Taq DNA Polymerase E (high efficiency)

The High-Fidelity Pwo proofreading DNA polymerase from Genaxxon bioscience is a thermostable, highly processive enzyme possessing 5'-3' DNA polymerase with additional 3'-5' proofreading exonuclease activity, which enables the correction of nucleotide incorporation errors. It has no 5'→3' exonuclease activity. The High-Fidelity Pwo DNA polymerase is a recombinant form of the hyperthermophilic archaebacteria Pyrococcus woesei. Pwo proofreading DNA polymerase shows an increased thermostability and a 10-times higher accuracy compared to Taq DNA polymerase. A mixture of Taq DNA Polymerase and Pwo DNA polymerase provides more robust synthesis of longer amplification products (Barnes, 1994. Proc. Natl. Acad. Sci. USA 91:2216-2220). Test sample available at a special price! The test sample price will be refunded on the first official order of the product. Features: 10-times higher accuracy compared to Taq DNA polymerase High-Fidelity polymerase Proofreading function (3' - 5' exonuclease activity) High thermo stability Generates blunt-end PCR products Generates PCR products for cloning and expression More High-Fidelity Proofreading Polymerases from Genaxxon bioscience:- M3003 ReproFast Proofreading Polymerase- M3004 Pfu Proofreading Polymerase- M3012 ReproHot (KOD) Proofreading Polymerase - AQ97 High Fidelity proofreading Polymerase With our high quality dNTPs as Set (M3015) or Mix (M3016) or our DNA Ladders and our favourable standard agarose (M3044) we can offer additional products for your PCR.

ReproFast DNA Polymerase – Maximum Reliability for Your Standard PCRs ReproFast DNA Polymerase – the robust all-round solution for efficient and precise amplification in your daily laboratory workflows. This innovative blend of classic Taq DNA polymerase and a proofreading polymerase in a finely tuned buffer delivers superior results across nearly all standard PCR applications – with minimal optimization effort. ReproFast reliably handles standard PCRs up to 5 kb from human genomic DNA and up to 7 kb from lambda DNA with consistently high yield. Save time and reduce costs: uniform reaction conditions cover a broad range of template types, from routine assays to challenging targets. The All-Round Polymerase – Key Advantages Perfect balance: Taq for power and proofreading for accuracy – the ideal combination of speed, yield, and fidelity Universally applicable: Optimized buffer for diverse standard PCR applications, even with varying templates Proven performance: Up to 5 kb (human gDNA) / 7 kb (lambda DNA) – validated in routine and research workflows Workflow boost: Fewer enzymes, less optimization, more results Practical Benefits for the Laboratory Streamlined PCR setups for cloning, genotyping, and screening High reproducibility for service laboratories and high-throughput applications A single enzyme reduces storage needs and workflow complexity – a smart choice for efficient lab work Extended Options Hot Start PCR: Available as Hot-Start version (ReproHot Proofreading Polymerase, M3012) : increased specificity, room-temperature setup, and immediate start without activation – ideal for sensitive reactions. With ReproFast DNA Polymerase, you have a universal enzyme covering a wide range of applications – powerful, reliable, and efficient. More High-Fidelity Proofreading Polymerases from Genaxxon bioscience: - M3002 Pwo Proofreading Polymerase- M3004 Pfu Proofreading Polymerase- M3012 ReproHot (KOD) Proofreading Polymerase- AQ97 High Fidelity proofreading Polymerase With our high quality dNTPs as Set (M3015) or Mix (M3016) or our DNA Ladders and our favourable standard agarose (M3044) we can offer additional products for your PCR.

