dntpack  (Millipore)

 
  • Logo
  • About
  • News
  • Press Release
  • Team
  • Advisors
  • Partners
  • Contact
  • Bioz Stars
  • Bioz vStars
  • 93
    Name:
    FastStart High Fidelity PCR System
    Description:
    The enzyme dNTPack comprises the FastStart High Fidelity PCR System and a ready to use solution of PCR grade nucleotides The FastStart High Fidelity PCR System is a blend of FastStart Taq DNA Polymerase and a thermostable proofreading protein which is also chemically modified This protein mediates proofreading activity but carries no polymerase activity Both proteins are inactive below 75C and are activated by heating to 95C for two minutes The FastStart R High Fidelity PCR System is the ideal tool for all amplification experiments where a combination of high specificity sensitivity accuracy and yield is needed Hot start protocols have improved specificity sensitivity and PCR yield Heat labile blocking groups on amino acid residues of FastStart Taq DNA Polymerase make the enzyme inactive at room temperature There is no elongation at low temperatures when primers bind nonspecifically
    Catalog Number:
    FHIFIN-RO
    Price:
    None
    Applications:
    The FastStart(R) High Fidelity PCR System - an optimized enzyme blend for hot start PCR - combines all the benefits of FastStart Taq DNA Polymerase with fourfold higher accuracy and the ability to amplify fragments up to 5kb.. Hot start PCR. Multiplex PCR. Hot Start RT-PCR. GC-rich template amplification. Difficult or challenging PCRsThe FastStart High Fidelity PCR System is the product of choice for multiplex PCR.
    Buy from Supplier


    Structured Review

    Millipore dntpack
    FastStart High Fidelity PCR System
    The enzyme dNTPack comprises the FastStart High Fidelity PCR System and a ready to use solution of PCR grade nucleotides The FastStart High Fidelity PCR System is a blend of FastStart Taq DNA Polymerase and a thermostable proofreading protein which is also chemically modified This protein mediates proofreading activity but carries no polymerase activity Both proteins are inactive below 75C and are activated by heating to 95C for two minutes The FastStart R High Fidelity PCR System is the ideal tool for all amplification experiments where a combination of high specificity sensitivity accuracy and yield is needed Hot start protocols have improved specificity sensitivity and PCR yield Heat labile blocking groups on amino acid residues of FastStart Taq DNA Polymerase make the enzyme inactive at room temperature There is no elongation at low temperatures when primers bind nonspecifically
    https://www.bioz.com/result/dntpack/product/Millipore
    Average 93 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    dntpack - by Bioz Stars, 2021-07
    93/100 stars

    Images

    Related Articles

    Polymerase Chain Reaction:

    Article Title: Naturally occurring SARS-CoV-2 gene deletions close to the spike S1/S2 cleavage site in the viral quasispecies of COVID19 patients.
    Article Snippet: The second round of amplification (nested) was done using overlapping internal primer pairs to amplify fragments 470bp to 313bp in length. .. The FastStart High-Fidelity PCR System dNTPack (Sigma, St. Louis, MO, CA) was used for this purpose, as follows: activation at 94ºC for 4 min, followed by 30 cycles with denaturation at 94ºC for 30 sec, annealing at 55ºC for 30 sec, and elongation at 72ºC for 40 sec, ending with a single elongation step at 72ºC for 7 min. PCR products were purified using the QIAquick Gel Extraction Kit (Qiagen, Hilden, Germany) with QG buffer, following the manufacturers’ instructions, and eluted DNA was quantified by fluorometry using the QUBIT dsDNA BR Assay Kit (ThermoFisher, MA, USA). .. For each patient, PCR products were normalized to 1.5 ng/µL, pooled in a single tube, and purified using KAPA Pure Beads (KapaBiosystems, Roche, Pleasanton, CA, USA) to ensure that no short DNA fragments were present in the library.

