Phusion cannot incorporate dutp and is not recommended for use with uracilcontaining primers or template. Pdf singlestep purification of t7 rna polymerase with a. Both buffers contain mgcl 2 at a concentration of 7. Promega t7 rna polymerase dnadependent rna polymerase for in vitro transcription. Gotaq dna polymerasea,b is a taq dna polymerase supplied in a proprietary formulation. Wang department of pathology stanford university school of medicine stanford, california 943055324 enzymatic properties and characteristics that distinguish each dna polymerase during the past decade, five dna polymerases pol have been charac terized in eukaryotic cells. A protocol for gotaq flexi dna polymerase which provides robust amplification equal to and in some cases superior to conventional taq dna polymerase. The enzyme is a taq dna polymerase supplied in a proprietary formulation containing 50% glycerol with buffers designed for enhanced amplification.
Crimson taq reaction buffer contains a density reagent, which allows direct loading of pcr products onto a gel. Which hot start polymerase for multiplex str amplification. Recombinase polymerase amplification rpa, combined with lateral flow lf strips, is a novel. The gotaq pcr core system is supplied with gotaq dna polymerase, a proprietary formulation of taq dna polymerase designed for enhanced amplification, and gotaq flexi buffers, magnesium free proprietary buffer formulations supplied at ph 8. Gotaq g2 dna polymerase is supplied with 5x green gotaq reaction buffer and 5x colorless gotaq reaction buffer. Promega manufacturing and delivery systems continue to be fully operational during the covid19 outbreak. Pdf a novel recombinase polymerase amplification rpa. Crimson taq dna polymerase offers superior performance in its newly formulated pcr buffer. Gotaq mdx hot start polymerase contains gotaq mdx dna polymerase bound to a proprietary antibody that blocks polymerase activity, which is restored during the initial denaturation step when amplification reactions are heated at 9495c for two minutes, allowing hotstart pcr. Only t7 dna or dna cloned downstream from a t7 promoter can serve as a template for t7 rna polymerase directed rna synthesis. Luciferase assay systems quick protocol card keywords. Readytouse master mix format with gotaq dna polymerase. Restriction enzyme activity in promega 10x buffers, reaction temperature and heat inactivation. Endpoint and realtime pcr products feature gotaq taq dna polymerase in convenient master mixes, kits and flexible enzyme formulations for basic pcr, hotstart pcr, longrange pcr, qpcr, and rt.
Our teams are in regular contact with suppliers and distributors worldwide and are taking all steps necessary to address both demands for diagnostic tools and reliable delivery of all products as quickly as possible. T7 rna polymerase is a dnadependent phage rna polymerase that exhibits extremely high specificity for its cognate promoter sequence. This item has been discontinued by the manufacturer and is no longer available. Email promega technical services if you have questions on use of this system.
Pdf characterization of dna polymerase from pyrococcus. Promega corporation 2800 woods hollow road madison, wi 537115399 usa telephone 6082744330 toll free 8003569526 fax 6082772516 internet. Assay for taq dna polymerase activity to assay for taq dna polymerase activity, students perform pcr using the supernatant fraction as the dna polymerase component. Luminescent signals generated in the dualluciferase.
T7 rna polymerase does not recognize sp6 or t3 rna polymerase promoter sequences as a start site for transcription. Gotaq dna polymerase product information promega corporation. Highperformance gotaq g2 dna polymerase with mgfree buffer system. Mytaq dna polymerase is a high performance polymerase that exhibits more robust amplification than other commonly used polymerases, delivering very high yield over a wide range of pcr templates and making it the ideal choice for most pcr assays. Although pcr is simple in theory, it can benefit from optimization of several parameters. Promega taq dna polymerase 10x reaction buffers life.
From endpoint pcr to quantitative realtime pcr and rtpcr, we offer a complete portfolio of highquality products to meet your amplification needs. The dna polymerase gene from the archaeon pyrococcus sp. The enzyme is a fulllength form of taq dna polymerase that exhibits 5. Recombinant taq is a 94kda enzyme with an optimal 53 polymerase activity between 7580 c in the presence of a magnesium cofactor.
Promega notes number 83 2003 21 article subhead abstract gotaq dna polymerase, promegas new formulation of taq dna polymerase, is supplied with two buffersgreen gotaq reaction buffer and colorless gotaq reaction buffer. Dna amplified using gotaq dna polymerase with green. Gotaq flexi dna polymerase offers robust amplification equal to and in some cases superior to conventional taq dna polymerase, allows optimization of mgcl 2 concentration, and facilitates directtogel analysis of pcr products. New formulation of taq dna polymerase with enhanced performance over. Assay advantages include broad dynamic range, ease of use and high sensitivity. Gotaq flexi dna polymerase m829 protocol pdf 207 kb english. Protocol phusion highfidelity pcr master mix with hf buffer. Free of detectable levels of contaminating rnase and dnase activity incorporates 32p, 33p, 3h and 35s nucleoside triphosphates. A protocol for a vector containing five copies of an nfkappab response element nfkappabre that drives transcription of the luciferase reporter gene luc2p, a synthetically derived luciferase sequence with humanized codon optimization, designed for high expression and reduced anomalous transcription. Pdf direct inhibition of rna polymerase ii transcription. Gotaq mdx dna polymerase is a fulllength form of taq dna polymerase that exihibits 5.
Making taq dna polymerase in the undergraduate biology. Promega corporation2800 woods hollow road madison, wi 537115399 usa toll free in usa 8003569526 telephone 6082744330 fax 6082772601 printed in usa. To be the most responsive supplier of biological reagents and reagent systems used in research and applied technology applications worldwide. I used to use pfu turbo from stratagene for proof reading 95 degree for 2min initial denaturation and 30sec for each cycle, 5 degree below the primer tm for annealing,72 degree extension for 1kbmin, and 10 min 72 degree extra extentionand everything works fine.
