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PCR Brochure, March 2002

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PCR Brochure, March 2002 novagen PCR ENZYMES O VERVIEW 2 www.novagen.com PCR ENZYME S ELECTION G UIDE Enzyme PCR Product Elongation Specificity Fidelity GC-rich Yield PCR Product Size Rate Templates Ends KOD HiFi DNA Polymerase < 6 kbp 120 bases/s ● ■■blunt KOD Hot Start DNA Polymerase < 21 kbp 120 bases/s ■■ ■■blunt KOD XL DNA Polymerase < 30 kbp 120 bases/s ● ■■■Mixed (blunt and 3'-dA) NovaTaq™ DNA Polymerase < 5 kbp 60 bases/s ●● ●3'-dA NovaTaq Hot Start DNA Polymerase < 5 kbp 60 bases/s ■ ● ▲ 3'-dA NovaTaq DNA Polymerase + Taq Antibody < 5 kbp 60 bases/s ■ ● ▲ 3'-dA ● Satisfactory ▲ Good ■ Excellent Novagen offers a complete selection of enzymes and kits for PCR, featuring KOD HiFi DNA Polymerase. This unique proofreading enzyme, isolated from the extreme thermophile Thermococcus kodakaraensis KOD1, possess- es superior processivity and fidelity that enable faster, more accurate PCR amplification than with conventional KOD DNA Polymerase enzymes, including Pfu DNA polymerase (1). KOD HiFi DNA Polymerase is also available in a Hot Start version Faster than Taq... for high specificity and increased read length (2), and as a blend (KOD XL DNA Polymerase) recommended for very long templates (3). More accurate than Pfu... NovaTaq™ DNA Polymerase is a high-purity, recombi- nant enzyme suitable for any application requiring premi- um quality Taq DNA polymerase. For increased specifici- ty and convenience with standard PCR, we also offer NovaTaq™ Hot Start DNA Polymerase and the Taq Antibody.NovaTaq Hot Start DNA Polymerase is a chem- ically modified form of Taq DNA polymerase that 1. Takagi, M., Nishioka, M., Kakihara, H., Kitabayashi, M., Inoue, H., becomes active when heated at 95°C for 7–10 minutes. Kawakami, B., Oka, M., and Imanaka, T. (1997) Appl. Environ. Microbiol. 63, The Taq Antibody is available as an alternative means to 4504–4510. provide hot start capability to NovaTaq DNA Polymerase 2. Mizuguchi, H., Nakatsuji, M., Fujiwara, S., Takagi, M., and Imanaka, T. (1999) J. Biochem. (Tokyo) 126, 762–768. as well as other sources of Taq DNA polymerase. Please 3. Nishioka, M., Mizuguchi, H., Fujiwara, S., Komatsubara, S., Kitabayashi, M., refer to the table above as a guide to select the appro- Uemura, H., Takagi, M., and Imanaka, T. (2001) J. Biotechnol. 88, 141–149. priate enzyme combination for your application. 3 T ABLE O F C ONTENTS A MPLIFY (see page 6) KOD POLYMERASES ■ KOD DNA Polymerase is a new, high performance DNA polymerase KOD HiFi DNA Polymerase – Faster than Taq,more accurate than Pfu KOD Hot Start DNA Polymerase – A heat activatable form of KOD HiFi DNA Polymerase for automated PCR set-up KOD XL DNA Polymerase – Recombinant forms of KOD Polymerase blended for long and accurate PCR GENOMIC DNA Genomic DNA – High quality, high integrity human, animal, and yeast DNA RT-PCR TOOLS First Strand cDNA Synthesis Kit – Reliable preparation of templates for RT-PCR One Step RT-PCR Kit – Convenient, one-enzyme, single buffer system for RT-PCR TAQ POLYMERASES AND PCR KITS ■ NovaTaq™ DNA Polymerase is a premium quality recombinant form of Thermus aquaticus DNA polymerase NovaTaq DNA Polymerase – A molecular biology lab essential 10 mM dNTP Mix NovaTaq PCR Kit – Everything for PCR except DNA and primers NovaTaq PCR Kit PLUS – What