideum
    Article Snippet: After precipitation, the RNA was incubated with 20 units of T4 polynucleotide kinase (Fermentas) and 20 units of RiboLock RNase Inhibitor (Fermentas) in 30 μl of 50 m m Tris-HCl (pH 7.6), 10 m m MgCl2 , 5 m m DTT, 100 μ m spermidine, 1 m m ATP for 30 min at 37 °C. .. After protein extraction with phenol and chloroform, RNA was precipitated with ethanol.

    CRISPR:

    Article Title: The three major types of CRISPR-Cas systems function independently in CRISPR RNA biogenesis in Streptococcus thermophilus
    Article Snippet: The 3′ ligated RNAs were gel purified away from free adaptor and treated with T4 polynucleotide kinase (Ambion) before 5′ ligation to an additional adaptor with T4 ssRNA Ligase 1 (NEB). .. For assessing cas gene mRNA expression profiles associated with each of the four CRISPR-Cas loci, including possible readthrough of Cas gene expression into each CRISPR locus , strand-specific RNA-seq libraries were prepared using prerelease Directional RNA-Seq Library Kits (Illumina).

    cDNA Library Assay:

    Article Title: Four Methods of Preparing mRNA 5? End Libraries Using the Illumina Sequencing Platform
    Article Snippet: Preparation of cDNA libraries using the CapSMART method Libraries were constructed using both ExactSTART Eukyaryotic mRNA 5′- & 3′- RACE (epicentre) and SMART cDNA library Construction Kits (Clontech), with modified SMART oligonucleotides and STOP oligos ( , ). .. The products were then treated with T4 Polynucleotide Kinase to add mono-phosphate to non-capped mRNA to ready it for ligation; a reaction mixture consisting of 1 µl of T4 Polynucleotide Kinase (Fermentas, # EK0032), 2 µl of RNA Ligase Reaction Buffer (New England Biolabs), 0.5 µl of RNaseOUT (Invitrogen, #10777-019), 1 µl of 100 mM ATP solution (Fermentas, #R0441), and 15.5 µl of alkaline phosphatase-treated RNA was incubated for 30 minutes at 37°C.

    Sample Prep:

    Article Title: Characterization of oligodeoxyribonucleotide synthesis on glass plates
    Article Snippet: Paragraph title: Sample preparation and 32 P-labeling ... A portion of the sample (3 µl) was labeled with [γ-32 P]ATP (5 µCi, 3000 Ci/mmol) using T4 polynucleotide kinase (1 U) and the conditions recommended by the manufacturer (Gibco).

    In Vitro:

    Article Title: Cleavage of poly(A) tails on the 3?-end of RNA by ribonuclease E of Escherichia coli
    Article Snippet: .. Four to five pmoles of RNA that had been either chemically synthesised or transcribed in vitro and then dephosphorylated was radiolabelled at the 5′-end by incubating with 23 pmol (160 µCi) [γ-32 P]ATP (ICN) and 10 U T4 polynucleotide kinase (MBI Fermentas) as described previously ( ). .. The reactions were quenched with urea loading buffer [7 M urea, 0.1% (w/v) bromophenol blue and xylene cyanol] and the 5′-labelled RNAs were gel purified ( ).

    Ethanol Precipitation:

    Article Title: DNA-guided DNA interference by a prokaryotic Argonaute
    Article Snippet: Nucleic acids were separated from protein content using Roti phenol/chloroform/isoamyl alcohol pH 7.5–8.0 (Carl Roth GmbH) and further purified by ethanol precipitation. .. Purified nucleic acids were [γ-32 P]ATP labelled with T4 PNK (Fermentas) in exchange- or forward-labelling reactions and thereafter separated from free [γ-32 P] ATP using a Sephadex G-25 column (GE).

    Ancient DNA Assay:

    Article Title: Road blocks on paleogenomes--polymerase extension profiling reveals the frequency of blocking lesions in ancient DNA
    Article Snippet: .. PEP assays and sequencing For blunt end repair, ∼15 ng of PCR product pool, 2 ng of fragmented horse DNA, 4 ng of UV-irradiated horse DNA, 10 µl ancient DNA extract or a water sample were incubated for 15 min at 12°C and 15 min at 25°C in a 40 µl reaction containing in final concentrations 1× Tango buffer, 0.1 U/µl T4 DNA polymerase, 0.5 U/µl T4 polynucleotide kinase (all Fermentas), 1 mM ATP and 0.1 mM dNTP. .. Reactions were purified using the MinElute PCR Purification kit.

    Produced:

    Article Title: The three major types of CRISPR-Cas systems function independently in CRISPR RNA biogenesis in Streptococcus thermophilus
    Article Snippet: RNA libraries used to examine crRNA expression were produced as described previously ( ) except that no size-selection was done prior to RNA manipulation. .. The 3′ ligated RNAs were gel purified away from free adaptor and treated with T4 polynucleotide kinase (Ambion) before 5′ ligation to an additional adaptor with T4 ssRNA Ligase 1 (NEB).

    Article Title: Detection of Ligation Products of DNA Linkers with 5?-OH Ends by Denaturing PAGE Silver Stain
    Article Snippet: When these manufacturers were questioned, they stated that their T4 DNA ligases had very high quality and it was very unlikely that there would be PNK in their ligases because their ligases were produced by using E. coli cells and production lines that were different from those for T4 PNK. .. A quality inspection report of T4 DNA ligase from Fermentas showed that T4 PNK could not be detected in their T4 DNA ligase ( ); (iii) PNK could not be detected in T4 DNA ligase (Fermentas) by using mass spectrometry (MS) analysis ( and ); (iv) PNK is abundant in mammalian cells but absent in E. coli cells .

    Concentration Assay:

    Article Title: DNA-guided DNA interference by a prokaryotic Argonaute
    Article Snippet: Proteinase K (Ambion) and CaCl2 (final concentration, 5 mM) were added to purified proteins and samples were incubated for 1 h at 37 °C. .. Purified nucleic acids were [γ-32 P]ATP labelled with T4 PNK (Fermentas) in exchange- or forward-labelling reactions and thereafter separated from free [γ-32 P] ATP using a Sephadex G-25 column (GE).

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  • 99
    Thermo Fisher phage t4 polynucleotide kinase
    Phage T4 Polynucleotide Kinase, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 2 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/phage t4 polynucleotide kinase/product/Thermo Fisher
    Average 99 stars, based on 2 article reviews
    Price from $9.99 to $1999.99
    phage t4 polynucleotide kinase - by Bioz Stars, 2020-04
    99/100 stars
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