GoTaq Amplification Maximum performance, flexibility, control and convenience

Transcription

1 GoTaq Amplification Maximum performance, flexibility, control and convenience GoTaq products and formulations designed to improve your amplification needs

2 Reverse Transcription GoScript Reverse Transcriptase 3 The Gold Standard M-MLV Reverse Transcriptase, RNase H Minus, Point Mutant 4 Cost effective, quality RT Real-Time PCR How to choose your system 5 GoTaq 1-Step RT-qPCR 6 Brighter signal for earlier quantification GoTaq 2-Step RT-qPCR 7 Ultra active, ultra bright GoTaq qpcr Master Mix 8 Higher sensitivity for detecting a single copy gene GoTaq Probe qpcr and RT-qPCR Master Mixes 9-11 The reliability and performance of GoTaq in a truly affordable probe format End Point PCR Hot Start GoTaq G2 Flexi Hot Start Polymerase 12 Higher specificity, greater sensitivity GoTaq G2 Hot Start Green Master Mix 12 GoTaq Hot Start with the convenience of a tracking dye GoTaq G2 Hot Start Colourless Master Mix 12 GoTaq Hot Start with the convenience of a loading buffer End Point PCR Standard PCR GoTaq G2 Flexi DNA Polymerase 13 Flexi DNA Polymerase for performance optimisation GoTaq G2 DNA Polymerase 13 Robust, enhanced, reliable amplification GoTaq Green Master Mix 13 Pre-mixed, ready-to-use with tracking dye GoTaq Colourless Master Mix 13 Pre-mixed, ready-to-use with loading buffer Special PCR GoTaq Long PCR Master Mix 14 Proven, robust amplification of GoTaq Polymerase in long range PCR (up to 30kb) Pfu DNA Polymerase 14 High fidelity, lowest error rate of any thermostable DNA Polymerase Related Products QuantiFluor dsdna, ssdna & RNA dyes 15 Fluorometers 15 More sensitive and economical Diamond Nucleic Acid Dye 16 PCR Cloning, dntps, RNasin Plus RNase Inhibitor 16 CXR Reference Dye 16 Reverse Transcription GoScript Reverse Transcriptase Specifically optimised for RT-qPCR ensuring robust, reliable cdna synthesis even with the most difficult templates. Detect both rare and abundant transcripts with ease Transcribe long mrnas Work with RNA with complex secondary structure, e.g. G-C rich Nocturnin Synthesise cdna in the presence of inhibitors, e.g % ethanol Save money compared to leading RTs GoScript Reverse Transcription System GoScript Reverse Transcriptase ** ** GoScript Reverse Transcriptase utilises M-MLV and state-of-the-art buffer technology designed for qpcr to deliver robust, reliable cdna synthesis of a full range of rare and abundant transcripts, even in the presence of inhibitors. 50 reactions 500 reactions A5000 A5001 A5003 A The GOLD Standard in reverse transcription Reverse Transcription 2 3

3 Reverse Transcription M-MLV Reverse Transcriptase, RNase H Minus, Point Mutant Ideal for the preparation of full-length cdna from long RNA templates and libraries containing a high percentage of full-length cdna Thermostability of this point mutant prevents problems associated with secondary structure More tolerance to variations in enzyme and substrate concentration means improved consistency in performance Real-Time PCR Real-Time PCR - how to choose your system Product Details Technology Dye-Based Detection GoTaq Dye-Based qpcr and RT-qPCR Uses dsdna binding dye, BRYT Green Dye, to detect PCR product as it accumulates during PCR, SYBR Green alternative Probe-Based Detection GoTaq Probe-Based qpcr and RT-qPCR Uses hydrolosis probes specific to target gene to detect product as it accumulates during PCR Specificity Medium High Sensitivity-Low Copies Variable 1-10 copies Reproductibility Medium High Multiplexing Melt-Curve For Specificity Yes No Cost M-MLV Reverse Transcriptase, RNase H Minus, Point Mutant Thermal stability and synthesis of large transcripts. cdnas were synthesised from 1µg of total human RNA using M-MLV (H ), Point Mutant, (Panel A) or SuperScript II (Panel B) reverse transcriptase at 37 C, 42 C and 50 C. GoTaq Long PCR amplifications were performed on each transcription reaction using five primer sets, amplifying products from 9.4kb to 1.5kb. Ten microlitres of the 9.4, 6.9, and 5.2kb PCR products and 1µl of the 3.2 and 1.5kb PCR products were separated on a 1% agarose gel. Lane M. BenchTop 1kb DNA Ladder (Cat.No. G7541). 2,500u 10,000u 50,000u M3681 M3682 M Applications Select 1-Step if: Gene Expression, DNA Quantification, CHiP Selecting the most suitable format You do not need to store cdna You will dispose of samples after a few uses You have many samples with one or few targets You are using a liquid-handling robot 1-Step Advantages Reduced chance of cross-contamination during the procedure You will dispose of sample after a few uses Faster results Disadvantages Increased risk of primer-dimers cdna is not available for other assays Select 2-Step if: Gene Expression, DNA Quantification, CHiP SNP Genotyping, Copy Number Variation, Pathway Analysis, Mutation Detection, Protein Analysis, Multiplexing You need to store the cdna You have a limited amount of sample You are assaying many targets per sample 2-Step Advantages Optimised performance of both RT and PCR steps cdna is available for other procedures Disadvantages RT enzymes and buffers can inhibit real-time PCR Less convenient, more time consuming Contamination risk is higher than 1-Step method 1-Step or 2-Step formats? 4 5

