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KAPA2G Robust PCR Kits

For Research Use Only. Not for use in diagnostic procedures.

KAPA2G Robust PCR

Overview

KAPA2G Robust PCR Kits contain a second-generation DNA polymerase engineered to solve inconsistent amplification across a broad range of amplicon types (GC- and AT-rich).  The high performance of the DNA polymerase is ideally suited for challenging PCR applications and difficult samples, eliminating the need for optimization using multiple enzymes and protocols.

Benefits of KAPA2G Robust PCR Kits

  • High processivity and specific activity, translating to robust PCR performance and high sensitivity
  • Improved tolerance to common PCR inhibitors

Product highlights

Efficient amplification of GC- and AT-rich targets

  • Robust performance across a wide range of GC- and AT-rich templates
  • Increased PCR success rates

View data showing 100% success* in PCR amplifications of amplicons with high GC content (up to 84%) using KAPA2G Robust PCR Kits and 66% success using wild-type Taq polymerase.

Figure 1. Half of each of the PCR products obtained with 72 of the 96 primer sets used in this study were electrophoresed in a 1% TBE-agarose gel. Amplicons were loaded in order of increasing GC content, with the lowest GC content (27%, blue) at the top left hand side and the highest GC content (84%, red) at the bottom right hand side of each composite gel image. Primers selected for this study had variable primer lengths, sequence composition, theoretical melting temperatures and other design features. Some primers contained 5'-tails for post-PCR sequencing using M13 or other standard sequencing primers. KAPA2G Robust HotStart ReadyMix reactions (25 µL) were performed as outlined in the User Guide. All reactions contained 25 ng human genomic DNA. 5% DMSO was included in all reactions (KAPA2G Robust and Taq Kits) targeting amplicons with a GC content >70%.

image 1

Figure 2. Half of each of the PCR products obtained with 72 of the 96 primer sets used in this study were electrophoresed in a 1% TBE-agarose gel. Amplicons were loaded in order of increasing GC content, with the lowest GC content (27%, blue) at the top left hand side and the highest GC content (84%, red) at the bottom right hand side of each composite gel image. Primers selected for this study had variable primer lengths, sequence composition, theoretical melting temperatures and other design features. Some primers contained 5'-tails for post-PCR sequencing using M13 or other standard sequencing primers. KAPA2G Robust HotStart ReadyMix reactions (25 µL) were performed as outlined in the User Guide. All reactions contained 25 ng human genomic DNA. 5% DMSO was included in all reactions (KAPA2G Robust and Taq Kits) targeting amplicons with a GC content >70%.

image 2

Improved tolerance to common PCR inhibitors

  • Efficient amplification from crude samples
  • Higher yield and sensitivity per unit of enzyme

View data showing more efficient amplification of a plasmid DNA sample in the presence of PCR inhibitors such as NaCl, urea, ethanol and SDS using KAPA2G Robust PCR Kits compared to wild type Taq polymerase.

image 3

 

 

Figure 3. Amplification of a 1.5 kb fragment from 1 pg plasmid DNA in the presence of four common PCR inhibitors using the KAPA2G Robust HotStart PCR kit and wild-type hot start <i>Taq</i> polymerase. All reactions contained 0.5 units of enzyme per 25 µL reaction. KAPA2G Robust HotStart Buffer B was used throughout, with the addition of KAPAEnhancer 1 for reactions containing SDS. Cycling was performed with an Eppendorf Mastercycler epgradient S, using a standard 3-step cycling profile (35 cycles) with an annealing temperature of 64ºC and 1.5 min extension time per cycle for all enzymes.

Superior performance in colony PCR compared to wild type Taq

  • Higher yields and improved consistency of PCR directly from E. coli and yeast cells

View data showing superior PCR amplification of E. coli and yeast colonies using KAPA2G Robust PCR Kits compared to wild-type Taq polymerase.

Figure 4. Amplification of a cloned 2.7 kb insert from four commonly used <i>E. coli</i> strains (DH5a, DH10B, JM109 or BL21) using KAPA2G Robust HotStart (top) or wild-type Taq (bottom). Colonies (grown on LB-agar + Amp plates) were either resuspended directly in individual PCR reactions (left) or first resuspended in PCR grade water and then added to PCR reaction mixes (middle). For overnight cultures (prepared in LB + Amp), 1 μL was added directly to the PCR mix

image 4

Figure 5. Amplification of a 2.5 kb (left) or 1.6 kb (right) fragment from the GSH1 gene from three commonly used 

 

 

 

image 5

*Data on file.

Specifications

  • Starting Material

    Genomic DNA, cDNA, plasmid, lambda

  • Input Amount

    < 100 ng for genomic DNA, cDNA < 10 ng for less complex DNA

Kits can be stored for up to 12 months at -20˚C.

Standard kits include KAPA2G Robust (5 U/μL), KAPA2G Buffer A, KAPA2G Buffer B and KAPA2G GC Buffer (all at 5X, with 1.5 mM MgCl2 at 1X), KAPA Enhancer 1 (5X), and supplemental MgCl2 (25 mM). Kit options include HotStart and dNTP Mix (10 mM each).

ReadyMix formulations (2X) of KAPA2G Robust HotStart DNA Polymerase are also available. These contain DNA Polymerase, reaction buffer (with or without dye), dNTPs and MgCl2 (at a final concentration of 2 mM).

Contents

Contents
Components
KAPA2G Robust Standard or HotStart DNA Polymerase (5 U/µL)
5X KAPA2G Buffer A with MgCl2
5X KAPA2G Buffer B with MgCl2
5X KAPA2G GC Buffer with MgCl2
5X KAPA Enhancer 1
MgCl2 (25 mM)
dNTP Mix (10 mM each dNTP; optional)

For purchasing or general questions

For online ordering

KAPA Code
Roche Cat. No
Description
Kit Size
How to buy
KK5004
07960743001
With dNTPs 100 U
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KK5005
07960751001
With dNTPs 250 U
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KK5023
07960859001
Standard kit 100 U
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KK5024
07960867001
Standard kit 250 U
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KK5515
07961057001
HotStart PCR Kit 250 U
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KK5516
07961065001
HotStart PCR Kit, with dNTPs 250 U
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KK5517
07961073001
HotStart PCR Kit 500 U
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KK5518
07961081001
HotStart PCR Kit, with dNTPs 500 U
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KK5522
07961103001
HotStart PCR Kit 100 U
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KK5525
07961120001
HotStart PCR Kit 2500 U
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KK5532
07961197001
HotStart PCR Kit, with dNTPs 100 U
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KAPA Code
Roche Cat. No
Description
Kit Size
How to buy
KK5701
07961332001
HotStart ReadyMix, 100 x 25 µL reactions 1.25 mL
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KK5702
07961359001
HotStart ReadyMix, 500 x 25 µL reactions 6.25 mL
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KK5704
07961375001
HotStart ReadyMix with dye, 100 x 25 µL reactions 1.25 mL
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KK5705
07961383001
HotStart ReadyMix with dye, 500 x 25 µL reactions 6.25 mL
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For purchasing or general questions

For online ordering