taq polymerase  (TaKaRa)

 
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    Name:
    TaKaRa Taq DNA Polymerase
    Description:
    TaKaRa Taq DNA Polymerase is a recombinant version Taq polymerase derived from the Thermus aquaticus YT 1 strain and is suitable for routine PCR applications
    Catalog Number:
    r001c
    Price:
    None
    Size:
    3 000 Units
    Category:
    Takara Taq and premix Takara Taq products Standard PCR PCR
    		Buy from Supplier

    Structured Review

    TaKaRa taq polymerase
    PCR amplicons digested by restriction endonucleases <t>Taq</t> I. Lanes 1–53: random samples from 56 samples; P: PCR amplicon; M: DL-2000 <t>DNA</t> marker.
    TaKaRa Taq DNA Polymerase is a recombinant version Taq polymerase derived from the Thermus aquaticus YT 1 strain and is suitable for routine PCR applications
    https://www.bioz.com/result/taq polymerase/product/TaKaRa
    Average 99 stars, based on 1618 article reviews
    Price from $9.99 to $1999.99
    taq polymerase - by Bioz Stars, 2020-07
    99/100 stars

    Related Products / Commonly Used Together

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    dntp
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    pcr buffer
    cdna
    kcl
    template dna
    pcr reactions
    tris-hcl
    genomic dna
    pcr mixture
    pcr amplification
    deoxynucleoside triphosphate
    dntp mix

    Images

    1) Product Images from "Molecular Identification of Ancylostoma caninum Isolated from Cats in Southern China Based on Complete ITS Sequence"

    Article Title: Molecular Identification of Ancylostoma caninum Isolated from Cats in Southern China Based on Complete ITS Sequence

    Journal: BioMed Research International

    doi: 10.1155/2013/868050

    PCR amplicons digested by restriction endonucleases Taq I. Lanes 1–53: random samples from 56 samples; P: PCR amplicon; M: DL-2000 DNA marker.
    Figure Legend Snippet: PCR amplicons digested by restriction endonucleases Taq I. Lanes 1–53: random samples from 56 samples; P: PCR amplicon; M: DL-2000 DNA marker.

    Techniques Used: Polymerase Chain Reaction, Amplification, Marker

    2) Product Images from "Identification of a Gene Essential for Sheathed Structure Formation in Sphaerotilus natans, a Filamentous Sheathed Bacterium"

    Article Title: Identification of a Gene Essential for Sheathed Structure Formation in Sphaerotilus natans, a Filamentous Sheathed Bacterium

