Advantage GC Genomic LA Polymerase Mix
Advantage GC Genomic LA Polymerase Mix
Brand: Takara Bio.
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Advantage GC Genomic LA Polymerase Mix
Advantage GC Genomic LA Polymerase Mix
Advantage Genomic LA Polymerase Mix and Advantage GC Genomic LA Polymerase Mix are designed for long and accurate (LA) PCR amplification. These mixes offer 6.5X higher fidelity than wild-type Taq DNA polymerase, and allow synthesis of products up to 30 kb for complex templates such as genomic DNA, and up to 48 kb for less complex templates such as λ DNA. Both versions of Advantage Genomic GC Polymerase Mix are composed of a full-length Taq DNA polymerase, a small amount of proofreading enzyme, and a hot-start antibody, and are supplied with optimized buffers.
The Advantage GC Genomic LA Polymerase Mix includes a 2X Advantage GC-Melt buffer to allow efficient amplification of GC-rich templates, which have strong secondary structures that resist denaturation and prevent efficient primer annealing. This kit can handle GC-rich sequences (up to 90% GC) that resist standard PCR amplification techniques, and enables synthesis of PCR products up to 20 kb from GC-rich human genomic DNA.
Overview
- Designed for 'long and accurate' PCR amplification—amplify products as large as 48 kb
- 6.5X higher fidelity than wild-type Taq DNA polymerase—generates virtually error-free PCR products
- High sensitivity—amplify regions of GC-rich DNA, such as a 1.9-kb region of the TGF-beta gene (GC content = 69%), from as little as 10 pg of template
- Antibody-mediated hot start—increase specificity by preventing nonspecific amplification before thermal cycling
- Advantage GC-Melt Buffer system weakens base pairing in GC-rich sequences—optimized for use with complex, GC-rich genomic templates
Applications
- Long PCR
- GC-rich PCR
- Difficult PCR
Removal of incorrectly incorporated bases by 3' to 5' exonuclease activity
Removal of incorrectly incorporated bases by 3' to 5' exonuclease activity. The 3' to 5' exonuclease removes incorrect bases, allowing product extension to proceed quickly and efficiently.
Conceptualized GC-Melt mechanism
Conceptualized GC-Melt mechanism. The GC-Melt buffer reduces the strength of GC base pairing, enabling efficient amplification of DNA sequences that have a GC-content of up to 90%.
Advantage Genomic LA Polymerase Mix amplifies a wide range of target sizes
Advantage Genomic LA Polymerase Mix amplifies a wide range of target sizes. Advantage Genomic LA was used to PCR-amplify a range of targets from human genomic DNA (100 ng/reaction; Panel A) and lambda DNA (1 ng/ reaction; Panel B). Amplicons of the following sizes were produced: Lane 1: 4 kb; Lane 2: 8 kb; Lane 3: 15 kb; Lane 4: 20 kb; Lane 5: 24 kb; Lane 6: 31 kb; Lane 7: 4 kb; Lane 8: 8 kb; Lane 9: 15 kb; Lane 10: 20 kb; Lane 11: 30 kb; Lane 12: 40 kb; Lane 13: 43 kb; Lane M: lambda-HindIII DNA size marker.
Advantage GC Genomic LA exhibits high sensitivity on GC-rich targets
Advantage GC Genomic LA exhibits high sensitivity on GC-rich targets. Advantage GC Genomic LA was used to amplify a 1.9 kb, GC-rich region (GC content: 69%) of the TGF-beta gene. The amount of template used in each reaction is as follows: Lane 1: 100 ng; Lane 2: 10 ng; Lane 3: 1 ng; Lane 4: 100 pg; Lane 5: 10 pg; Lane 6: No template control. Lane M: Mixture (1:1) of lambda-HindIII and phi-X174-HaeIII digests.
Increased yields from GC-rich templates with Advantage GC Genomic LA Polymerase Mix
Increased yields from GC-rich templates with Advantage GC Genomic LA Polymerase Mix. Reaction yields from three templates of differing size and GC content, using Advantage Genomic LA enzyme system with either the Advantage GC-Melt buffer (Lanes 1, 3, and 5) or the Advantage Genomic LA buffer (Lanes 2, 4, and 6) are compared. PCR reactions were diluted fivefold, and 5 µl was loaded in each well. Lanes 1 and 2: c-jun, 1.255 kb, 65% GC; Lanes 3 and 4: TGF-beta, 1.9 kb, 69% GC; Lanes 5 and 6: IGFR2, 0.5 kb, contains a 100 bp region that is 90% GC-rich; Lane 7: no template control, Lane M: Mixture (1:1) of lambda-HindIII and phi-Χ174-HaeIII digests
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Removal of incorrectly incorporated bases by 3' to 5' exonuclease activity
Removal of incorrectly incorporated bases by 3' to 5' exonuclease activity. The 3' to 5' exonuclease removes incorrect bases, allowing product extension to proceed quickly and efficiently.
Conceptualized GC-Melt mechanism
Conceptualized GC-Melt mechanism. The GC-Melt buffer reduces the strength of GC base pairing, enabling efficient amplification of DNA sequences that have a GC-content of up to 90%.
Advantage Genomic LA Polymerase Mix amplifies a wide range of target sizes
Advantage Genomic LA Polymerase Mix amplifies a wide range of target sizes. Advantage Genomic LA was used to PCR-amplify a range of targets from human genomic DNA (100 ng/reaction; Panel A) and lambda DNA (1 ng/ reaction; Panel B). Amplicons of the following sizes were produced: Lane 1: 4 kb; Lane 2: 8 kb; Lane 3: 15 kb; Lane 4: 20 kb; Lane 5: 24 kb; Lane 6: 31 kb; Lane 7: 4 kb; Lane 8: 8 kb; Lane 9: 15 kb; Lane 10: 20 kb; Lane 11: 30 kb; Lane 12: 40 kb; Lane 13: 43 kb; Lane M: lambda-HindIII DNA size marker.
Advantage GC Genomic LA exhibits high sensitivity on GC-rich targets
Advantage GC Genomic LA exhibits high sensitivity on GC-rich targets. Advantage GC Genomic LA was used to amplify a 1.9 kb, GC-rich region (GC content: 69%) of the TGF-beta gene. The amount of template used in each reaction is as follows: Lane 1: 100 ng; Lane 2: 10 ng; Lane 3: 1 ng; Lane 4: 100 pg; Lane 5: 10 pg; Lane 6: No template control. Lane M: Mixture (1:1) of lambda-HindIII and phi-X174-HaeIII digests.
Increased yields from GC-rich templates with Advantage GC Genomic LA Polymerase Mix
Increased yields from GC-rich templates with Advantage GC Genomic LA Polymerase Mix. Reaction yields from three templates of differing size and GC content, using Advantage Genomic LA enzyme system with either the Advantage GC-Melt buffer (Lanes 1, 3, and 5) or the Advantage Genomic LA buffer (Lanes 2, 4, and 6) are compared. PCR reactions were diluted fivefold, and 5 µl was loaded in each well. Lanes 1 and 2: c-jun, 1.255 kb, 65% GC; Lanes 3 and 4: TGF-beta, 1.9 kb, 69% GC; Lanes 5 and 6: IGFR2, 0.5 kb, contains a 100 bp region that is 90% GC-rich; Lane 7: no template control, Lane M: Mixture (1:1) of lambda-HindIII and phi-Χ174-HaeIII digests