The High-Fidelity Pfu proofreading DNA polymerase from Genaxxon bioscience is a thermostable, highly processive enzyme possessing 5'-3' DNA polymerase with additional 3'-5' proofreading exonuclease activity, which enables the correction of nucleotide incorporation errors. It has no 5'→3' exonuclease activity. The High-Fidelity Pfu DNA polymerase is a recombinant form of the hyperthermophilic archaebacteria Pyrococcus furiosus (Pfu). Pfu proofreading DNA polymerase shows an increased thermostability and a 10-times higher accuracy compared to Taq DNA polymerase. A mixture of Taq DNA Polymerase and Pfu DNA polymerase provides more robust synthesis of longer amplification products (Barnes, 1994. Proc. Natl. Acad. Sci. USA 91:2216-2220). Test sample available at a special price! The test sample price will be refunded on the first official order of the product. Features: 10-times higher accuracy compared to Taq DNA polymerase High-Fidelity polymerase Proofreading function (3' - 5' exonuclease activity) High thermo stability Generates blunt-end PCR products Generates PCR products for cloning and expression More High-Fidelity Proofreading Polymerases from Genaxxon bioscience:- M3003 ReproFast Proofreading Polymerase- M3002 Pwo Proofreading Polymerase- M3012 ReproHot (KOD) Proofreading Polymerase - AQ97 High Fidelity proofreading Polymerase With our high quality dNTPs as Set (M3015) or Mix (M3016) or our DNA Ladders and our favourable standard agarose (M3044) we can offer additional products for your PCR.

High-quality Taq DNA Polymerase from Genaxxon is a highly processive 5' - 3' DNA Polymerase, lacking 3' - 5' exonuclease activity. The high processivity and fidelity of Genaxxon bioscience Taq Polymerase allows amplification of DNA fragment of up to >7 kb. Genaxxon bioscience Taq Polymerase is delivered with 10X reaction buffer and separate MgCL2.The enzyme is delivered with our buffer component 'Buffer-S'. The buffer is optimised for high specificity amplification of DNA-templates. Our complete buffer contains 15mM MgCl2. Free Taq DNA Polymerase test sample available!No shipping costs within Germany. With our high quality dNTPs as Set (M3015.4100 and M3015.0250) or Mix (M3016.1010) or our DNA Ladders and our favourable standard agarose (M3044) we can offer additional high quality products for your PCR.

High-quality Taq DNA Polymerase from Genaxxon is a highly processive 5' - 3' DNA Polymerase, lacking 3' - 5' exonuclease activity. The high processivity and fidelity of Genaxxon bioscience Taq Polymerase allows amplification of DNA fragment of >7 kb. Genaxxon bioscience Taq Polymerase is delivered with 10X reaction buffer and separate MgCL2.The enzyme is delivered with our buffer component 'Buffer-E'. The buffer is optimised for high yield amplification of DNA-templates. Our complete buffer contains 25mM MgCl2. Taq DNA Polymerase test sample available! No shipping costs within Germany. With our high quality dNTPs as Set (M3015.4100 and M3015.0250) or Mix (M3016.1010) or our DNA Ladders and our favourable standard agarose (M3044) we can offer additional high quality products for your PCR.

SuperHot Taq DNA Polymerase is a superior DNA Polymerase for Real Time PCR, Hot-Start PCR, low-copy number PCR, or PCR of difficult templates. The enzyme is developed to enhance the specificity, sensitivity and yield of DNA amplification. SuperHot Taq DNA polymerase is a chemicaly modified form of thermostable DNA polymerase Taq, which is activated by heat treatment. A chemical moiety is attached to the enzyme at the active site, which renders the enzyme inactive at room temperature. Thus, during setup and before the first PCR cycle, the enzyme is not active and misprimed primers are not extended. As a result specifity and efficiency are increased by far compared to standard Taq DNA polymerase. Additionally, difficult targets with high GC-content can be amplified. For realtime PCR Genaxxon offers specially optimized 2-times qPCR mastermix >, respective a special Multiplex-PCR mastermix >. Features of Genaxxons SuperHot DNA Polymerase chemically modified increased specifity increased sensitivity for difficult templates with high GC-content Please have also a look on our SuperHot Taq 2-times Mastermix >. Even more convenient than the SuperHot Taq. With our high quality dNTPs as Set (M3015.4100 and M3015.0250) > or Mix (M3016.1010) > or our DNA Ladders > and favourable standard agarose (M3044) > we can offer additional products for your PCR.