    Article Title: DNA methylation clocks as a predictor for ageing and age estimation in naked mole-rats, Heterocephalus glaber
    Article Snippet: DNA was bisulfite converted in a 96 well plate format using the EZ-96 DNA Methylation™ Kit (Zymo, Cat. D5003). .. Target amplification was performed using the FastStart High Fidelity PCR System, dNTPack (Sigma-Aldrich, Cat. 4738284001) in the 48.48 layout on the Fluidigm® C1 system (Fluidigm®, USA), a microfluidics platform. .. Library preparation was performed using the same kit including 4 μl of Access Array Barcode Library Primer and 1 μl of PCR product diluted 1:100.

    Article Title: Naturally occurring SARS-CoV-2 gene deletions close to the spike S1/S2 cleavage site in the viral quasispecies of COVID19 patients
    Article Snippet: The second round of amplification (nested) was done using overlapping internal primer pairs to amplify fragments 470bp to 313bp in length. .. The FastStart High-Fidelity PCR System dNTPack (Sigma, St. Louis, MO, CA) was used for this purpose, as follows: activation at 94°C for 4 min, followed by 30 cycles with denaturation at 94°C for 30 sec, annealing at 55°C for 30 sec, and elongation at 72°C for 40 sec, ending with a single elongation step at 72°C for 7 min. PCR products were purified using the QIAquick Gel Extraction Kit (Qiagen, Hilden, Germany) with QG buffer, following the manufacturers’ instructions, and eluted DNA was quantified by fluorometry using the QUBIT dsDNA BR Assay Kit (ThermoFisher, MA, USA). .. For each patient, PCR products were normalized to 1.5 ng/μL, pooled in a single tube, and purified using KAPA Pure Beads (KapaBiosystems, Roche, Pleasanton, CA, USA) to ensure that no short DNA fragments were present in the library.

    Article Title: Whole-genome characterization and resistance-associated substitutions in a new HCV genotype 1 subtype
    Article Snippet: .. All primers were numbered according to isolate H77 (accession number AF009606) The external fragments were amplified using the Transcriptor One Step RT-PCR Kit (#04655885001 Sigma, Roche), and for the internals, the FastStart High Fidelity PCR System, dNTPack (#04738292001 Sigma, Roche), adapting the various PCR programs to the lengths and temperatures of each specific primer pair. .. Due to the high variability of the region corresponding to internal fragment number 11, a random priming (#11034731001 Sigma, Roche) PCR was necessary to perform between the RT-PCR and the nested PCR with the specific primers, to obtain enough DNA for analysis.

    Article Title: Whole-genome characterization and resistance-associated substitutions in a new HCV genotype 1 subtype
    Article Snippet: To characterize any naturally occurring baseline RAS in the putative new subtype taking into account that any isolate is composed by a population of sequences, four new internal primers pairs were designed, corresponding to the three DAA-targeted regions of the HCV genome (1 NS3, 1 NS5A and 2 NS5B); using this time the isolate's full genome sequence, obtained with Sanger ssequencing, was used as reference (Table S1). .. To amplify these new fragments, we used the products of the external RT-PCRs obtained previously on the FastStart High Fidelity PCR System, dNTPack (#04738292001 Sigma, Roche), adapting the thermocycler programs to the characteristics of each different primer pair. .. Again all fragments were checked and purified on 1.5% agarose gel, and were deep-sequenced using the MiSeq Reagent Kit v3 (600 cycle) (#MS-102–3003, Illumina) according to a previously described method.