    Journal: Applied and Environmental Microbiology

    doi: 10.1128/AEM.68.1.365-371.2002

    Schematic representation of a method for monitoring sthA expression in vivo. (A) Construction of pSTH16, which was used to insert gfp and the Km r cassette downstream of sthA by double-crossover integration. A 2.5-kb fragment containing sthA (open box) and its upstream region (shaded box) was amplified from genomic DNA by using PyroBEST polymerase and primers UP1 (5′-ATGAGCGATACCCACTTCGGCTTC-3′) and UP2 (5′-TGTGGCGTGCACGACATGTTCGAT-3′). Adenine was added to the 3′ termini of the PCR products by using Taq polymerase, and the products were cloned into pT7Blue by the TA cloning procedure. A plasmid in which the orientation of the insert was opposite that of lacZ was then selected and designated pSTH11. A 0.8-kb Xba I- Hin dIII fragment, including promoterless gfp , was prepared from pUTminiTn 5 gfp and ligated into pSTH11 which had been digested with Xba I and Hin dIII to generate pSTH12. A 1.5-kb fragment that included the region downstream of the sthA gene (ORF and shaded boxes) was amplified from genomic DNA by using PyroBEST polymerase with primers DP3 (5′-TGAAGGCCTGCGGGACGACAGAAGC-3′) and DP4 (5′-TGCTCGTCGATGAAGCGGCTGGCCT-3′), and the PCR product was cloned into pT7Blue as described above. A plasmid in which the orientation of the insert was opposite that of lacZ was selected and designated pSTH13. A 1.3-kb Hin dIII- Sma I fragment carrying Km r prepared from pSUP5011 was blunted and ligated into pSTH13 which had been digested with Bam HI and blunted in order to generate pSTH14. pSTH12 was digested with Eco RI and Hin dIII, and the 3.3-kb Eco RI- Hin dIII fragment containing sthA-gfp of pSTH12 was blunted and ligated into the Sma I site of pSTH14. A plasmid in which the insert was oriented properly was selected and designated pSTH15. Finally, a 1.8-kb Bam HI fragment carrying the mob site prepared from pSUP5011 was inserted into the Bam HI site of pSTH15, yielding pSTH16, in which gfp and the Km r cassette were situated between sthA and its downstream ORF, without disrupting either gene. pol., polymerase. (B) Strategy for insertion of gfp and the Km r cassette between sthA and its downstream ORF.
    Figure Legend Snippet: Schematic representation of a method for monitoring sthA expression in vivo. (A) Construction of pSTH16, which was used to insert gfp and the Km r cassette downstream of sthA by double-crossover integration. A 2.5-kb fragment containing sthA (open box) and its upstream region (shaded box) was amplified from genomic DNA by using PyroBEST polymerase and primers UP1 (5′-ATGAGCGATACCCACTTCGGCTTC-3′) and UP2 (5′-TGTGGCGTGCACGACATGTTCGAT-3′). Adenine was added to the 3′ termini of the PCR products by using Taq polymerase, and the products were cloned into pT7Blue by the TA cloning procedure. A plasmid in which the orientation of the insert was opposite that of lacZ was then selected and designated pSTH11. A 0.8-kb Xba I- Hin dIII fragment, including promoterless gfp , was prepared from pUTminiTn 5 gfp and ligated into pSTH11 which had been digested with Xba I and Hin dIII to generate pSTH12. A 1.5-kb fragment that included the region downstream of the sthA gene (ORF and shaded boxes) was amplified from genomic DNA by using PyroBEST polymerase with primers DP3 (5′-TGAAGGCCTGCGGGACGACAGAAGC-3′) and DP4 (5′-TGCTCGTCGATGAAGCGGCTGGCCT-3′), and the PCR product was cloned into pT7Blue as described above. A plasmid in which the orientation of the insert was opposite that of lacZ was selected and designated pSTH13. A 1.3-kb Hin dIII- Sma I fragment carrying Km r prepared from pSUP5011 was blunted and ligated into pSTH13 which had been digested with Bam HI and blunted in order to generate pSTH14. pSTH12 was digested with Eco RI and Hin dIII, and the 3.3-kb Eco RI- Hin dIII fragment containing sthA-gfp of pSTH12 was blunted and ligated into the Sma I site of pSTH14. A plasmid in which the insert was oriented properly was selected and designated pSTH15. Finally, a 1.8-kb Bam HI fragment carrying the mob site prepared from pSUP5011 was inserted into the Bam HI site of pSTH15, yielding pSTH16, in which gfp and the Km r cassette were situated between sthA and its downstream ORF, without disrupting either gene. pol., polymerase. (B) Strategy for insertion of gfp and the Km r cassette between sthA and its downstream ORF.

    Techniques Used: Expressing, In Vivo, Amplification, Polymerase Chain Reaction, Clone Assay, TA Cloning, Plasmid Preparation

    3) Product Images from "A Novel Universal Primer-Multiplex-PCR Method with Sequencing Gel Electrophoresis Analysis"

    Article Title: A Novel Universal Primer-Multiplex-PCR Method with Sequencing Gel Electrophoresis Analysis

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0022900

    Optimization of the UP-M-PCR. Lane A, B, C, D, E, amplicon fragments by UP (500 nmol L −1 ) and compound specific primer hpt-839, nptII-508, pat-262, bar-226 and sps-110 at a series concentrations of 500 nmol L −1 , 50 nmol L −1 , 25 nmol L −1 , 5 nmol L −1 , 0.5 nmol L −1 ; lane F1, amplicon fragments by UP at 500 nmol L −1 and all compound specific primers at 25 nmol L −1 ; lane F2, amplicon fragments by UP at 500 nmol L −1 and all compound specific primers at the optimized concentration; lane G1,G2,G3, amplicon fragments by all primers at the optimized concentration with TaKaRa Taq ™, Phire™ Hot Start DNA polymerase, iProof™ High-Fidelity DNA polymerase; lane H1, amplicon fragments by all primers at the optimized concentration with Phire™ Hot Start DNA polymerase under the common amplification conditions; lane H2, amplicon fragments by all primers at the optimized concentration with Phire™ Hot Start DNA polymerase under the optimized amplification conditions; lane M, 100 bp DNA Marker.
    Figure Legend Snippet: Optimization of the UP-M-PCR. Lane A, B, C, D, E, amplicon fragments by UP (500 nmol L −1 ) and compound specific primer hpt-839, nptII-508, pat-262, bar-226 and sps-110 at a series concentrations of 500 nmol L −1 , 50 nmol L −1 , 25 nmol L −1 , 5 nmol L −1 , 0.5 nmol L −1 ; lane F1, amplicon fragments by UP at 500 nmol L −1 and all compound specific primers at 25 nmol L −1 ; lane F2, amplicon fragments by UP at 500 nmol L −1 and all compound specific primers at the optimized concentration; lane G1,G2,G3, amplicon fragments by all primers at the optimized concentration with TaKaRa Taq ™, Phire™ Hot Start DNA polymerase, iProof™ High-Fidelity DNA polymerase; lane H1, amplicon fragments by all primers at the optimized concentration with Phire™ Hot Start DNA polymerase under the common amplification conditions; lane H2, amplicon fragments by all primers at the optimized concentration with Phire™ Hot Start DNA polymerase under the optimized amplification conditions; lane M, 100 bp DNA Marker.

