High-Fidelity PCR Kits

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PCR Master Mixes and Kits

Overview

Exclusive Sso7d Fusion Polymerase Technology to Empower Your PCR

Sso7d, a 7 kD protein obtained from Sulfolobus solfataricus, binds to dsDNA without any sequence specificity.

Sso7d is covalently linked to a DNA polymerase and is able to stabilize the polymerase-template complex without compromising the structural integrity, thermal stability, or catalytic activity of the enzyme.

The unique structure of this Sso7d fusion polymerase provides several significant advantages over ordinary DNA polymerase:

  • Increased processivity
  • Effective amplification in GC-rich regions or strong secondary structure
  • Efficient amplification of longer products
  • Superior tolerance to PCR inhibitors
Sso7D Technology
iProof™ Polymerase Benefits

Comparison of common thermostable DNA polymerases. iProof Polymerase exhibits the highest fidelity of any commercially available thermostable polymerase, >50-fold higher than ordinary Taq polymerase. A common high-fidelity polymerase was ~8-fold higher than Taq, and an ultrahigh-fidelity polymerase was ~32-fold higher.

iProof Polymerase demonstrates unrivaled speed, leading to dramatically shorter overall reaction times. The reaction protocol for iProof Polymerase was compared to the recommended protocols for two competing polymerases. Each protocol was designed to amplify 1, 8, or 15 kb products in 30 cycles. Reactions with iProof Polymerase used a two-step protocol with a combined annealing and extension step, while the other reactions used three-step protocols with the minimum recommended extension times. Overall reaction times include temperature ramping times.

iProof™ Polymerase Performance Data

Comparison of amplification using iProof Polymerase and another supplier's fast- cycling enzyme. Manufacturer's recommended cycling conditions were used for both enzymes. Left, a 1 kb fragment was amplified from 10 ng human genomic DNA using 20 U/ml of iProof Polymerase (lanes 6–9) or 50 U/ml of the other enzyme (lanes 20–24) in a standard-ramping thermal cycler. Total run time for protocols using a 30 sec annealing/extension step was 48 min (lanes 6, 7, 20, and 21). Total run time for protocols using a 15 sec annealing/extension step was 39 min (lanes 8 and 9) or 40 min (lanes 22 and 23). Lane 5, 500 bp EZ Load marker. Right, the same 1 kb fragment was amplified from 100 ng human genomic DNA on a fast-ramping thermal cycler, using 50 U/ml of iProof Polymerase (lane 3) or 100 U/ml of the other enzyme (lane 4), and using an annealing/extension step of 30 sec. Total run time on the fast ramping cycler was 30 min. Lane 2, 500 bp EZ Load Marker.

Comparison of amplification using iProof Polymerase and another supplier's fast- cycling enzyme. Manufacturer's recommended cycling conditions were used for both enzymes. Left, a 1 kb fragment was amplified from 10 ng human genomic DNA using 20 U/ml of iProof Polymerase (lanes 6–9) or 50 U/ml of the other enzyme (lanes 20–24) in a standard-ramping thermal cycler. Total run time for protocols using a 30 sec annealing/extension step was 48 min (lanes 6, 7, 20, and 21). Total run time for protocols using a 15 sec annealing/extension step was 39 min (lanes 8 and 9) or 40 min (lanes 22 and 23). Lane 5, 500 bp EZ Load marker. Right, the same 1 kb fragment was amplified from 100 ng human genomic DNA on a fast-ramping thermal cycler, using 50 U/ml of iProof Polymerase (lane 3) or 100 U/ml of the other enzyme (lane 4), and using an annealing/extension step of 30 sec. Total run time on the fast ramping cycler was 30 min. Lane 2, 500 bp EZ Load Marker.

iProof Polymerase demonstrates increased speed with minimal amounts of enzyme. The gel images below compare the performance of three thermostable polymerases in amplifying a 1 kb DNA fragment using a two-step protocol that combined annealing and extension times ranging from 7 to 240 sec. iProof Polymerase successfully amplified the target with the shortest extension time (15 sec). Competing enzymes required an annealing/extension step of 30–60 sec or more and 5- to 10-fold more enzyme to produce comparable yields.