    Activation Assay:

    Article Title: Naturally occurring SARS-CoV-2 gene deletions close to the spike S1/S2 cleavage site in the viral quasispecies of COVID19 patients.
    Article Snippet: The second round of amplification (nested) was done using overlapping internal primer pairs to amplify fragments 470bp to 313bp in length. .. The FastStart High-Fidelity PCR System dNTPack (Sigma, St. Louis, MO, CA) was used for this purpose, as follows: activation at 94ºC for 4 min, followed by 30 cycles with denaturation at 94ºC for 30 sec, annealing at 55ºC for 30 sec, and elongation at 72ºC for 40 sec, ending with a single elongation step at 72ºC for 7 min. PCR products were purified using the QIAquick Gel Extraction Kit (Qiagen, Hilden, Germany) with QG buffer, following the manufacturers’ instructions, and eluted DNA was quantified by fluorometry using the QUBIT dsDNA BR Assay Kit (ThermoFisher, MA, USA). .. For each patient, PCR products were normalized to 1.5 ng/µL, pooled in a single tube, and purified using KAPA Pure Beads (KapaBiosystems, Roche, Pleasanton, CA, USA) to ensure that no short DNA fragments were present in the library.

    Article Title: Naturally occurring SARS-CoV-2 gene deletions close to the spike S1/S2 cleavage site in the viral quasispecies of COVID19 patients
    Article Snippet: The second round of amplification (nested) was done using overlapping internal primer pairs to amplify fragments 470bp to 313bp in length. .. The FastStart High-Fidelity PCR System dNTPack (Sigma, St. Louis, MO, CA) was used for this purpose, as follows: activation at 94°C for 4 min, followed by 30 cycles with denaturation at 94°C for 30 sec, annealing at 55°C for 30 sec, and elongation at 72°C for 40 sec, ending with a single elongation step at 72°C for 7 min. PCR products were purified using the QIAquick Gel Extraction Kit (Qiagen, Hilden, Germany) with QG buffer, following the manufacturers’ instructions, and eluted DNA was quantified by fluorometry using the QUBIT dsDNA BR Assay Kit (ThermoFisher, MA, USA). .. For each patient, PCR products were normalized to 1.5 ng/μL, pooled in a single tube, and purified using KAPA Pure Beads (KapaBiosystems, Roche, Pleasanton, CA, USA) to ensure that no short DNA fragments were present in the library.

    Purification:

    Article Title: Naturally occurring SARS-CoV-2 gene deletions close to the spike S1/S2 cleavage site in the viral quasispecies of COVID19 patients.
    Article Snippet: The second round of amplification (nested) was done using overlapping internal primer pairs to amplify fragments 470bp to 313bp in length. .. The FastStart High-Fidelity PCR System dNTPack (Sigma, St. Louis, MO, CA) was used for this purpose, as follows: activation at 94ºC for 4 min, followed by 30 cycles with denaturation at 94ºC for 30 sec, annealing at 55ºC for 30 sec, and elongation at 72ºC for 40 sec, ending with a single elongation step at 72ºC for 7 min. PCR products were purified using the QIAquick Gel Extraction Kit (Qiagen, Hilden, Germany) with QG buffer, following the manufacturers’ instructions, and eluted DNA was quantified by fluorometry using the QUBIT dsDNA BR Assay Kit (ThermoFisher, MA, USA). .. For each patient, PCR products were normalized to 1.5 ng/µL, pooled in a single tube, and purified using KAPA Pure Beads (KapaBiosystems, Roche, Pleasanton, CA, USA) to ensure that no short DNA fragments were present in the library.

    Article Title: Naturally occurring SARS-CoV-2 gene deletions close to the spike S1/S2 cleavage site in the viral quasispecies of COVID19 patients
    Article Snippet: The second round of amplification (nested) was done using overlapping internal primer pairs to amplify fragments 470bp to 313bp in length. .. The FastStart High-Fidelity PCR System dNTPack (Sigma, St. Louis, MO, CA) was used for this purpose, as follows: activation at 94°C for 4 min, followed by 30 cycles with denaturation at 94°C for 30 sec, annealing at 55°C for 30 sec, and elongation at 72°C for 40 sec, ending with a single elongation step at 72°C for 7 min. PCR products were purified using the QIAquick Gel Extraction Kit (Qiagen, Hilden, Germany) with QG buffer, following the manufacturers’ instructions, and eluted DNA was quantified by fluorometry using the QUBIT dsDNA BR Assay Kit (ThermoFisher, MA, USA). .. For each patient, PCR products were normalized to 1.5 ng/μL, pooled in a single tube, and purified using KAPA Pure Beads (KapaBiosystems, Roche, Pleasanton, CA, USA) to ensure that no short DNA fragments were present in the library.