you need for successful PCR and PCR optimization NovaTaq PCR Master Mix – A ready-to-use 2X concentrated PCR reagent mixture NovaTaq Hot Start DNA Polymerase – For increased specificity and convenient reaction assembly Taq Antibody – Monoclonal antibody for automated Hot Start PCR O RDERING I NFORMATION (see page 24) 4 www.novagen.com P URIFY (see page 14) DNA PURIFICATION SpinPrep™ PCR Clean-up Kit – Rapid purification of PCR products for downstream procedures SpinPrep Gel DNA Kit – Rapid, efficient extraction of DNA from agarose gels SpinPrep Plasmid Kit – Rapid, high quality plasmid minipreps SpinPrep Master Kit – Combo Kit; 20 plasmid preps and 20 DNA gel extractions PELLET PAINT® CO-PRECIPITANTS Pellet Paint Co-Precipitant – Rapid, quantitative precipitation of DNA and RNA, including PCR clean-up Pellet Paint NF Co-Precipitant – Non-fluorescent visible DNA co-precipitant for sequencing and other automated applications C LONE (see page 18) CLONING KITS Perfectly Blunt® Cloning Kits – Efficient “universal” cloning of DNA amplified by any polymerase AccepTor™ Vector Kits – Rapid, direct cloning of DNA amplified with non-proofreading DNA polymerases T RANSFORM (see page 20) COMPETENT CELLS NovaBlue Singles™ Competent Cells – High efficiency transformation of E. coli in less than 8 minutes HT96™ NovaBlue Competent Cells – High-efficiency competent cells pre-dispensed in a 96-well format HT96 Isothermal Block – Efficient thermal transfer for uniform incubations A NALYZE (see page 22) DNA LADDERS AND MARKERS Perfect DNA™ Ladders and Markers PCR Markers 5 KOD POLYMERASES Pure recombinant high fidelity DNA polymerases from Thermococcus kodakaraensis KOD HiFi DNA Polymerase Ultra high fidelity DNA polymerase KOD HiFi DNA Polymerase is a recombinant form of Thermococcus ADVANTAGES kodakaraensis KOD1 DNA polymerase. It is the most efficient •Higher fidelity than Pfu DNA Polymerase (1) thermostable DNA polymerase, exhibiting higher accuracy, elongation rate and processivity than any other commercially available DNA poly- •Higher processivity – sequential nucleotide polymerization merase. KOD HiFi DNA Polymerase possesses 3' ← 5' is between 10 and15X greater than Pfu DNA Polymerase and exonuclease-dependent proofreading activity that enables the Deep Vent* DNA Polymerase (1) polymerase to correct nucleotide misincorporation. The enzyme •Greater yield – extension speed is 2X faster than Taq DNA generates blunt-ended PCR fragments that are suitable for cloning polymerase and 5X faster than Pfu DNA polymerase (1) with Novagen’s Perfectly Blunt® Cloning Kits. • More accurate PCR in a shorter time Each order also includes 10X KOD HiFi DNA Polymerase Buffer 1 and 10X KOD HiFi DNA Polymerase Buffer 2, plus separate vials of 25 mM •No truncated amplification products in PCR reaction MgCl2 and 2mM dNTP Mix. 123 4 KOD HiFi Pfu Taq bp 1 Perfect DNA™ Markers, 0.5–12 kbp Enzyme DNA Polymerase DNA Polymerase DNA Polymerase 12,000 – 2 5.4 kbp PCR product (lambda DNA) 10,000 – Species Thermococcus Pyrococcus Thermus 8000 – 3 2.0 kbp PCR product (plasmid DNA) 6000 – kodakaraensis furiosis aquaticus YT-1 41.6 kbp PCR product (human genomic DNA) 4000 – † –3 –3 –2 Fidelity 3.5 x 10 3.9 x 10 1.3 x 10 3000 – Elongation rate 106–138 25 61 2000 – (bases/second) 1500 – Processivity > 300 bases < 20 bases not determined (nucleotide bases) 1000 – 1. Takagi, M., et al., (1997) Applied and