    Techniques Used: Polymerase Chain Reaction, Amplification, Concentration Assay, Marker

    4) Product Images from "A Novel Universal Primer-Multiplex-PCR Method with Sequencing Gel Electrophoresis Analysis"

    Article Title: A Novel Universal Primer-Multiplex-PCR Method with Sequencing Gel Electrophoresis Analysis

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0022900

    Optimization of the UP-M-PCR. Lane A, B, C, D, E, amplicon fragments by UP (500 nmol L −1 ) and compound specific primer hpt-839, nptII-508, pat-262, bar-226 and sps-110 at a series concentrations of 500 nmol L −1 , 50 nmol L −1 , 25 nmol L −1 , 5 nmol L −1 , 0.5 nmol L −1 ; lane F1, amplicon fragments by UP at 500 nmol L −1 and all compound specific primers at 25 nmol L −1 ; lane F2, amplicon fragments by UP at 500 nmol L −1 and all compound specific primers at the optimized concentration; lane G1,G2,G3, amplicon fragments by all primers at the optimized concentration with TaKaRa Taq ™, Phire™ Hot Start DNA polymerase, iProof™ High-Fidelity DNA polymerase; lane H1, amplicon fragments by all primers at the optimized concentration with Phire™ Hot Start DNA polymerase under the common amplification conditions; lane H2, amplicon fragments by all primers at the optimized concentration with Phire™ Hot Start DNA polymerase under the optimized amplification conditions; lane M, 100 bp DNA Marker.
    Figure Legend Snippet: Optimization of the UP-M-PCR. Lane A, B, C, D, E, amplicon fragments by UP (500 nmol L −1 ) and compound specific primer hpt-839, nptII-508, pat-262, bar-226 and sps-110 at a series concentrations of 500 nmol L −1 , 50 nmol L −1 , 25 nmol L −1 , 5 nmol L −1 , 0.5 nmol L −1 ; lane F1, amplicon fragments by UP at 500 nmol L −1 and all compound specific primers at 25 nmol L −1 ; lane F2, amplicon fragments by UP at 500 nmol L −1 and all compound specific primers at the optimized concentration; lane G1,G2,G3, amplicon fragments by all primers at the optimized concentration with TaKaRa Taq ™, Phire™ Hot Start DNA polymerase, iProof™ High-Fidelity DNA polymerase; lane H1, amplicon fragments by all primers at the optimized concentration with Phire™ Hot Start DNA polymerase under the common amplification conditions; lane H2, amplicon fragments by all primers at the optimized concentration with Phire™ Hot Start DNA polymerase under the optimized amplification conditions; lane M, 100 bp DNA Marker.

    Techniques Used: Polymerase Chain Reaction, Amplification, Concentration Assay, Marker

    Related Articles

    Amplification:

    Article Title: Modified Proofreading PCR for Detection of Point Mutations, Insertions and Deletions Using a ddNTP-Blocked Primer
    Article Snippet: .. The results showed that the use of 0.1–0.15 U of high-fidelity DNA polymerase combined with 1 U of Taq DNA polymerase in per 20 μL reaction resulted in the best discrimination and amplification effects ( ). .. Thus, a mix of 0.1–0.15 U of high-fidelity DNA polymerase with 1 U of Taq DNA polymerase in per 20 μL reaction was selected for the subsequent experiments.

    Mutagenesis:

    Article Title: Modified Proofreading PCR for Detection of Point Mutations, Insertions and Deletions Using a ddNTP-Blocked Primer
    Article Snippet: .. Development of a modified PR-PCR method using the dideoxynucleotide-blocked primer and a mixture of Taq DNA polymerase and high-fidelity DNA polymerase The experimental results above indicate that two major factors affect the efficiency of the PR-PCR in mutation detection. .. One is that the modification of the 3'-end of a primer with-NH2 or-Pi is unable to completely block primer extension ( ).

    Isolation:

    Article Title: Involvement of GluR2 and GluR3 subunit C-termini in the trigeminal spinal subnucleus caudalis and C1-C2 neurons in trigeminal neuropathic pain
    Article Snippet: .. PCR was performed on the isolated genomic DNA using taq DNA polymerase (TaKaRa Ex Taq™, Takara, Otsu, Japan) and primer sets (primer #21–23 for GluR2 delta7: #21, 5′-ACA GAG GAA GGT AGT GGA AGG GAG-3′; #22, 5′-CTT GGT TTG GTT GTT GGT CAT AGC-3′; #23, 5′-CTA GTG AAC CTC TTC GAG GGA C-3′; primer #31–33 for GluR3 delta7: #31, 5′-CCA ATA CTC CAC AGG GGC AAT TTA TC-3′; #32, 5′-CCG TTG ACT GTT TTG AAT CTC ACA CC-3′; #33, 5′-CTA GTG AAC CTC TTC GAG GGA C-3′). .. Size of the amplification product was estimated by gel electrophoresis for genotyping (210 bp for wild-type and 350 bp for GluR2 delta7; 440 bp for wild-type and 300 bp for GluR3 delta7).