4 Brighter signal for earlier quantification of low and high copy RNA targets in a convenient 1-Step RT-qPCR format. GoTaq 2-Step RT-qPCR kit delivers optimised, highly sensitive quantification of a full range of RNA targets, combining GoScript Reverse Transcriptase with the ultra bright fluorescence of GoTaq qpcr Master Mix. High efficiency full length cdna synthesis Sensitive detection of rare to abundant RNA targets Linear quantification over a wide range of samples 2.00 Contains BRYT Green Dye GoTaq 2-Step RT-qPCR system offers the flexibility of a 2-step format with optimised and proven components, which combined with the recommended protocol, enable high efficiency, full length cdna synthesis via the GoScript RT and the separate GoTaq qpcr module. This enables multiple amplifications for different targets to be undertaken from a single cdna preparation if desired, unlike 1-Step RT-qPCR systems. Sensitive quantification of GAPDH across an extended linear range of total RNA input with detection down to 10 fg. Product GoTaq 1-Step RT-qPCR System Size Cat.No. Price 200 reactions (50 µl) A GoTaq 2-Step RT-qPCR provides sensitive detection down to 1fg across an extended linear range of RNA input. Product GoTaq 2-Step RT-qPCR System SYBR GreenER Two-Step qrt-pcr Kit Universal ( ) Robust activity in the presence of inhibitors 3.00 SYBR GreenER Two-Step qrt-pcr Kit for ABI PRISM ( ) 1-Step reverse transcription and real-time quantitative PCR (RT-qPCR) in a single tube format. Combines BRYT Green Dye and optimised RT buffers. Price/Reac on 4.00 SuperScript III Pla num Two-Step qrt-pcr Kit with SYBR Green ( ) Compatible with all real-time PCR instruments in both fast and standard cycling conditions Room temperature set-up makes it suitable for automation SuperScript III Pla num SYBR Green Two-Step qrt-pcr Kit w/rox ( ) BRYT Green Fluorescent Dye, a novel dsdna binding dye and optimised buffer formulation improve data accuracy and sensitivity of low expressing targets or targets present in low amounts GoTaq 2-Step RT-qPCR GoTaq 2-Step RT-qPCR System (A6010) Ultra active cdna synthesis combined with ultra bright qpcr detection GoTaq 1-Step RT-qPCR Size Cat.No. Price 50 RT reactions x 50 µl or x 25 µl PCR reactions A