Genaxxon's FAST DNA polymerase is a robust enzyme, ideally suited for applications like genotyping and screening, amplifying with extreme speed of up to 2 seconds per 1kb, as well as high yield and consistency. FAST DNA polymerase has 5’-3’ exonuclease activity, but no 3’-5’ exonuclease (proofreading) activity. PCR products generated with this enzyme are A-tailed and can thus be cloned into TA cloning vectors. The extreme speed of Genaxxons' FAST DNA polymerase allows the use of an extension rate of 4-8 kb/min. Genaxxon's FAST DNA Polymerase is delivered with 5X reaction buffer already containing MgCL2, dNTPs, enhancers and stabilizers for an optimal PCR result. Genaxxon's FAST DNA polymerase is also available as a convenient 2x master mix (M3286), including an inert red tracking dye for electrophoresis, containing all required components for fast PCR, except specific primers. Example of a typical PCR protocol (graphical presentation)* Ta is the annealing temperature (Ta should be 2°C above Tm**)  ** Tm is the melting temperature of a primer which is defined as the temperature at which 50% of the primer bind to the complementary sequence of the target DNA. Please note: By increasing Ta above Tm, this percentage decreases, however, primers will still anneal up to a certain degree and initiate extension. Therefore, PCR would still work with a Ta of several degrees higher than the Tm but with a dramatically reduced efficiency. Hence, we recommend optimizing the Ta by performing a temperature gradient (e.g., starting at the lowest Tm or a few degrees below and increasing with 2°C increments). *** Largest tested amplicon size: 5kb. Please note that longer extension times may be required for targets larger than 5kb!

Especially suitable for use in microbiology, the DNA free Taq Polymerase DF Taq S for high specificity is virtually free of foreign DNA. The special purification procedure guarantees DF Taq Polymerase free of any DNA impurities, especially free of DNA from the conservative region of the 16S ribosomal gene. DNA free DF Taq DNA Polymerase from Genaxxon is a highly processive 5' - 3' DNA Polymerase, lacking 3' - 5' exonuclease activity. The high processivity and fidelity of Genaxxon bioscience DNA free Taq Polymerase allows amplification of DNA fragments >7 kb. Genaxxon bioscience Taq Polymerase is delivered with 10X reaction buffer S and separate MgCl2. The buffer is optimised for high specificity amplification of DNA-templates. Our complete buffer S contains 15mM MgCl2. With our high quality dNTPs as Set (M3015.4100 and M3015.0250) or Mix (M3016.1010) or our DNA Ladders and our favourable standard agarose (M3044) we can offer additional high quality products for your PCR.

Taq DNA Hot Start Polymerase with Aptamer technology. The specific aptamer inhibits DNA polymerase activity below temperatures of 50°C to 55°C, thus enabling hot start PCR. At higher temperatures, the aptamer falls off the active site of Taq polymerase. The Hot Start Taq Polymerase can be used like a normal Taq without having to lengthen the first denaturation step.The binding of aptamer and polymerase prevents the formation of primer-dimer constructs and other artefacts revealed by non-specific primer binding. Check also our SuperHot Mastermix with or without dye. With our high quality dNTPs as Set or Mix or our DNA Ladders and our favourable standard agarose we can offer additional products for your PCR.

AQ97 High Fidelity DNA Polymerase ist eine proofreading Polymerase mit hoher Amplifikationsgeschwindigkeit und sehr hoher Genauigkeit (>60-fach) gegenüber Taq DNA Polymerase. AQ97 ist ideal für schwierige Targets mit sehr niedrigem oder hohem GC-Gehalt geeignet. Neben diesen Eigenschaften zeichnet sich die AQ97 auch durch Ihre Eignung, lange Targets bis zu 18 kb, zu amplifizieren aus. Dies führt zu sehr genauen und zuverlässigen PCR-Ergebnissen.  AQ97 High Fidelity DNA Polymerase besitzt sowohl eine 5'→3' DNA Polymerase-Aktivität als auch eine 3'→5' proofreading Exonuklease-Aktivität, die es dieser Polymerase ermöglicht, Basenpaar-Fehlpaarungen zu korrigieren. AQ97 High Fidelity DNA Polymerase ist ein fusionierter Proteinkomplex aus DNA-Polymerase mit einer prozessivitätssteigernden DNA-Bindungsdomäne. Neben einer sehr schnellen und robusten Amplifikation komplexer und langer Zielmoleküle zeichnet sich die AQ97 High Fidelity DNA Polymerase durch eine hohe Genauigkeit aus, was dazu führt, dass auch für lange Amplicons eine genaue Amplifikation gewährleistet ist. Dadurch eignet sich die AQ97 besonders für PCR-Experimente, die eine Amplifikation mit sehr niedrigen Fehlerraten erfordern, wie z. B. Klonen/Subklonen, NGS-Anwendungen, SNP-Analysen und Mutagenese. Eigenschaften: • High Fidelity: >60x Taq fidelity• High elongation rate: 10 sec/kb (bis 6000bp pro Minute)• Long range amplification: 18 kb for human gDNA• 5'-3' polymerase acitivity and 3'-5' exonuclease activity.