Dna and rna extraction, plasmid purification, reporter assays, pcr, qpcr, cloning enzymes, cell viability assays, apoptosis detection systems, luminometers, protein expression systems and more. T3 rna polymerase is a dnadependent phage rna polymerase that exhibits extremely high specificity for its cognate promoter sequence. Links to the entire contents of the promega product catalog are accessible from this page. T7 rna polymerase is a dnadependent rna polymerase that exhibits extremely high specificity for its cognate promoter sequences. Dualluciferase reporter assay system harvard university. Does anybody here use pfu polymerase from promega for pcr amplification please. Commercial producers have sinced cloned and expressed the enzyme in e. Allows optimization of mgcl 2 concentration and facilitates directtogel analysis of pcr products. This is due in part to the fact that the stabilization free energy for klentaq polymerase, at 27. Gotaq mdx dna polymerase is a fulllength form of taq dna polymerase that exhibits.
T3 rna polymerase is available in the ribomax and riboprobe systems. Ampliqon taq dna polymerase has a molecular weight of 95 kda and exhibits both a 53 dna polymerase and a 53 exonuclease activity. Secondgeneration, highperformance gotaq g2 dna polymerase with mg free buffers. T3 rna polymerase does not recognize sp6 or t7 rna polymerase promoter sequences as a start site for transcription. In many cases, good activity is also obtained using one of the.
Dnadependent rna polymerase specific for t7 promoter sequences 90% pure as determined by sds polyacrylamide gel electrophoresis. The chk1 kinase enzyme system can be purchased with or without the adpglo kinase assay reagents. Amplification of difficult targets, such as those with gcrich sequences or secondary structure, may be improved by the presence. Size units p207b 1,000 p207e 5,000 p407a high conc. Green master mix, 2x gotaq hot start master mixes are allinone master mixes just add template and primers that contain the highperformance gotaq dna polymerase bound to a proprietary antibody that blocks polymerase activity until the reaction is heated at 9495c for two. Pdf characterization of dna polymerase x from thermus. For example, only t7 dna or dna cloned downstream from an t7 promoter can serve. Page 4 revised 806 0 10 20 30 40 50 60 70 80 90 100 02468 10 12 activity % peak time sec firefly renilla figure 2. T7 rna polymerase is available in promegas ribomax and riboprobe systems. Taq polymerase makes a huge difference when multiplexing. Instructions for use of products m8291, m8295, m8296, m8297, m8298.
At 1x concentration, phusion master mix provides 1. The development of a method to rapidly diagnose neospora caninum infection is highly desirable. Product components and storage conditions smallscale isolation minipreps product size cat. Promega corporation 2800 woods hollow road madison, wi 537115399 usa toll free in usa 8003569526 telephone 6082744330 fax 6082772601 printed in usa. Characterization of dna polymerase x from thermus thermophilus hb8 reveals the polxc and php. Promega m3005 gotaq dna polymerase, 4 x 1ml 5x green gotaq reaction buffer and 4 x 1ml 5x colorless gotaq reaction buffer, mgcl2 at a concentration of 7. Gotaq flexi dna polymerasea,b is supplied in a proprietary. The 53 exonuclease activity leaves a 3da overhang on the pcr product, which are convenient for direct ta cloning. Luciferase assay systems quick protocol card, fb026 author. Enhanced performance over standard taq dna polymerase. Sp6, t3 and t7 rna polymerases are dnadependent rna polymerases that exhibit extremely high specificity for their cognate promoter sequences. Making taq dna polymerase in the undergraduate biology laboratory.
No protein species are observed from an uninduced control culture where no iptg was added figure 2, lane 1. Restriction enzyme activity in promega 10x buffers, reaction. The glycerol free product formulation is further purified to remove glycerol, making the enzyme suitable for further manufacture processing and lyophilization. I have tested at least 8 different taqs in side by side experiments and two performed the best. English canada download sdspdf 103 kb english united states. Taq dna polymerase is a thermostable dna polymerase that possesses a 5. The 10x reaction buffer supplied with each restriction enzyme is optimized to give 100% activity. Promega corporation 2800 woods hollow road madison, wi 537115399 usa telephone 6082744330 toll free 8003569526 fax 6082772516 internet pcr satisfaction guarantee promega pcr systems, enzymes and reagents are proven in pcr to ensure reliable, highperformance results. Download fulltext pdf direct inhibition of rna polymerase ii transcription by recql5 received for publication, may 1, 2009, and in revised form, june. Gotaq mdx dna polymerase, glycerolfree, product information pdf 210 kb english. Certificate of analysis gotaq flexi dna polymerase. The enzyme also exhibits 35 exonuclease proofreading activity, 53 exonuclease activity, and ribonuclease h activity. Restriction enzyme activity in promega 10x buffers.
Promega corporation is a worldwide leader in applying biochemistry and molecular biology to the development of innovative, highvalue products for the life sciences. Pfu dna polymerase is tested for performance in the polymerase chain reaction pcr using 1. Gotaq g2 dna polymerase reliably amplifies a wide range of pcr templates and provides. Thermo scientific dna polymerase i, a templatedependent dna polymerase, catalyzes 53 synthesis of dna. Gotaq dna polymerase contains gotaq dna polymerase, 5x green gotaq reaction buffer and 5x colorless gotaq reaction buffer. Dualluciferase reporter assay system protocol promega.