5 GoTaq qpcr Master Mix GoTaq Probe qpcr Real-time PCR Systems Higher sensitivity, superior stability BRYT Green Dye gives higher sensitivity and earlier detection Early Cq and detection of low copy number targets Superior dye-based real-time PCR in a 2x mix GoTaq qpcr Master Mix s proprietary, fluorescent DNA binding dye exhibits greater fluorescence than SYBR Green I. Combined with GoTaq Hot Start and an optimised buffer, it provides robust, real-time PCR with increased reliability, reproducibility and sensitivity. GoTaq qpcr Master Mix is a pre-mixed, ready-touse solution containing all the necessary components formulated to achieve superior dye-based, real-time PCR in a 2x mix. Enhanced stability for room-temperature set-up Direct substitute for SYBR Green I Robust reliable performance of GoTaq Hot Start Six commercially available dye-based master mixes were compared to GoTaq qpcr Master Mix amplifying the GAPDH gene from 1ng of human DNA GoTaq qpcr Master Mix 200 reactions (50 µl) A GoTaq qpcr Master Mix 1,000 reactions (50 µl) A The reliability and performance of GoTaq in a truly affordable probe format! Compatible with any real-time PCR instrument, both fast and standard cycling methods and with Taqman and other probe assays such as molecular beacons Sensitive detection on any real-time instrument Ready-to-use, stabilised 2x master mix that simplifies the preparation of reactions for qpcr using hydrolysis probe detection. Optimised for quantitative PCR assays, and designed for sensitive sequence detection and quantification, gene expression analysis, detection of sequence variants and fluorescent detection. This master mix employs rapid hot-start activation and processive enzymes. The master mix does not contain a reference dye; however, a separate tube of carboxy-x-rhodamine (CXR) reference dye is included with these systems, allowing users to add reference dye to amplification reactions if desired. Exceptional room-temperature set-up makes it suitable for automation and high-throughput detection High-efficiency, full-length cdna synthesis in the presence of inhibitors GoTaq Probe 2-Step RT-qPCR System. cdna was generated from 100ng of human pancreas total RNA, using the GoScript Reverse Transcription System. The human insulin gene (INS) was detected from five-fold serial dilutions of cdna (100ng to 6.4pg), using GoTaq Probe qpcr Master Mix for amplification. The resulting standard curve is shown in the inset (R2 = 0.998,slope = 3.33). 8 9

6 GoTaq Probe 1-Step RT-qPCR System GoTaq Probe qpcr Master Mix Real - Time PCR For 1-Step RT-qPCR from RNA templates For the detection and relative quantification of RNA expression levels using a one-step RT-qPCR method, this system combines GoScript Reverse Transcriptase and GoTaq Probe qpcr Master Mix in single-step real-time amplification reaction. The GoTaq Probe qpcr Master Mix is formulated with dutp. When dutp is incorporated into the amplification products, the amplicons are susceptible to degradation by uracil-dna glycosylase (UNG); this allows users to incorporate UNG into subsequent reactions for control of possible carryover contamination. Ready-to-use, stabilised 2x formulation with all of the components for qpcr, just add template, primers and probe. GoTaq Probe qpcr Master Mix is designed to provide resistance to a wide range of PCR inhibitors. Using antibody-mediated hot-start chemistry, reaction set-up can be performed at room- temperature. The hot-start activation and processive enzymes make it suitable for both standard and fast instrument cycling programmes. For qpcr from cdna templates GoTaq Probe qpcr Master Mix 200 reactions (20 µl) 1,000 reactions (20 µl) A6101 A GoTaq Probe 1-Step RT-qPCR System 200 reactions (20 µl) A UNG not included in the kit GoTaq Probe 2-Step RT-qPCR System For 2-step RT-qPCR from RNA templates Two-step RT-qPCR method utilising GoScript Reverse Transcription System. Optimised reaction buffer and reverse transcriptase designed to enable efficient synthesis of first-strand cdna in preparation for PCR amplification. The cdna product may be added directly to downstream qpcr amplification reactions. GoTaq Probe 2-Step RT-qPCR System 200 reactions (20 µl) A