• generates blund ends.Picture 1: Comparison figures of fidelity values for AQ97 DNA Polymerase, AccuPol DNA Polymerase, two well-recognized high fidelity DNA polymerases P and Q and Taq DNA Polymerase were determined through NGS-based analysis of nucleotide misincorporation during PCR. Initially, PCR amplification was performed on a ~ 200 bp synthetic DNA target, generating PCR products for each of the tested polymerases (using recommended setup conditions).  Picture 2: Distribution of substitution errors. PCR was performed using Taq DNA Polymerase, AQ97 High Fidelity DNA Polymerase, high fidelity DNA Polymerase Q and high fidelity DNA Polymerase P. The PCR was followed by NGS sequencing of the PCR products. The number of substitutions at each PCR target position was calculated and plotted in diagram A. Substitutions include misincorporated nucleotides and deletions at each position. Non-polymerase errors are subtracted from the total number of errors to revel true polymerase errors. Non-polymerase errors include mutations caused by thermocycling-induced DNA damage, pre-NGS sample preparation and sequencing errors. In these diagrams the average number of substitutions for Taq DNA Polymerase (Taq average) and for AQ97 High Fidelity DNA Polymerase (AQ97 average) is also plotted. Diagram B magnifies the area near the detection limit, displaying more information about the number of substitutions for AQ97 High Fidelity DNA Polymerase, high fidelity DNA Polymerase Q and high fidelity DNA polymerase P. Click to enlarge diagrams.  

AQ97 Hot Start High Fidelity DNA Polymerase ist eine proofreading Polymerase mit hoher Amplifikationsgeschwindigkeit und sehr hoher Genauigkeit (>60-fach) gegenüber Taq DNA Polymerase. AQ97 ist ideal für schwierige Targets mit sehr niedrigem oder hohem GC-Gehalt geeignet. Neben diesen Eigenschaften zeichnet sich die AQ97 auch durch Ihre Eignung, lange Targets bis zu 18 kb, zu amplifizieren aus. Dies führt zu sehr genauen und zuverlässigen PCR-Ergebnissen.  AQ97 High Fidelity DNA Polymerase besitzt sowohl eine 5'→3' DNA Polymerase-Aktivität als auch eine 3'→5' proofreading Exonuklease-Aktivität, die es dieser Polymerase ermöglicht, Basenpaar-Fehlpaarungen zu korrigieren. AQ97 High Fidelity DNA Polymerase ist ein fusionierter Proteinkomplex aus DNA-Polymerase mit einer prozessivitätssteigernden DNA-Bindungsdomäne. Neben einer sehr schnellen und robusten Amplifikation komplexer und langer Zielmoleküle zeichnet sich die AQ97 High Fidelity DNA Polymerase durch eine hohe Genauigkeit aus, was dazu führt, dass auch für lange Amplicons eine genaue Amplifikation gewährleistet ist. Dadurch eignet sich die AQ97 besonders für PCR-Experimente, die eine Amplifikation mit sehr niedrigen Fehlerraten erfordern, wie z. B. Klonen/Subklonen, NGS-Anwendungen, SNP-Analysen und Mutagenese. Eigenschaften: • High Fidelity: >60x Taq fidelity• High elongation rate: 10 sec/kb (bis 6000bp per minute) • Built in hot start technology• Long range amplification: 18 kb for human gDNA• 5'-3' polymerase acitivity and 3'-5' exonuclease activity.• generates blund ends. Bild 1: Die Vergleichswerte der Genauigkeitswerte für die AQ97-DNA-Polymerase, die AccuPol-DNA-Polymerase, die beiden Wettbewerber High-Fidelity-DNA-Polymerasen "P" und "Q" und die Taq-DNA-Polymerase wurden durch eine NGS-basierte Analyse des Nukleotid-Fehleinbaus während der PCR ermittelt. Die PCR-Amplifikation wurde mit einem synthetischen DNA-Target von etwa 200 bp durchgeführt, wobei für jede der getesteten Polymerasen PCR-Produkte erzeugt wurden (unter Verwendung der empfohlenen Setup-Bedingungen). Bild 2: Acht verschiedene humane genomische DNA-Targets mit einer Länge von 400 bis 800 bp und einem GC-Gehalt zwischen 29% und 78% wurden mit dem AQ97 Hot Start High Fidelity DNA Polymerase 2x Master Mix amplifiziert. Für alle Targets wurde eine robuste Amplifikation beobachtet. Für Targets mit einem GC-Gehalt von mehr als 70% wurde der Reaktionsmischung 2 M Betain Enhancer Solution zugesetzt. Marker: M.  Bild 3: Es wurden fünf verschiedene Targets aus menschlicher genomischer DNA mit einer Länge von 2 kb bis 17,5 kb amplifiziert. Für alle Targets wurde eine robuste Amplifikation mit dem AQ97 Hot Start High Fidelity DNA Polymerase 2x Master Mix beobachtet, was die Fähigkeit des Master Mixes zur Amplifikation großer und komplexer Targets belegt. Die Amplikongrößen sind am oberen Rand des Gels angegeben. Marker M: High Range DNA-Leiter.