    Gel Extraction:

    Article Title: Naturally occurring SARS-CoV-2 gene deletions close to the spike S1/S2 cleavage site in the viral quasispecies of COVID19 patients.
    Article Snippet: The second round of amplification (nested) was done using overlapping internal primer pairs to amplify fragments 470bp to 313bp in length. .. The FastStart High-Fidelity PCR System dNTPack (Sigma, St. Louis, MO, CA) was used for this purpose, as follows: activation at 94ºC for 4 min, followed by 30 cycles with denaturation at 94ºC for 30 sec, annealing at 55ºC for 30 sec, and elongation at 72ºC for 40 sec, ending with a single elongation step at 72ºC for 7 min. PCR products were purified using the QIAquick Gel Extraction Kit (Qiagen, Hilden, Germany) with QG buffer, following the manufacturers’ instructions, and eluted DNA was quantified by fluorometry using the QUBIT dsDNA BR Assay Kit (ThermoFisher, MA, USA). .. For each patient, PCR products were normalized to 1.5 ng/µL, pooled in a single tube, and purified using KAPA Pure Beads (KapaBiosystems, Roche, Pleasanton, CA, USA) to ensure that no short DNA fragments were present in the library.

    Article Title: Naturally occurring SARS-CoV-2 gene deletions close to the spike S1/S2 cleavage site in the viral quasispecies of COVID19 patients
    Article Snippet: The second round of amplification (nested) was done using overlapping internal primer pairs to amplify fragments 470bp to 313bp in length. .. The FastStart High-Fidelity PCR System dNTPack (Sigma, St. Louis, MO, CA) was used for this purpose, as follows: activation at 94°C for 4 min, followed by 30 cycles with denaturation at 94°C for 30 sec, annealing at 55°C for 30 sec, and elongation at 72°C for 40 sec, ending with a single elongation step at 72°C for 7 min. PCR products were purified using the QIAquick Gel Extraction Kit (Qiagen, Hilden, Germany) with QG buffer, following the manufacturers’ instructions, and eluted DNA was quantified by fluorometry using the QUBIT dsDNA BR Assay Kit (ThermoFisher, MA, USA). .. For each patient, PCR products were normalized to 1.5 ng/μL, pooled in a single tube, and purified using KAPA Pure Beads (KapaBiosystems, Roche, Pleasanton, CA, USA) to ensure that no short DNA fragments were present in the library.

    Amplification:

    Article Title: DNA methylation clocks as a predictor for ageing and age estimation in naked mole-rats, Heterocephalus glaber
    Article Snippet: DNA was bisulfite converted in a 96 well plate format using the EZ-96 DNA Methylation™ Kit (Zymo, Cat. D5003). .. Target amplification was performed using the FastStart High Fidelity PCR System, dNTPack (Sigma-Aldrich, Cat. 4738284001) in the 48.48 layout on the Fluidigm® C1 system (Fluidigm®, USA), a microfluidics platform. .. Library preparation was performed using the same kit including 4 μl of Access Array Barcode Library Primer and 1 μl of PCR product diluted 1:100.

    Article Title: Whole-genome characterization and resistance-associated substitutions in a new HCV genotype 1 subtype
    Article Snippet: .. All primers were numbered according to isolate H77 (accession number AF009606) The external fragments were amplified using the Transcriptor One Step RT-PCR Kit (#04655885001 Sigma, Roche), and for the internals, the FastStart High Fidelity PCR System, dNTPack (#04738292001 Sigma, Roche), adapting the various PCR programs to the lengths and temperatures of each specific primer pair. .. Due to the high variability of the region corresponding to internal fragment number 11, a random priming (#11034731001 Sigma, Roche) PCR was necessary to perform between the RT-PCR and the nested PCR with the specific primers, to obtain enough DNA for analysis.