    Polymerase Chain Reaction:

    Article Title: Modified Proofreading PCR for Detection of Point Mutations, Insertions and Deletions Using a ddNTP-Blocked Primer
    Article Snippet: .. All reactions were performed in a total volume of 20 μL containing a mixture of 0.5 U of Taq DNA polymerase, 0.3 μM each of forward and reverse primer, 0.2 mM dNTPs, (1×) PCR Buffer and 5 ng (equal to approximately 1×106 copies) of template. .. The PCR cycling condition was pre-denaturation at 98°C for 2 min, followed by 35 cycles of denaturation at 98°C for 10 s, annealing at 55°C for 15 s and extension at 72°C for 15 s. The positive (TB-526PC and TB-531PC) and negative control plasmids were both amplified, regardless of whether WT-F or Mu-F was used.

    Article Title: Involvement of GluR2 and GluR3 subunit C-termini in the trigeminal spinal subnucleus caudalis and C1-C2 neurons in trigeminal neuropathic pain
    Article Snippet: .. PCR was performed on the isolated genomic DNA using taq DNA polymerase (TaKaRa Ex Taq™, Takara, Otsu, Japan) and primer sets (primer #21–23 for GluR2 delta7: #21, 5′-ACA GAG GAA GGT AGT GGA AGG GAG-3′; #22, 5′-CTT GGT TTG GTT GTT GGT CAT AGC-3′; #23, 5′-CTA GTG AAC CTC TTC GAG GGA C-3′; primer #31–33 for GluR3 delta7: #31, 5′-CCA ATA CTC CAC AGG GGC AAT TTA TC-3′; #32, 5′-CCG TTG ACT GTT TTG AAT CTC ACA CC-3′; #33, 5′-CTA GTG AAC CTC TTC GAG GGA C-3′). .. Size of the amplification product was estimated by gel electrophoresis for genotyping (210 bp for wild-type and 350 bp for GluR2 delta7; 440 bp for wild-type and 300 bp for GluR3 delta7).

    Article Title: Molecular Identification of Ancylostoma caninum Isolated from Cats in Southern China Based on Complete ITS Sequence
    Article Snippet: .. PCR-RFLP Both PCRs were performed in 25 μ L volume containing 2 μ L of the DNA sample, 0.2 μ L of Taq polymerase (TaKaRa, Dalian, China), 2.5 μ L of 10×Taq buffer (TaKaRa), 2 μ L of diethylnitrophenyl thiophosphate (dNTP, TaKaRa) mixture, 0.5 μ L of each primer (AF/AR or CAF/CAR, 50 mM), and 17.3 μ L of distilled water. .. PCR cycling parameters were as follows: 1 cycle at 96°C for 5 minutes, then 35 cycles of 96°C for 30 seconds, at 60°C for 30 seconds, and at 72°C for 90 seconds, followed by 1 cycle at 72°C for 7 minutes.

    Article Title: Characterization of porcine cytokine inducible SH2-containing protein gene and its association with piglet diarrhea traits
    Article Snippet: .. The polymerase chain reaction (PCR) reaction was 25 μL containing 100 ng templates, 0.5 μM of each primer, 0.5 mM dNTPs, 2.0 U Taq DNA polymerase and 2.5 μL buffer (TaKaRa, Tokyo, Japan). ..

    Modification:

    Article Title: Modified Proofreading PCR for Detection of Point Mutations, Insertions and Deletions Using a ddNTP-Blocked Primer
    Article Snippet: .. Development of a modified PR-PCR method using the dideoxynucleotide-blocked primer and a mixture of Taq DNA polymerase and high-fidelity DNA polymerase The experimental results above indicate that two major factors affect the efficiency of the PR-PCR in mutation detection. .. One is that the modification of the 3'-end of a primer with-NH2 or-Pi is unable to completely block primer extension ( ).

    Article Title: Modified Proofreading PCR for Detection of Point Mutations, Insertions and Deletions Using a ddNTP-Blocked Primer
    Article Snippet: .. In conclusion, we developed a novel modified PR-PCR method that uses a ddNTP-blocked primer and an enzyme combination of a routine amount of Taq DNA polymerase and a tiny amount of high-fidelity DNA polymerase. .. This method can be used for easy detection of various mutation types, including point mutations, insertions and deletions, with high selectivity and sensitivity.