Genaxxon’s FAST HotStart DNA polymerase is a robust enzyme with an antibody mediated hotstart function, ideally suited for applications like genotyping and screening, amplifying with extreme speed of up to 2 seconds per 1kb, as well as high yield and consistency. PCR products generated with this enzyme are A-tailed and can thus be cloned into TA cloning vectors. The extreme speed of Genaxxon’s FAST HotStart DNA polymerase allows the use of an extension rate of 4-8 kb/min. The buffer composition has been optimized via high-throughput screening and makes the enzyme particularly resistant to PCR inhibitors. Therefore, Genaxxon’s FAST HotStart DNA polymerase is suitable for direct PCR from unpurified samples, including blood, urine, and bacterial colonies, making this the ideal choice for consistent results in fast complex PCR amplifications. Genaxxon's FAST HotStart DNA polymerase is also available as a convenient 2x master mix (M3288), including an inert red tracking dye for electrophoresis, containing all required components for fast PCR, except specific primers. Example of a typical PCR protocol (graphical presentation)* Ta is the annealing temperature (Ta should be 2°C above Tm**)  ** Tm is the melting temperature of a primer which is defined as the temperature at which 50% of the primer bind to the complementary sequence of the target DNA. Please note: By increasing Ta above Tm, this percentage decreases, however, primers will still anneal up to a certain degree and initiate extension. Therefore, PCR would still work with a Ta of several degrees higher than the Tm but with a dramatically reduced efficiency. Hence, we recommend optimizing the Ta by performing a temperature gradient (e.g., starting at the lowest Tm or a few degrees below and increasing with 2°C increments). *** Largest tested amplicon size: 5kb. Please note that longer extension times may be required for targets larger than 5kb!

SNP Pol DNA polymerase for easy, reliable and rapid allele-specific discrimination, eg. CRISPR / Cas9 point mutations, for detecting incorrect CRISPR / Cas9 products, or validating sequencing results. The SNP Pol DNA polymerase distinguishes highly specific, whether a mismatch of the primer-template-complex is present or not. The mismatch (point mutation) must be at the 3 'end of the primer. Thus, mutant alleles can be distinguished exactly from wild-type alleles - without sequencing, since the polymerase simply does not amplify in the case of a mismatch. Just place your primer on the supposed point mutation (Important: The point mutation must be at the 3 'end) and the polymerase will detect a mismatch in this region with almost 100% accuracy: If the template base complementary to the 3' end of the primer shows the mutation and the primer does not, no amplification takes place - 100% certainty within a short time!The SNP Pol DNA polymerase can therefore be easily, time and cost-effectively used for the screening of point mutations. The variant SNP PolTaq DNA polymerase > has 5'-3 'nuclease activity and can therefore be used for specific hydrolysi probes such as Taqman® probes or Molecular beacons. For more information on our SNP DNA polymerases and applications, read our blog now. DescriptionSNP Pol DNA Polymerase is a highly selective DNA polymerase for the detection of Single Nucleotide Polymorphisms. It was developed specifically for allele-specific discrimination, where a very high discrimination rate is required e.g., in allele-specific PCR (ASA; AS-PCR), allele-specific primer extension (AS-PEX), SNP analysis, genotyping or in methylation-specific PCRs (MSP). Many other DNA polymerases tolerate mismatched primer-template complexes and are therefor not suitable. The SNP Pol DNA polymerase, on the other hand, distinguishes these specifically (high discrimination) and supplies only PCR products with perfectly matching primer pairs! The Genaxxon SNP Pol and SNP PolTaq DNA polymerases differ up to 100% by means of allele-specific PCR between the two alleles and, after a simple qPCR, give a clear result as to which allele is present. Thus, the principly great potential of the CRISPR/Cas9 technology can be used for human medicine and plant biotechnology especially together with the SNP PolTaq or SNP Pol DNA polymerase. Allele-specific