Annealing/extension time, sec
iProof High-Fidelity Polymerase Competitor High-Fidelity Polymerase Taq Polymerase

iProof DNA Polymerase delivers high yields with significantly less enzyme. A 1 kb fragment was amplified from 1.25 pM of lambda DNA template in 20 cycles, using combined annealing/extension step of variable duration as indicated. All enzymes were used according to manufacturer recommendations.



iProof™ High-Fidelity DNA Polymerase is a unique Pyrococcus-like proofreading enzyme fused to the Sso7d dsDNA-binding protein to create a thermostable fusion polymerase that accurately amplifies long products from a variety of DNA templates.

Features and Benefits of iProof High-Fidelity DNA Polymerase:

  • High fidelity — iProof DNA polymerase is 52-fold more accurate than Taq polymerase
  • Speed — high processivity dramatically reduces extension (15–30 sec/kb) and overall reaction times
  • Successful amplification of long products with higher yields — fragments up to 37 kb are amplified in less time and with less enzyme (0.25–1 unit/reaction)

Choose the best DNA polymerase for your application.

Key Features iProof DNA Polymerase iTaq DNA Polymerase
Speed
Processivity
Long fragment amplification
Proofreading Yes No
Hot-Start No Yes
 

Related Categories

Supporting Documentation

Storage at –20°C iProof™ Enzyme, Kit, and Buffers: guaranteed for 6 months at –20°C in a constant-temperature freezer
iProof Master Mixes: guaranteed for 3 months at –20°C in a constant-temperature freezer (multiple freeze/thaws not recommended)
Storage at +4°C Guaranteed for 3 months at 4°C
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iProof™ HF Master Mix, 100 x 50 µl rxns, 2.5 ml

1725310
100 x 50 µl reactions, premixed PCR reagents, includes 2x master mix (0.04 U/µl), DMSO; suitable for most templates

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$236.00
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iProof™ GC Master Mix, 100 x 50 µl rxns, 2.5 ml

1725320
100 x 50 µl reactions, premixed PCR reagents, includes 2x master mix (0.04 U/µl), DMSO; suitable for GC-rich templates

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$326.00
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iProof™ High-Fidelity DNA Polymerase, 20 U (2 U/µl), 10 µl

1725300
20 U, 2 U/µl, stand-alone enzyme, includes 5x reaction buffers, MgCl2 solution, DMSO

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$30.00
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iProof™ High-Fidelity DNA Polymerase, 100 U (2 U/µl), 50 µl

1725301
100 U, 2 U/µl, stand-alone enzyme, includes 5x reaction buffers, MgCl2 solution, DMSO

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$150.00
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iProof™ High-Fidelity DNA Polymerase, 500 U (2 U/µl), 250 µl

1725302
500 U, 2 U/µl, stand-alone enzyme, includes 5x reaction buffers, MgCl2 solution, DMSO

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$619.00
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iProof™ High-Fidelity PCR Kit, 50 U (2 U/µl), 25 µl

1725330
50 U, 2 U/µl, iProof polymerase PCR kit includes 5x reaction buffers, MgCl2 solution, DMSO, dNTPs, λ DNA, 1.3 and 10 kb primers, DNA standard

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$96.00
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iProof™ High-Fidelity PCR Kit, 200 U (2 U/µl), 100 µl

1725331
200 U, 2 U/µl, iProof DNA polymerase PCR kit includes 5x reaction buffers, MgCl2 solution, DMSO, dNTPs, λ DNA, 1.3 and 10 kb primers, DNA standard

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$336.00
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Number Description Options
10002299 Instruction Manual, iProof High-Fidelity Master Mix, Rev E Click to download
10002298 Instruction Manual, iProof High-Fidelity DNA Polymerase, Rev B Click to download
10002300 Instruction Manual, iProof High-Fidelity PCR Kit, Rev B Click to download
5211 iProof High-Fidelity DNA Polymerase Flier, Rev C Click to download [ Add to Cart (Free) ]
6252 Reagent Comparison Guide for Real-Time PCR Brochure, Rev C Click to download [ Add to Cart (Free) ]
6090 Amplification Reagents and Plastics Brochure, Rev E Click to download [ Add to Cart (Free) ]
5337 Long Inverse PCR Using iProof Polymerase, Rev A Click to download
5856 Mutational Analysis of the ATM Gene in Familial Breast Cancer Using iProof High-Fidelity DNA Polymerase, Rev A Click to download [ Add to Cart (Free) ]