    Chloramphenicol Acetyltransferase Assay:

    Article Title: Involvement of GluR2 and GluR3 subunit C-termini in the trigeminal spinal subnucleus caudalis and C1-C2 neurons in trigeminal neuropathic pain
    Article Snippet: .. PCR was performed on the isolated genomic DNA using taq DNA polymerase (TaKaRa Ex Taq™, Takara, Otsu, Japan) and primer sets (primer #21–23 for GluR2 delta7: #21, 5′-ACA GAG GAA GGT AGT GGA AGG GAG-3′; #22, 5′-CTT GGT TTG GTT GTT GGT CAT AGC-3′; #23, 5′-CTA GTG AAC CTC TTC GAG GGA C-3′; primer #31–33 for GluR3 delta7: #31, 5′-CCA ATA CTC CAC AGG GGC AAT TTA TC-3′; #32, 5′-CCG TTG ACT GTT TTG AAT CTC ACA CC-3′; #33, 5′-CTA GTG AAC CTC TTC GAG GGA C-3′). .. Size of the amplification product was estimated by gel electrophoresis for genotyping (210 bp for wild-type and 350 bp for GluR2 delta7; 440 bp for wild-type and 300 bp for GluR3 delta7).

    Activated Clotting Time Assay:

    Article Title: Involvement of GluR2 and GluR3 subunit C-termini in the trigeminal spinal subnucleus caudalis and C1-C2 neurons in trigeminal neuropathic pain
    Article Snippet: .. PCR was performed on the isolated genomic DNA using taq DNA polymerase (TaKaRa Ex Taq™, Takara, Otsu, Japan) and primer sets (primer #21–23 for GluR2 delta7: #21, 5′-ACA GAG GAA GGT AGT GGA AGG GAG-3′; #22, 5′-CTT GGT TTG GTT GTT GGT CAT AGC-3′; #23, 5′-CTA GTG AAC CTC TTC GAG GGA C-3′; primer #31–33 for GluR3 delta7: #31, 5′-CCA ATA CTC CAC AGG GGC AAT TTA TC-3′; #32, 5′-CCG TTG ACT GTT TTG AAT CTC ACA CC-3′; #33, 5′-CTA GTG AAC CTC TTC GAG GGA C-3′). .. Size of the amplification product was estimated by gel electrophoresis for genotyping (210 bp for wild-type and 350 bp for GluR2 delta7; 440 bp for wild-type and 300 bp for GluR3 delta7).

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    • taq dna polymerase  (TaKaRa)
    99
    TaKaRa taq dna polymerase
    (A) Genotype in each generation of mutant mice confirmed by PCR analysis on isolated genomic <t>DNA.</t> PCR was performed on the isolated genomic DNA using <t>taq</t> DNA polymerase and primer sets (#1 and #2 are for wild-type and GluR2 delta7 KI, #3 and #4 are for
    Taq Dna Polymerase, supplied by TaKaRa, used in various techniques. Bioz Stars score: 99/100, based on 2134 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/taq dna polymerase/product/TaKaRa
    Average 99 stars, based on 2134 article reviews
    Price from $9.99 to $1999.99
    taq dna polymerase - by Bioz Stars, 2020-07
    99/100 stars
    		  Buy from Supplier
    la dna polymerase  (TaKaRa)
    99
    TaKaRa la dna polymerase
    ( A ) Gel electrophoresis of near full-length genome PCR products produced from four long amplifying <t>DNA</t> polymerases; KlenTaq LA (discontinued), AccuTaq LA, PrimeSTAR GXL and <t>Takara</t> LA Taq (separate gel) per manufacturer’s instructions for samples with high GC/secondary structures. M, HyperLadder 1 kb; 1, CVB3 Nancy; 2, CVB5 Faulkner; 3, H 2 O control. ( B ) Gel electrophoresis of near full-length genome PCR products produced from Takara LA Taq DNA polymerase. M, HyperLadder 1 kb; 1–4, known EV positives from NSW Health Pathology East virology diagnostic lab; 5, CVB3 Nancy; 6, H 2 .
    La Dna Polymerase, supplied by TaKaRa, used in various techniques. Bioz Stars score: 99/100, based on 667 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/la dna polymerase/product/TaKaRa
    Average 99 stars, based on 667 article reviews
    Price from $9.99 to $1999.99
    la dna polymerase - by Bioz Stars, 2020-07
    99/100 stars
    		  Buy from Supplier

    Image Search Results


    (A) Genotype in each generation of mutant mice confirmed by PCR analysis on isolated genomic DNA. PCR was performed on the isolated genomic DNA using taq DNA polymerase and primer sets (#1 and #2 are for wild-type and GluR2 delta7 KI, #3 and #4 are for