PCR can be used to quantify the mutation rate in a pool or background of wild-type sequences. The verification of mutation frequencies determined by NGS can also be verified by means of allele-specific PCR and SNP Pol DNA polymerase. The SNP PolTaq DNA polymerase is very suitable for the analysis of liquid biopsy samples. With SNP PolTaq, the presence and frequency of cancer mutations can be analyzed and quantified very well. Picture below: Application note SNP Pol DNA Polymerase   We recommend short amplicon length (about 60-200 bp) for best results, but also longer amplicon lengths are possible. The addition of additional Magnesium (+0.5 - 1.5mM) might be needed in case of longer amplicons >500 bp.Trouble shooting: No bands after PCR of 30 cycles! Optimisation procedure for missing or weak bands.Annealing temperature is too low! Attention: The SNP Pol is an aptamer inhibited DNA polymerase. The aptamer oligo used reversibly inhibits the polymerase at temperatures <55°C! Therefore, the primers should ideally have an annealing temperature of >57°C. - realtime PCR approach - Check the dNTP concentration. This should be between 200 and 300µM (in the PCR). - Increase the number of cycles (at least 35, preferably equal to 40) Test optimisation - number of cyclesIn endpoint detection, a cycle count of 30 is not always sufficient to generate a clearly visible band. For example, with less than 200 DNA copies as a starting value. The test should therefore be run using real-time PCR, or five parallel PCRs should be used, one of which is taken from the PCR device after each of five different cycles (example below). Endpoint detection: Set up five identical PCR reactions in parallel using the SNP Pol DNA polymerase. Remove one of the PCR tubes from the cycler at each of 20, 25, 30, 35 and 40 cycles and apply all five reactions to an agarose gel at the end. This makes it easy to recognise which is the optimum number of cycles in the PCR for the given application (starting material, target, primer). SNP Pol PCR with cell lysates Animal cells and E.coli can be used directly for PCR with the SNP Pol DNA polymerase! No separate lysis or proteinase K digestion is necessary. PCR procedure:1. 50 - 500 cells are required per PCR batch! 2. prepare PCR mix with primers and buffer and place on ice! 3. add the picked colony directly to 10-20µL water (no separate lysis and no proteinase K digestion), vortex briefly (5 seconds), then add this cell suspension directly to the PCR reaction mixture, e.g. 10µL cell suspension for a PCR preparation with a total volume of 20µL. An initial denaturation time of 2-3 minutes is sufficient is sufficient, can be extended to 5 minutes if necessary. Genomic DNA per reaction max. 100 copies is relatively low, but should still work. The remainder of this cell suspension can also be frozen away in order to carry out further tests. Optional: 4. if possible: do real-time PCR!The SNP Pol DNA polymerase (M3009 > or M3061 >) can be used together with non-specific fluorescent dyes (eg Genaxxon's Green DNA Dye > or SybrGreen®) in real-time PCR. When working with specific PCR probes, only the SNP PolTaq DNA polymerase can be used, since only these have a 5'-3 'exonuclease activity. With our high quality dNTPs as Set (M3015.4100) > or Mix (M3016.1010) > or our DNALadders > and our favourable standard agarose (M3044) > we can offer additional products for your PCR. Application areas for SNP Pol DNA and SNP Pol DNA polymerase- Monitoring, verification and detection of point mutations- Identification of correct or wrong CRISPR/Cas9 products- Verification/validation of sequencing results- Quantification of mutations (e.g. NGS results)- SNP-detection by allele-specific amplification (ASA) / Allele-specific PCR- Methylation specific PCRs (MSP) after bisulfite treated DNA (CpG methylation sides)- HLA genotyping- micro sequencing- realtime PCR with hydrolysis probes- realtime multiplex PCRs - DamID-seq data in C. elegans. As an alternative to ChIP, DNA adenine methyltransferase identification by sequencing (DamID-seq) was recently shown to be able to characterize binding sites in single mammalian cells. Additionally, DamID can be achieved for cell-type specific analysis by expressing Dam fusion proteins under tissue specific promoters in a controlled manner. In this report, we present a user-friendly pipeline to analyse DamID-seq data in C. elegans. Sharma R, Ritler D, Meister P.Tools for DNA adenine methyltransferase identification analysis of nuclear organization during C. elegans development. Genesis. 