7 End Point PCR Hot Start End Point PCR Standard PCR GoTaq G2 Flexi Hot Start Polymerase and Master Mixes GoTaq G2 DNA Polymerase End Point PCR Hot Start Next generation GoTaq New, optimised taq and buffers Prepare your reactions at room-temperature, not on ice Eliminate non-specific amplification with Hot Start technology Simplify reaction set-up and save time with a ready-to-use master mix Improve performance across many sample types with a PCR inhibitor resistant system Reliable amplification Performance optimisation GoTaq G2 Flexi DNA Polymerase allows you to optimise enzyme and magnesium concentrations in your PCR GoTaq G2 Flexi DNA Polymerase Separate MgCl 2 is supplied GoTaq G2 DNA Polymerase has the convenience of reaction buffers containing magnesium 100u 500u 2,500u 10,000u M7801 M7805 M7806 M Next generation performance for all your standard PCR needs End Point PCR Hot Start GoTaq G2 DNA Polymerase 100u 500u 2,500u M7841 M7845 M GoTaq G2 Hot Start Polymerase GoTaq G2 Hot Start Green Master Mix GoTaq G2 Hot Start Colourless Master Mix GoTaq G2 Hot Polymerase produces excellent specificity and higher yield with difficult targets. The 1.5kb fragment of the Corynephage omega gene, a hot-start model of specificity, was amplified from 500ng of plasmid DNA using GoTaq G2 Hot Start Polymerase with Green (G) and colourless (C) Buffers and the leading competitors (R, Q, L1, L2, L3). 100u 500u 2,500u 10,000u 1,000 reactions 1,000 reactions M7401 M7405 M7406 M7408 M7422 M7423 M7432 M , GoTaq Master Mixes Green or colourless buffers Pre-mixed and ready-to-use for standard PCR GoTaq Green Master Mix GoTaq Colourless Master Mix 1,000 reactions 1,000 reactions M7122 M7123 M7132 M

8 Special PCR Related Products GoTaq Long PCR Master Mix QuantiFluor dsdna, ssdna and RNA Dye Quantification Systems Allows increased elongation resulting in longer DNA amplification Amplifies up to 30kb of human genomic DNA and 40kb of lower complexity targets Proof-reading enzyme repairs DNA mismatches in the presence of a highly processive enzyme allowing increased DNA elongation resulting in longer DNA amplification Ideal for cloning genes, mutational analysis, DNA sequencing and next-gen sequencing PCR products varying in size from 5.2kb (α-1 antitrypsin) up to 30kb (β-globin) were amplified from 200ng of human genomic DNA using GoTaq Long PCR Master Mix. Very economical and compatible with most detection instruments. QuantiFluor dsdna Quantification System 1ml E QuantiFluor ssdna Quantification System 1ml E QuantiFluor RNA Quantification System 1ml E More sensitive and more economical than leading dyes Special PCR Ready-to-use blend of GoTaq Hot Start Polymerase with a proprietary, thermostable, proof-reading polymerase for enhanced processivity and proof reading capabilities. Amplification of targets up to 40kb in length from 2.5ng of Lambda DNA using GoTaq Long PCR Master Mix. GoTaq Long PCR Master Mix M Fluorometers Fully integrated system for dsdna quantification. Affordable, easy-to-use, sensitive fluorometers designed for quick and accurate fluorescence measurements. QuantiFluor-ST Handheld Fluorometer with UV/Blue Channels QuantiFluor-P Handheld Fluorometer with Green/Blue Channels 1 each E6090 1,878 1 each E6100 1,430 Related Products Pfu DNA Polymerase High-fidelity, thermostable enzyme, replicates DNA at 75 C, catalysing the polymerisation of nucleotides into duplex DNA in the 5 3 direction in the presence of magnesium. It possesses 3 5 exonuclease, proof-reading activity and is recommended for use in PCR and primer extension reactions that require high-fidelity synthesis. QuantiFluor-P Handheld Fluorometer with UV/Blue Channels 1 each E6105 1,430 Product Size Concentration Cat.No. Price Pfu DNA Polymerase 100u 500u 2-3u/μl 2-3u/μl M7741 M

9 Diamond Nucleic Acid Dye Room temperature stable, sensitive fluorescent dye to stain and visualise nucleic acids in gels. Does not require pre-washing or destaining of gels. Cost-effective alternative to ethidium bromide and less expensive than SYBR Gold. Diamond Nucleic Acid Dye 500μl H PCR Cloning Subclone PCR and RT-PCR products rapidly and affordably using Promega pgem T-Vector family. dntps Promega is a primary manufacturer of dntps, offering bulk and custom options. Choose the dntp mixes or purchase individual tubes. Promega UK Ltd Delta House Southampton Science Park Southampton SO16 7NS RNasin Plus RNase Inhibitor Recombinant mammalian RNase inhibitor capable of inhibiting eukaryotic RNases (e.g. RNase A and RNase B) similarly to human placental RNase inhibitor. Tested in RT-PCR and compatible with enzymes such as AMV, M-MLV and ImProm-II Reverse Transcriptases or Taq and Tfl DNA Polymerases. CXR Reference Dye For use as a passive reference dye in real-time quantitative PCR. 16