    Reverse Transcription Polymerase Chain Reaction:

    Article Title: Whole-genome characterization and resistance-associated substitutions in a new HCV genotype 1 subtype
    Article Snippet: .. All primers were numbered according to isolate H77 (accession number AF009606) The external fragments were amplified using the Transcriptor One Step RT-PCR Kit (#04655885001 Sigma, Roche), and for the internals, the FastStart High Fidelity PCR System, dNTPack (#04738292001 Sigma, Roche), adapting the various PCR programs to the lengths and temperatures of each specific primer pair. .. Due to the high variability of the region corresponding to internal fragment number 11, a random priming (#11034731001 Sigma, Roche) PCR was necessary to perform between the RT-PCR and the nested PCR with the specific primers, to obtain enough DNA for analysis.

    Similar Products

  • Logo
  • About
  • News
  • Press Release
  • Team
  • Advisors
  • Partners
  • Contact
  • Bioz Stars
  • Bioz vStars
  • 94
    Millipore taq dna polymerase
    Strand-displacement by mutant enzymes with D732N. (A) Half-dumbbell test. Catalyzed <t>DNA</t> synthesis begins at the primer (arrow) and proceeds to and through the loop and back again, only if the polymerase is capable of strand displacement, to make a product 2.4 kb in size. The two D732N mutant versions of <t>Taq</t> (full-length A111 and Klentaq1) are marked with asterisk (*). Bst pol was expected to be able to strand-displace, and was included as a positive control. We explain unextended 1.2 kb template as due to inefficiency of the half-dumbbell construction at unknown point(s) during its preparation. (B) Discovery of LAMP ability. A multiply mutant form of Taq pol carrying D732N, named KTflnC4RR (lanes KT*, see text) was tested compared Manta (lanes M), a brand of the large fragment of Bst DNA polymerase (Enzymatics). The banded amplification product which is typical for LAMP appeared at 3 h incubation at 60°C for both enzymes but only for KT* enzyme at 70°C. The middle lane size standards were 1, 2, 4 kb.
    Taq Dna Polymerase, supplied by Millipore, used in various techniques. Bioz Stars score: 94/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/taq dna polymerase/product/Millipore
    Average 94 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    taq dna polymerase - by Bioz Stars, 2021-07
    94/100 stars
      Buy from Supplier

    93
    Millipore faststart high fidelity pcr system dntpack
    Strand-displacement by mutant enzymes with D732N. (A) Half-dumbbell test. Catalyzed <t>DNA</t> synthesis begins at the primer (arrow) and proceeds to and through the loop and back again, only if the polymerase is capable of strand displacement, to make a product 2.4 kb in size. The two D732N mutant versions of <t>Taq</t> (full-length A111 and Klentaq1) are marked with asterisk (*). Bst pol was expected to be able to strand-displace, and was included as a positive control. We explain unextended 1.2 kb template as due to inefficiency of the half-dumbbell construction at unknown point(s) during its preparation. (B) Discovery of LAMP ability. A multiply mutant form of Taq pol carrying D732N, named KTflnC4RR (lanes KT*, see text) was tested compared Manta (lanes M), a brand of the large fragment of Bst DNA polymerase (Enzymatics). The banded amplification product which is typical for LAMP appeared at 3 h incubation at 60°C for both enzymes but only for KT* enzyme at 70°C. The middle lane size standards were 1, 2, 4 kb.
    Faststart High Fidelity Pcr System Dntpack, supplied by Millipore, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/faststart high fidelity pcr system dntpack/product/Millipore
    Average 93 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    faststart high fidelity pcr system dntpack - by Bioz Stars, 2021-07
    93/100 stars
      Buy from Supplier