2016 Feb 4. doi: 10.1002/dvg.22925. - Minisequencing SNP genotyping with SNPase DNA Polymerase can be carried out by the procedure described in: Lovmar L, Fredriksson M, Liljedahl U, Sigurdsson S, Syvänen AC. Quantitative evaluation by minisequencing and microarrays reveals accurate multiplexed SNP genotyping of whole genome amplified DNA. Nucleic Acids Res. 2003;31:e129. - Allel specific mismatch selectivity by the HiDi DNA polymerase Drum M, Kranaster R, Ewald C, Blasczyk R, Marx A (2014) Variants of a Thermus aquaticus DNA Polymerase with Increased Selectivity for Applications in Allele- and Methylation-Specific Amplification. PLoS ONE 9(5): e96640. doi:10.1371/journal.pone.0096640

SNP Pol DNA polymerase for easy, reliable and rapid allele-specific discrimination, eg. CRISPR / Cas9 point mutations, for detecting incorrect CRISPR / Cas9 products, or validating sequencing results. The SNP Pol DNA polymerase distinguishes highly specific, whether a mismatch of the primer template complex is present or not. The mismatch (point mutation) must be at the 3 'end of the primer. Thus, mutant alleles can be distinguished exactly from wild-type alleles - without sequencing since the polymerase simply does not amplify in the case of a mismatch. Just place your primer on the supposed point mutation (Important: The point mutation must be at the 3 'end) and the polymerase will detect a mismatch in this region with almost 100% accuracy: If the template base complementary to the 3' end of the primer shows the mutation and the primer does not, no amplification takes place - 100% certainty within a short time!The SNP Pol DNA polymerase can therefore be easily, time and cost-effectively used for the screening of point mutations. SNP PolTaq DNA polymerase shows also 5'-3'-nuclease activity and is therefore suitable for hydrolysis probe-based assays (Taqman®, molecular beacons, etc.). For more information on our SNP DNA polymerases and applications, read our blog now. DescriptionSNP PolTaq DNA Polymerase is a highly selective DNA polymerase for the detection of Single Nucleotide Polymorphisms. It was developed specifically for allele-specific discrimination, where a very high discrimination rate is required e.g., in allele-specific PCR (ASA; AS-PCR), allele-specific primer extension (AS-PEX), SNP analysis, genotyping or in methylation-specific PCRs (MSP). Many other DNA polymerases tolerate mismatched primer-template complexes and are therefor not suitable. The SNP Pol DNA polymerase, on the other hand, distinguishes these specifically (high discrimination) and supplies only PCR products with perfectly matching primer pairs! The Genaxxon SNP Pol and SNP PolTaq DNA polymerases differ up to 100% by means of allele-specific PCR between the two alleles and, after a simple qPCR, give a clear result as to which allele is present. Thus, the principle great potential of the CRISPR/Cas9 technology can be used for human medicine and plant biotechnology especially together with the SNP PolTaq or SNP Pol DNA polymerase. Allele-specific PCR can be used to quantify the mutation rate in a pool or background of wild-type sequences. The verification of mutation frequencies determined by NGS can also be verified by means of allele-specific PCR and SNP Pol DNA polymerase. The SNP PolTaq DNA polymerase is very suitable for the analysis of liquid biopsy samples. With SNP PolTaq , the presence and frequency of cancer mutations can be analyzed and quantified very well. Picture below: Application note SNP PolTaq DNA Polymerase   We recommend short amplicon length (about 60-200 bp) for best results, but also longer amplicon lengths are possible. The addition of additional Magnesium (+0.5 - 1.5mM) might be needed in case of longer amplicons >500 bp.Trouble shooting: No bands after PCR of 30 cycles! Optimisation procedure for missing or weak bands.Annealing temperature is too low! Attention: The SNP PolTaq is an aptamer inhibited DNA polymerase. The aptamer oligo used reversibly inhibits the polymerase at