    Journal: Neuroscience letters

    Article Title: Involvement of GluR2 and GluR3 subunit C-termini in the trigeminal spinal subnucleus caudalis and C1-C2 neurons in trigeminal neuropathic pain

    doi: 10.1016/j.neulet.2010.12.060

    Figure Lengend Snippet: (A) Genotype in each generation of mutant mice confirmed by PCR analysis on isolated genomic DNA. PCR was performed on the isolated genomic DNA using taq DNA polymerase and primer sets (#1 and #2 are for wild-type and GluR2 delta7 KI, #3 and #4 are for

    Article Snippet: PCR was performed on the isolated genomic DNA using taq DNA polymerase (TaKaRa Ex Taq™, Takara, Otsu, Japan) and primer sets (primer #21–23 for GluR2 delta7: #21, 5′-ACA GAG GAA GGT AGT GGA AGG GAG-3′; #22, 5′-CTT GGT TTG GTT GTT GGT CAT AGC-3′; #23, 5′-CTA GTG AAC CTC TTC GAG GGA C-3′; primer #31–33 for GluR3 delta7: #31, 5′-CCA ATA CTC CAC AGG GGC AAT TTA TC-3′; #32, 5′-CCG TTG ACT GTT TTG AAT CTC ACA CC-3′; #33, 5′-CTA GTG AAC CTC TTC GAG GGA C-3′).

    Techniques: Mutagenesis, Mouse Assay, Polymerase Chain Reaction, Isolation

    Blocking efficiency of three kinds of blocked primers in PCR amplifications mediated by Taq DNA polymerase and high-fidelity DNA polymerase. (a) Primer and template sets. WT and Mu indicate wild-type and mutant types, respectively. The 3'-blocked forward primer completely matches the WT sequence but forms a mismatch with the Mu sequence (Mu*1) at the 3'-end. The reverse primer matches both the WT and Mu sequences. The 3'-terminal nucleotide of the blocked primer is blocked with-Pi or-Amino C6, or replaced with ddCTP. (b) Blocking efficiency of-Pi,-Amino C6 or-ddC in PCR mediated by Taq DNA polymerase. (c) Blocking efficiency of-Pi,-Amino C6 or-ddC in PCR mediated by high-fidelity DNA polymerase (KOD FX). (d) Discrimination efficiency of the typical proofreading-PCR medicated by various amounts (0.5, 1, 1.5 and 2 U) of KOD FX DNA polymerase using the ddC-blocked primer. All PCR amplifications were performed in a total volume of 20 μL containing 1 U of Taq or KOD FX DNA polymerase (Panel b and c) or various amounts of KOD FX DNA polymerase (Panel d). The cycling conditions consisted of pre-denaturation at 94°C for 2 min, followed by 30 cycles of denaturation at 94°C for 20 s, annealing at 56°C for 20 s and extension at 72°C for 25 s. NTC: no-template control.

    Journal: PLoS ONE

    Article Title: Modified Proofreading PCR for Detection of Point Mutations, Insertions and Deletions Using a ddNTP-Blocked Primer

    doi: 10.1371/journal.pone.0123468

    Figure Lengend Snippet: Blocking efficiency of three kinds of blocked primers in PCR amplifications mediated by Taq DNA polymerase and high-fidelity DNA polymerase. (a) Primer and template sets. WT and Mu indicate wild-type and mutant types, respectively. The 3'-blocked forward primer completely matches the WT sequence but forms a mismatch with the Mu sequence (Mu*1) at the 3'-end. The reverse primer matches both the WT and Mu sequences. The 3'-terminal nucleotide of the blocked primer is blocked with-Pi or-Amino C6, or replaced with ddCTP. (b) Blocking efficiency of-Pi,-Amino C6 or-ddC in PCR mediated by Taq DNA polymerase. (c) Blocking efficiency of-Pi,-Amino C6 or-ddC in PCR mediated by high-fidelity DNA polymerase (KOD FX). (d) Discrimination efficiency of the typical proofreading-PCR medicated by various amounts (0.5, 1, 1.5 and 2 U) of KOD FX DNA polymerase using the ddC-blocked primer. All PCR amplifications were performed in a total volume of 20 μL containing 1 U of Taq or KOD FX DNA polymerase (Panel b and c) or various amounts of KOD FX DNA polymerase (Panel d). The cycling conditions consisted of pre-denaturation at 94°C for 2 min, followed by 30 cycles of denaturation at 94°C for 20 s, annealing at 56°C for 20 s and extension at 72°C for 25 s. NTC: no-template control.

    Article Snippet: The results showed that the use of 0.1–0.15 U of high-fidelity DNA polymerase combined with 1 U of Taq DNA polymerase in per 20 μL reaction resulted in the best discrimination and amplification effects ( ).