Environmental Microbiology 63, 4504–4510. PCR products amplified using KOD HiFi DNA Polymerase † Fidelity was measured by the authors as mutation frequency in PCR products using DNA fragments from various templates were amplified using 2.5 a sensitive blue/white phenotypic assay with a 5.2 kbp lacZ plasmid as template. units KOD HiFi DNA Polymerase in a standard 100 µl reaction. * Deep Vent is a trademark of New England Biolabs, Inc. Portions of each amplification reaction were analyzed by agarose gel electrophoresis (1.2% TAE). 6 www.novagen.com AMPLIFY KOD Hot Start DNA Polymerase Heat-activatable form of KOD HiFi DNA Polymerase for automated PCR set-up KOD Hot Start DNA Polymerase is a premixed complex of KOD HiFi ADVANTAGES DNA Polymerase and two monoclonal antibodies that inhibit the ← •Highest accuracy, yield and processivity among enzyme’s DNA polymerase and 3' 5' exonuclease activities during PCR commercially available proofreading DNA polymerases (1) assembly. KOD Hot Start DNA Polymerase combines the high fidelity, fast extension speed, and outstanding processivity of KOD HiFi DNA • Amplifies genomic DNA templates up to 12 kbp Polymerase with the high yield and specificity of the antibody-based Hot • Amplifies plasmid DNA templates up to 21 kbp Start technology. KOD Hot Start DNA Polymerase generates blunt-ended PCR fragments that are suitable for cloning with Novagen’s Perfectly • Successfully amplifies GC-rich sequences Blunt® Cloning Kits. • Eliminates mispriming and primer-dimer formation Each order also includes 10X KOD Hot Start DNA Polymerase Buffer • Convenient room temperature set-up plus separate vials of 25 mM MgSO4 and 2mM dNTP Mix. M 8.4 12.3 (kbp) M 1 2 4 6 8 10 12 15 21 M (kbp) Genomic DNA amplification The human myosin heavy chain Lambda DNA amplification gene (8.4 kbp) and human ß-globin The indicated fragments of gene (12.3 kbp) were amplified lambda DNA were amplified using KOD Hot Start DNA with KOD Hot Start DNA Polymerase Polymerase M=Markers M=Markers PCR h i g h f i d e l i t y For ordering information, see page 24. KOD POLYMERASES KOD XL DNA Polymerase High performance enzyme blend for long and accurate PCR KOD XL DNA Polymerase is an optimized blend of KOD HiFi DNA ADVANTAGES Polymerase and a mutant form of KOD HiFi that is deficient in 3' 5'← •Ideal for amplification of large DNA fragments exonuclease activity. The enzyme mixture is designed for reliable from purified DNA or crude samples amplification of long, complex targets with robust yield and high accuracy. KOD XL DNA Polymerase generates PCR products suitable • Amplifies DNA templates up to 30 kbp for cloning with Novagen’s AccepTor™ Vector and Perfectly Blunt® • Successfully amplifies GC-rich sequences Cloning Kits. •Efficiently incorporates derivatized dNTPs Each order also includes 10X KOD XL DNA Polymerase Buffer and 2 mM dNTP Mix. Performance comparison λ DNA fragments were amplified using Taq DNA Polymerase and KOD XL DNA Polymerase M λHind III DNA Markers 58 kbp PCR product 11 kbp PCR product 6 10 kbp PCR product 22 kbp PCR product 7 12 kbp PCR product 34 kbp PCR product 8 15 kbp PCR product 46 kbp PCR product Genomic DNA High molecular weight, high purity DNA for any application ORGANISM SOURCE Bovine blood Novagen’s genomic DNAs are qualified for genomic analysis including Cat blood PCR and library construction.
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