Phusion Polymerase Error Rate
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4.4 X 10-7 in Phusion HF buffer and 9.5 X 10-7 in GC buffer as reported by Finnzymes/Thermo Scientific. Links to taq polymerase error rate this resource Products: Phusion High-Fidelity PCR Master Mix with GC Buffer, Phusion
Pcr Error Rate Calculator
High-Fidelity DNA Polymerase, Phusion High-Fidelity PCR Master Mix with HF Buffer, Phusion High-Fidelity PCR Kit Products Product CatalogNew dna polymerase error rate ProductsSpecial Offers My NEB Products Save your favorite products by clicking Add to My NEB, making re-ordering and remembering what you need quick and simple. Tools & Resources Returning to use
Rna Polymerase Error Rate
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development of high-fidelity polymerases has for many years been a
Pcr Fidelity Calculator
key focus at New England Biolabs (NEB). Highfidelity amplification is
Taq Polymerase Proofreading
essential for experiments whose outcome depends upon the correct DNA sequence (e.g., cloning, SNP analysis, NGS dna polymerase fidelity comparison applications). Whereas traditional fidelity assays are sufficient for Taq and other moderately faithful enzymes, Q5, an ultra highfidelity enzyme, pushes the limits of current methods https://www.neb.com/faqs/2012/09/06/what-is-the-error-rate-of-phusion-reg-high-fidelity-dna-polymerase used to assess this critical feature of DNA polymerases. John A. Pezza, Ph.D., Rebecca Kucera, M.S., Luo Sun, Ph.D., New England Biolabs, Inc. Introduction: What is fidelity? The fidelity of a DNA polymerase is the result of accurate replication of a desired template. Specifically, this involves multiple steps, including the ability https://www.neb.com/tools-and-resources/feature-articles/polymerase-fidelity-what-is-it-and-what-does-it-mean-for-your-pcr to read a template strand, select the appropriate nucleoside triphosphate and insert the correct nucleotide at the 3´ primer terminus, such that Watson-Crick base pairing is maintained. In addition to effective discrimination of correct versus incorrect nucleotide incorporation, some DNA polymerases possess a 3´→5´ exonuclease activity. This activity, known as “proofreading”, is used to excise incorrectly incorporated mononucleotides that are then replaced with the correct nucleotide. High-fidelity PCR utilizes DNA polymerases that couple low misincorporation rates with proofreading activity to give faithful replication of the target DNA of interest. When is fidelity important? Fidelity is important for applications in which the DNA sequence must be correct after amplification. Common examples include cloning/subcloning DNA for protein expression, SNP analysis and next generation sequencing applications. Fidelity is less important for many diagnostic applications where the read-out is simply the presence or absence of a product. Figure 1. DNA Replication with a Proofreading Polymerase Exte
$0.97 link Phusion HF The following table lists a variety of polymerases, some immediately available in lab and some not, and useful information about each of them. Taq Platinum Taq KOD PFU Vent Phusion Species http://barricklab.org/twiki/bin/view/Lab/ProtocolsTaq Thermophilis Aquaticus Mix of Taq, Pyrococcus species GB-D polymerase and Platinum® Taq Antibody Thermococcus kodakaraensis Pyrococcus furiosis Thermococcus litoralis Pyrococcus + Processivity domain Fidelity (err/bp) 10–4 to 10–5 10–5 to 10–6 2.6x10–6 1-3 x 10–6 10–6 10–6 to 10–7 Elongation Rate 1 min/kb 1 min/kb 20-30 sec/kb 1 min/kb 1 min/kb 20-30 sec/kb Processivity ~ 50 bases > 50 bases >300 bases 20 ? 35 Nicking Activity None None None ? < error rate 10% ? Amplicon Size < 5kbp Up to 20 kbp Plasmid/Linear = 6kbp Genome = 2kbp Vector: Up to 15 kbp Genome = Up to 19 kbp ? Genome = 10 kbp Plasmid/Linear = 20 kbp 3' → 5' exonuclease? (proofreading) No Yes (Pyrococcus) Yes Yes Yes Yes 5' → 3' exonuclease? Yes Yes No No No No Strand displacing? Yes Yes No No Yes No Overhang? 3' - A 3' - A polymerase error rate No No No No GC-Rich Performance Low Low High Medium High Medium Hot Start? No Yes Available Available No Available Applications Standard Taq Diagnostic PCR Since Taq is the cheapest polymerase available, it is the most appropriate for diagnostic applications requiring multiple reactions, such as checking if your cloning or genomic insertion worked via PCR of the intended insert region. Colony PCR Taq is especially well-suited to PCR on the unpurified genome, as in colony PCR. Taq will more successfully (and less expensively) amplify a single product off a small number of cells than Phusion. Phusion High Fidelity High-fidelity PCR The error rate of Phusion DNA Polymerase is 4.4 x 10^-7; therefore, Phusion DNA Polymerase is suitable for all PCR applications requiring great accuracy. Cloning Phusion DNA Polymerase amplifies templates with an accuracy and speed previously unattainable with a single enzyme. This makes it a superior choice for cloning. Phusion DNA Polymerase possesses 5' - 3' polymerase activity and 3'-5' exonuclease activity and will generate blunt ended products. Specialized Cloning Techniques (Gibson, CPEC, SLIC, Overlap PCR) All of these specialized cloning techniques require that the polymerase being used does not have strand displacement or 5' → 3' exonuclease activity. As such, and given its speed and high fidelity, Phusion is an excellent choice for them. Long amplicons