    Techniques: Blocking Assay, Polymerase Chain Reaction, Mutagenesis, Sequencing

    Optimization of annealing temperature and amount of high-fidelity DNA polymerase in the modified PR-PCR. (a) Discrimination efficiency of the modified PR-PCR for known mutations using a mixture of 1 U of Taq DNA polymerase and various amount (0.05, 0.1 and 0.3 U) of KOD FX DNA polymerase in the reaction. The PCR amplifications were performed in a total volume of 20 μL under cycling conditions consisting of pre-denaturation at 94°C for 2 min, followed by 30 cycles of denaturation at 94°C for 20 s, annealing at 56°C for 20 s and extension at 72°C for 25 s. (b) Optimization of annealing temperature for the modified PR-PCR using a mixture of 1 U of Taq DNA polymerase and 0.1 U of KOD FX DNA polymerase. As the optimal discrimination efficiency was obtained when the annealing temperature was within a scope of 61.8°C–62.6°C, the results from 50°C to 61.2°C were not shown. The temperature 62.2°C was selected for subsequent experiments. (c) Optimization of amount of high-fidelity DNA polymerase for the modified PR-PCR with an input of 1 U of Taq DNA polymerase at an annealing temperature of 62.2°C.

    Journal: PLoS ONE

    Article Title: Modified Proofreading PCR for Detection of Point Mutations, Insertions and Deletions Using a ddNTP-Blocked Primer

    doi: 10.1371/journal.pone.0123468

    Figure Lengend Snippet: Optimization of annealing temperature and amount of high-fidelity DNA polymerase in the modified PR-PCR. (a) Discrimination efficiency of the modified PR-PCR for known mutations using a mixture of 1 U of Taq DNA polymerase and various amount (0.05, 0.1 and 0.3 U) of KOD FX DNA polymerase in the reaction. The PCR amplifications were performed in a total volume of 20 μL under cycling conditions consisting of pre-denaturation at 94°C for 2 min, followed by 30 cycles of denaturation at 94°C for 20 s, annealing at 56°C for 20 s and extension at 72°C for 25 s. (b) Optimization of annealing temperature for the modified PR-PCR using a mixture of 1 U of Taq DNA polymerase and 0.1 U of KOD FX DNA polymerase. As the optimal discrimination efficiency was obtained when the annealing temperature was within a scope of 61.8°C–62.6°C, the results from 50°C to 61.2°C were not shown. The temperature 62.2°C was selected for subsequent experiments. (c) Optimization of amount of high-fidelity DNA polymerase for the modified PR-PCR with an input of 1 U of Taq DNA polymerase at an annealing temperature of 62.2°C.

    Article Snippet: The results showed that the use of 0.1–0.15 U of high-fidelity DNA polymerase combined with 1 U of Taq DNA polymerase in per 20 μL reaction resulted in the best discrimination and amplification effects ( ).

    Techniques: Modification, Polymerase Chain Reaction

    Sensitivity and selectivity of the modified PR-PCR for mutation detection using a fusion-blocked primer and adaptor. (a) Diagram of the fusion-blocked primer and adaptor used in the reactions. (b) Primer and adaptor sequences. (c) Sensitivity of the modified PR-PCR. (d) Selectivity of the modified PR-PCR. All reactions were performed in a total volume of 25 μL containing a mixture of 1 μL of gDNA template at 50 ng/μL, 2.5 μL of (10×) Taq PCR buffer, 0.2 mM dNTPs, 0.15 U of PrimeSTAR HS DNA polymerase, 0.75 U of Taq DNA polymerase, 1 μL of DMSO and 0.3 μM each of the reverse primer, fusion-blocked forward primer and adaptor. The DNA templates were prepared by mixing different amounts of MCF-7 (WT) gDNA (from 0 to 50 ng) among HCC1937 (mutant type, MT) gDNA (from 50 to 0 ng) with concentrations from 0 to 100%. The reactions were performed under cycling conditions consisting of pre-denaturation at 94°C for 2 min, followed by 40 cycles of denaturation at 98°C for 10 s, annealing at 59°C for 30 s and extension at 72°C for 20 s.

    Journal: PLoS ONE

    Article Title: Modified Proofreading PCR for Detection of Point Mutations, Insertions and Deletions Using a ddNTP-Blocked Primer

    doi: 10.1371/journal.pone.0123468

    Figure Lengend Snippet: Sensitivity and selectivity of the modified PR-PCR for mutation detection using a fusion-blocked primer and adaptor. (a) Diagram of the fusion-blocked primer and adaptor used in the reactions. (b) Primer and adaptor sequences. (c) Sensitivity of the modified PR-PCR. (d) Selectivity of the modified PR-PCR. All reactions were performed in a total volume of 25 μL containing a mixture of 1 μL of gDNA template at 50 ng/μL, 2.5 μL of (10×) Taq PCR buffer, 0.2 mM dNTPs, 0.15 U of PrimeSTAR HS DNA polymerase, 0.75 U of Taq DNA polymerase, 1 μL of DMSO and 0.3 μM each of the reverse primer, fusion-blocked forward primer and adaptor. The DNA templates were prepared by mixing different amounts of MCF-7 (WT) gDNA (from 0 to 50 ng) among HCC1937 (mutant type, MT) gDNA (from 50 to 0 ng) with concentrations from 0 to 100%. The reactions were performed under cycling conditions consisting of pre-denaturation at 94°C for 2 min, followed by 40 cycles of denaturation at 98°C for 10 s, annealing at 59°C for 30 s and extension at 72°C for 20 s.