    93
    Millipore expand long range dntpack
    Strand-displacement by mutant enzymes with D732N. (A) Half-dumbbell test. Catalyzed <t>DNA</t> synthesis begins at the primer (arrow) and proceeds to and through the loop and back again, only if the polymerase is capable of strand displacement, to make a product 2.4 kb in size. The two D732N mutant versions of <t>Taq</t> (full-length A111 and Klentaq1) are marked with asterisk (*). Bst pol was expected to be able to strand-displace, and was included as a positive control. We explain unextended 1.2 kb template as due to inefficiency of the half-dumbbell construction at unknown point(s) during its preparation. (B) Discovery of LAMP ability. A multiply mutant form of Taq pol carrying D732N, named KTflnC4RR (lanes KT*, see text) was tested compared Manta (lanes M), a brand of the large fragment of Bst DNA polymerase (Enzymatics). The banded amplification product which is typical for LAMP appeared at 3 h incubation at 60°C for both enzymes but only for KT* enzyme at 70°C. The middle lane size standards were 1, 2, 4 kb.
    Expand Long Range Dntpack, supplied by Millipore, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/expand long range dntpack/product/Millipore
    Average 93 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    expand long range dntpack - by Bioz Stars, 2021-07
    93/100 stars
      Buy from Supplier

    Image Search Results


    Strand-displacement by mutant enzymes with D732N. (A) Half-dumbbell test. Catalyzed DNA synthesis begins at the primer (arrow) and proceeds to and through the loop and back again, only if the polymerase is capable of strand displacement, to make a product 2.4 kb in size. The two D732N mutant versions of Taq (full-length A111 and Klentaq1) are marked with asterisk (*). Bst pol was expected to be able to strand-displace, and was included as a positive control. We explain unextended 1.2 kb template as due to inefficiency of the half-dumbbell construction at unknown point(s) during its preparation. (B) Discovery of LAMP ability. A multiply mutant form of Taq pol carrying D732N, named KTflnC4RR (lanes KT*, see text) was tested compared Manta (lanes M), a brand of the large fragment of Bst DNA polymerase (Enzymatics). The banded amplification product which is typical for LAMP appeared at 3 h incubation at 60°C for both enzymes but only for KT* enzyme at 70°C. The middle lane size standards were 1, 2, 4 kb.

    Journal: Frontiers in Bioengineering and Biotechnology

    Article Title: A Single Amino Acid Change to Taq DNA Polymerase Enables Faster PCR, Reverse Transcription and Strand-Displacement

    doi: 10.3389/fbioe.2020.553474

    Figure Lengend Snippet: Strand-displacement by mutant enzymes with D732N. (A) Half-dumbbell test. Catalyzed DNA synthesis begins at the primer (arrow) and proceeds to and through the loop and back again, only if the polymerase is capable of strand displacement, to make a product 2.4 kb in size. The two D732N mutant versions of Taq (full-length A111 and Klentaq1) are marked with asterisk (*). Bst pol was expected to be able to strand-displace, and was included as a positive control. We explain unextended 1.2 kb template as due to inefficiency of the half-dumbbell construction at unknown point(s) during its preparation. (B) Discovery of LAMP ability. A multiply mutant form of Taq pol carrying D732N, named KTflnC4RR (lanes KT*, see text) was tested compared Manta (lanes M), a brand of the large fragment of Bst DNA polymerase (Enzymatics). The banded amplification product which is typical for LAMP appeared at 3 h incubation at 60°C for both enzymes but only for KT* enzyme at 70°C. The middle lane size standards were 1, 2, 4 kb.

    Article Snippet: We have found that deletions and mutations of Taq DNA polymerase have various levels of Kornberg units per ug.