temperatures <55°C! Therefore, the primers should ideally have an annealing temperature of >57°C. - realtime PCR approach - Check the dNTP concentration. This should be between 200 and 300µM (in the PCR). - Increase the number of cycles (at least 35, preferably equal to 40) Test optimisation - number of cyclesIn endpoint detection, a cycle count of 30 is not always sufficient to generate a clearly visible band. For example, with less than 200 DNA copies as a starting value. The test should therefore be run using real-time PCR, or five parallel PCRs should be used, one of which is taken from the PCR device after each of five different cycles (example below). Endpoint detection: Set up five identical PCR reactions in parallel using the SNP Pol DNA polymerase. Remove one of the PCR tubes from the cycler at each of 20, 25, 30, 35 and 40 cycles and apply all five reactions to an agarose gel at the end. This makes it easy to recognise which is the optimum number of cycles in the PCR for the given application (starting material, target, primer). SNP Pol PCR with cell lysates Animal cells and E.coli can be used directly for PCR with the SNP Pol DNA polymerase! No separate lysis or proteinase K digestion is necessary. PCR procedure:1. 50 - 500 cells are required per PCR batch! 2. prepare PCR mix with primers and buffer and place on ice! 3. add the picked colony directly to 10-20µL water (no separate lysis and no proteinase K digestion), vortex briefly (5 seconds), then add this cell suspension directly to the PCR reaction mixture, e.g. 10µL cell suspension for a PCR preparation with a total volume of 20µL. An initial denaturation time of 2-3 minutes is sufficient is sufficient, can be extended to 5 minutes if necessary. Genomic DNA per reaction max. 100 copies is relatively low, but should still work. The remainder of this cell suspension can also be frozen away in order to carry out further tests. Optional: 4. if possible: do real-time PCR!The SNP Pol DNA polymerase (M3009 > or M3061 >) can be used together with non-specific fluorescent dyes (eg Genaxxon's Green DNA Dye > or SybrGreen®) in real-time PCR. When working with specific PCR probes, only the SNP PolTaq DNA polymerase can be used, since only these have a 5'-3 'exonuclease activity. With our high quality dNTPs as Set (M3015.4100) > or Mix (M3016.1010) > or our DNALadders > and our favourable standard agarose (M3044) > we can offer additional products for your PCR. Application areas for SNP Pol DNA and SNP PolTaq DNA polymerase- Monitoring, verification and detection of point mutations- Identification of correct or wrong CRISPR/Cas9 products- Verification/validation of sequencing results- Quantification of mutations (e.g. NGS results)- SNP-detection by allele-specific amplification (ASA) / Allele-specific PCR- Methylation specific PCRs (MSP) after bisulfite treated DNA (CpG methylation sides)- HLA genotyping- micro sequencing- realtime PCR with hydrolysis probes- realtime multiplex PCRs - DamID-seq data in C. elegans. As an alternative to ChIP, DNA adenine methyltransferase identification by sequencing (DamID-seq) was recently shown to be able to characterize binding sites in single mammalian cells. Additionally, DamID can be achieved for cell-type specific analysis by expressing Dam fusion proteins under tissue specific promoters in a controlled manner. In this report, we present a user-friendly pipeline to analyse DamID-seq data in C. elegans. Sharma R, Ritler D, Meister P.Tools for DNA adenine methyltransferase identification analysis of nuclear organization during C. elegans development. Genesis. 2016 Feb 4. doi: 10.1002/dvg.22925. - Minisequencing SNP genotyping with SNPase DNA Polymerase can be carried out by the procedure described in: Lovmar L, Fredriksson M, Liljedahl U, Sigurdsson S, Syvänen AC.bQuantitative evaluation by minisequencing and microarrays reveals accurate multiplexed SNP genotyping of whole genome amplified DNA. Nucleic Acids Res. 2003;31:e129. - Allel specific mismatch selectivity by the HiDi DNA polymerase Drum M, Kranaster R, Ewald C, Blasczyk R, Marx A (2014) Variants of a Thermus aquaticus DNA Polymerase with Increased Selectivity for Applications in Allele- and Methylation-Specific Amplification. PLoS ONE 9(5): e96640. doi:10.1371/journal.pone.0096640