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    Friday, 01 March 2013 10:49

    AccuPower PyroHotStart PCR PreMix

    Rate this item
    (1 Vote)

    The AccuPower PyroHotStart Taq PCR PreMix is a PCR master mix containing a thermostable DNA polymerase, thermostable pyrophosphatase, reaction buffer, dNTPs, tracking dye, and patented stabilizer in a ready to use hotstart PCR master mix. 
    Gentaur uses a unique enzyme-mediated HotStart PCR system that provides robust, sensitive and reliable results. 
    Gentaur's Taq DNA Polymerase is inhibited at lower temperatures (< 70°C) by pyrophosphate. This prevents the formation of mis-primed products and primer-dimers during the reaction set up process resulting in improved PCR specificity. However, Taq DNA Polymerase is rendered fully active at temperatures above 70°C via pyrophosphate hydrolysis with a thermostable pyrophosphatase (Figure 1). AccuPower® PyroHotStart Taq PCR PreMix is ideal for nucleic acid amplification reactions involving complex genomic or cDNA templates, very low copy targets, and multiplex reactions.

    accupower-gentaur-antibodies-Enzyme-mediated-HotStart 

    Figure 1. The mechanism of an enzyme-mediated hotstart PCR

     

    Features and Benefits

    Ease of use: Just add template DNA, primers and water to start your reaction. 
    dNTPs, buffer and enzyme are provided.
    Robust Stability: Stable at room temperature for a month, at 4°C for one year and for 2 years in a -20°C freezer.
    Improved specificity: Unique enzyme mediated PyroHotstart system results in greater specificity and more robust reactions
    Reproducibility: Gentaur's strict quality controlled production system ensures that your results will be reproducible experiment after experiment.

     


    Specifications

    5' to 3' exonuclease activity: Yes
    3' to 5' exonuclease activity: No
    3' – A Overhang: Yes
    Fragment size: Up to 5 kb from human genomic DNA and 10 kb from Lambda DNA


    Application

    - High specificity PCR
    - High sensitivity PCR
    - Low-copy target PCR 
    - Multiplex PCR
    - cDNA amplification
    - TA cloning

    accupower-PyroHotstart Taq antibodies-gentaur-1

    Figure 1. Comparison of PCR amplification specificity between AccuPower® PyroHotStart Taq PCR PreMix from Gentaur and other suppliers' Hot start PCR master mix.

    PCR reactions were performed according to each supplier's protocol. The PrP gene was amplified from human genomic DNA with two different primer sets, separately. This data shows that AccuPower® PyroHotStart Taq PCR PreMix has higher amplification efficiency and specificity than other suppliers' Hot start PCR master mix.
    Lane M: 100 bp DNA Ladder(Gentaur, Cat. No.D-1030)
    Lane 1: 100 ng DNA, PrP primer set (500 bp)
    Lane 2: 10 ng DNA, PrP primer set (500 bp) 
    Lane 3: 100 ng DNA, PrP primer set (705 bp)
    Lane 4: 10 ng DNA, PrP primer set (705 bp)

    accupower-gentaur-antibodies-PyroHotstart Taq fig2

    Figure 2. Comparison of PCR amplification specificity between AccuPower PyroHotStart Taq PCR PreMix from Gentaur and other suppliers' Hot start PCR master mix.

    The ApoE gene was amplifed from 100 ng of human genomic DNA (The PCR product size is 268bp). This data shows thatAccuPower PyroHotStart Taq PCR PreMix has higher amplificition efficency and specificity than other suppliers' Hot start PCR master mix.
    Lane M: 100 bp DNA Ladder (Gentaur, Cat. No. D-1030)
    Lane 1: AccuPower PyroHotStart Taq PCR PreMix
    Lane 2: Supplier I Hotstart Taq PCR preMix
    Lane 3: Supplier S Hotstart Taq PCR masterMix
    Lane 4: Supplier T Hotstart Taq PCR masterMix
    Lane 5: Supplier Q Hotstart Taq PCR masterMix

    accupower-PyroHotstart Taq -gentaur-antibodies-fig3

    Figure 3. AccuPower PyroHotStart Taq PCR PreMix has high amplification efficiency and specificity.

    Specificity test was performed using 7 different sets of primers targeting the P53 gene. 10 ng of human genomic DNA was used for each PCR reaction. The cycling conditions were 95°C for 5min, 30 cycles of 95°C for 20 sec, 55°C for 40 sec, and 72°C for 1 min, and 72°C for 5 min for final extension.
    Lane M: 100bp DNA Ladder (Gentaur, Cat. No. D-1030)
    Lane 1: P75/73 primer set (139 bp)
    Lane 2: P55/53 primer set (211 bp)
    Lane 3: P55/63 primer set (447 bp)
    Lane 4: P75/83 primer set (618 bp)
    Lane 5: P55/73 primer set (1082 bp)
    Lane 6: P65/83 primer set (1296 bp)
    Lane 7: P55/83 primer set (1561 bp)

    accupower-gentaur-antibodies-PyroHotstart Taq fig4

    Figure 4. Comparison of PCR amplification between AccuPower PyroHotStart Taq PCR PreMix from Gentaur and other suppliers' Hot start PCR master mix.

    Sensitivity test was performed by amplifying the IRGC gene from a serial dilution of human genomic DNA. This data shows that AccuPower PyroHotStart Taq PCR PreMix has higher amplification efficiency than other suppliers' Hot start PCR master mix.
    Lane M: 100 bp DNA Ladder (Gentaur, Cat. No. D-1030)
    Lane 1: 10 ng human genomic DNA
    Lane 2: 1 ng human genomic DNA
    Lane 3: 100 pg human genomic DNA
    Lane 4: 10 pg human genomic DNA

    accupower-antibodies-gentaur-PyroHotstart Taq fig5

    Figure 5. Comparison of cDNA template amplification between AccuPower PyroHotStart Taq PCR PreMix and other suppliers' Hot start PCR master mix.

    cDNA synthesized from 10-fold serial-diluted human total RNA from 10 ng to 10 pg using AccuPower RocketScript™ Cycle RT PreMix (Gentaur, Cat. No. K-2201) was used as a template for PCR amplification. This data shows thatAccuPower® PyroHotStart Taq PCR PreMix has higher amplification efficiency than other suppliers' Hot start PCR master mix.
    Lane M: 100 bp DNA Ladder (Gentaur, Cat. No. D-1030)
    Lane 1: 10 ng human total cDNA
    Lane 2: 1 ng human total cDNA
    Lane 3: 100 pg human total cDNA
    Lane 4: 10 pg human total cDNA

     

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