    Article Snippet: The results showed that the use of 0.1–0.15 U of high-fidelity DNA polymerase combined with 1 U of Taq DNA polymerase in per 20 μL reaction resulted in the best discrimination and amplification effects ( ).

    Techniques: Modification, Polymerase Chain Reaction, Mutagenesis

    ( A ) Gel electrophoresis of near full-length genome PCR products produced from four long amplifying DNA polymerases; KlenTaq LA (discontinued), AccuTaq LA, PrimeSTAR GXL and Takara LA Taq (separate gel) per manufacturer’s instructions for samples with high GC/secondary structures. M, HyperLadder 1 kb; 1, CVB3 Nancy; 2, CVB5 Faulkner; 3, H 2 O control. ( B ) Gel electrophoresis of near full-length genome PCR products produced from Takara LA Taq DNA polymerase. M, HyperLadder 1 kb; 1–4, known EV positives from NSW Health Pathology East virology diagnostic lab; 5, CVB3 Nancy; 6, H 2 .

    Journal: Scientific Reports

    Article Title: Amplification and next generation sequencing of near full-length human enteroviruses for identification and characterisation from clinical samples

    doi: 10.1038/s41598-018-30322-y

    Figure Lengend Snippet: ( A ) Gel electrophoresis of near full-length genome PCR products produced from four long amplifying DNA polymerases; KlenTaq LA (discontinued), AccuTaq LA, PrimeSTAR GXL and Takara LA Taq (separate gel) per manufacturer’s instructions for samples with high GC/secondary structures. M, HyperLadder 1 kb; 1, CVB3 Nancy; 2, CVB5 Faulkner; 3, H 2 O control. ( B ) Gel electrophoresis of near full-length genome PCR products produced from Takara LA Taq DNA polymerase. M, HyperLadder 1 kb; 1–4, known EV positives from NSW Health Pathology East virology diagnostic lab; 5, CVB3 Nancy; 6, H 2 .

    Article Snippet: First round PCR was performed by adding cDNA (5 µL) to a reaction mix (50 µL) containing Takara LA Buffer (1X), dNTPs (100 µM each), forward primer vir24 (0.2 µM), reverse primer vir20 (0.2 µM), nuclease-free water (29.5 µL) and Takara LA DNA polymerase (2.5 U).

    Techniques: Nucleic Acid Electrophoresis, Polymerase Chain Reaction, Produced, Diagnostic Assay

    Optimization of the multiplex PCR components. (A) Annealing temperature gradients, M: DL2000 marker. Lanes 1–8: 64, 63, 62, 61, 60, 59, 58, and 57°C, respectively. (B) Dose of Anaplasma primers, lane M: DL2000 marker; Lanes 1–7: 0.08, 0.16, 0.24, 0.32, 0.4, 0.48, and 0.56 μM, respectively; Lane N: negative control. (C) La Taq DNA polymerase. M: DL2000 marker; Lanes 1–7: 0.75, 1.0, 1.25, 1.5, 1.75, 2, and 2.25 U, respectively; Lane N: negative control.

    Journal: Frontiers in Microbiology

    Article Title: A Multiplex PCR Detection Assay for the Identification of Clinically Relevant Anaplasma Species in Field Blood Samples

    doi: 10.3389/fmicb.2020.00606

    Figure Lengend Snippet: Optimization of the multiplex PCR components. (A) Annealing temperature gradients, M: DL2000 marker. Lanes 1–8: 64, 63, 62, 61, 60, 59, 58, and 57°C, respectively. (B) Dose of Anaplasma primers, lane M: DL2000 marker; Lanes 1–7: 0.08, 0.16, 0.24, 0.32, 0.4, 0.48, and 0.56 μM, respectively; Lane N: negative control. (C) La Taq DNA polymerase. M: DL2000 marker; Lanes 1–7: 0.75, 1.0, 1.25, 1.5, 1.75, 2, and 2.25 U, respectively; Lane N: negative control.

    Article Snippet: Optimization of the Multiplex PCR After optimization, the optimum multiplex PCR assay was performed in a final volume of 25 μL, containing 2.5 μL of 10× PCR La buffer, 4 μL of dNTPs at 2.5 mM, 1.25 U of La Taq DNA polymerase, 0.32 μM of each primer, and 2 μL of the DNA template.

    Techniques: Multiplex Assay, Polymerase Chain Reaction, Marker, Negative Control