    Techniques: Mutagenesis, DNA Synthesis, Positive Control, Amplification, Incubation

    RT-LAMP of MS2 RNA using single-enzyme Taq-D732N or Klentaq1-D732N. Shown are real-time fluorescence traces of reactions with 1/3 μl test enzyme but no RT (reverse transcriptase) enzyme and no Mn ++ , using Eva Green fluorescence as indicator of double-strand DNA amplification. Blue circles/line: complete 26 μl reaction, with 3 ng MS2 RNA. Black lines: MS2 RNA preincubated with RNase I. Red lines, no MS2 RNA added. Taq and KT1 : Controls wild-type Taq (full-length) and Klentaq1 (N-terminal deletion of 278 a.a.) DNA polymerase. Buffer conditions are 50 mM Tris–HCl pH 8.55, 8 mM ammonium sulfate, 200 μM each dNTP, 0.75 M betaine, 0.025% Brij-58, 68°C. Melt curves for this experiment are shown in Supplementary Figure 2 .

    Journal: Frontiers in Bioengineering and Biotechnology

    Article Title: A Single Amino Acid Change to Taq DNA Polymerase Enables Faster PCR, Reverse Transcription and Strand-Displacement

    doi: 10.3389/fbioe.2020.553474

    Figure Lengend Snippet: RT-LAMP of MS2 RNA using single-enzyme Taq-D732N or Klentaq1-D732N. Shown are real-time fluorescence traces of reactions with 1/3 μl test enzyme but no RT (reverse transcriptase) enzyme and no Mn ++ , using Eva Green fluorescence as indicator of double-strand DNA amplification. Blue circles/line: complete 26 μl reaction, with 3 ng MS2 RNA. Black lines: MS2 RNA preincubated with RNase I. Red lines, no MS2 RNA added. Taq and KT1 : Controls wild-type Taq (full-length) and Klentaq1 (N-terminal deletion of 278 a.a.) DNA polymerase. Buffer conditions are 50 mM Tris–HCl pH 8.55, 8 mM ammonium sulfate, 200 μM each dNTP, 0.75 M betaine, 0.025% Brij-58, 68°C. Melt curves for this experiment are shown in Supplementary Figure 2 .

    Article Snippet: We have found that deletions and mutations of Taq DNA polymerase have various levels of Kornberg units per ug.

    Techniques: Fluorescence, Amplification

    RT-PCR catalyzable by single enzyme Taq D732N but not wild-type Taq. No reverse transcriptase was included, nor was any manganese ion. Two MS2 phage RNA targets, P and K were amplified from 5 pg phage RNA with 0.1 or 0.05 μl D732N mutant or twice as much volume of wild-type Taq (NEB) in 35 μl reactions for 35 PCR cycles. The amplified products were run in a 2.5% agarose gel stained with ethidium bromide, along with a 100 bp DNA ladder (white/black reversed for clarity as printed). As negative control, no RNA was included for the reactions analyzed on the bottom half of the gel. The size of the clearly and successfully amplified bands are 405 and 544 bp (primer-primer gap sizes 359 and 500).

    Journal: Frontiers in Bioengineering and Biotechnology

    Article Title: A Single Amino Acid Change to Taq DNA Polymerase Enables Faster PCR, Reverse Transcription and Strand-Displacement

    doi: 10.3389/fbioe.2020.553474

    Figure Lengend Snippet: RT-PCR catalyzable by single enzyme Taq D732N but not wild-type Taq. No reverse transcriptase was included, nor was any manganese ion. Two MS2 phage RNA targets, P and K were amplified from 5 pg phage RNA with 0.1 or 0.05 μl D732N mutant or twice as much volume of wild-type Taq (NEB) in 35 μl reactions for 35 PCR cycles. The amplified products were run in a 2.5% agarose gel stained with ethidium bromide, along with a 100 bp DNA ladder (white/black reversed for clarity as printed). As negative control, no RNA was included for the reactions analyzed on the bottom half of the gel. The size of the clearly and successfully amplified bands are 405 and 544 bp (primer-primer gap sizes 359 and 500).

    Article Snippet: We have found that deletions and mutations of Taq DNA polymerase have various levels of Kornberg units per ug.

    Techniques: Reverse Transcription Polymerase Chain Reaction, Amplification, Mutagenesis, Polymerase Chain Reaction, Agarose Gel Electrophoresis, Staining, Negative Control