RNA Quantification Using the GoTaq® RT-qPCR Systems

2 May 2017

Promega GoTaq^® products for RNA quantification offer two dye-based systems for accurate, sensitive, broad-range quantification of RNA targets. This application note demonstrates the successful amplification of 11 differentially expressed mRNA targets using the GoTaq^® 1-Step RT-qPCR System. One of these targets was also amplified to demonstrate equivalent performance between the GoTaq^® 1-Step and GoTaq^® 2-Step RT-qPCR Systems. It was found that reactions with the GoTaq^® RT-qPCR Systems have a higher change in fluorescence and earlier Cq values than other RT-qPCR systems.

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GoTaq® 2-Step RT-qPCR System
Promega Corp.
GoTaq®2-Step RT-qPCR System is a reagent system for quantitative analysis of RNA using a two-step reverse transcription-quantitative PCR (RT-qPCR) method. The first step consists of cDNA synthesis using the GoScript™ Reverse Transcription System. Optimized reagents and ultra-active, GoScript™ Reverse Transcriptase result in high-efficiency, full-length cDNA synthesis from rare and abundant targets with up to 5 µg of RNA input. In the second step, cDNA is quantified using the GoTaq®qPCR Master Mix. Formulated with a hot-start DNA polymerase, the GoTaq® qPCR Master Mix is optimized for reproducibly specific, sensitive, highly efficient qPCR using the standard or fast mode of a real-time PCR instrument. The GoTaq® qPCR Master Mix contains BRYT™ GREEN dye which can be twice as bright as of SYBR® Green I. The superior performance of these two steps allows researchers to observe earlier quantification cycle values over broader linear ranges. The GoTaq® 2-Step RT-qPCR System provides a ready-to-use kit for analyzing a wide range of RNA targets by combining the high-activity of GoScript™ Reverse Transcription System with the ultra-bright fluorescence of GoTaq® qPCR Master Mix. The system is robust, requires little optimization, is flexible for a variety of applications, and carries the PCR performance guarantee.
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GoTaq® Probe 1-Step RT-qPCR System
Promega Corp.
The GoTaq® Probe 1-Step RT-qPCR System is optimized for quantitative PCR assays in the hydrolysis probe detection format. The system enables detection and relative quantification of RNA expression levels using a one-step RT-qPCR method, combining GoScript™ Reverse Transcriptase and GoTaq® Probe qPCR Master Mix in single-step real-time amplification reactions. The GoScript™ RT Mix for 1-Step RT-qPCR (50X) combines optimized amounts of GoScript™ Reverse Transcriptase, RNasin® Plus RNase Inhibitor and additives to enhance single-step reactions. The GoTaq® Probe qPCR Master Mix is provided as a ready-to-use, stabilized 2X formulation that includes all components for qPCR (except template, primers and probe). This master mix does not contain a reference dye; however, a separate tube of carboxy-X-rhodamine (CXR) reference dye is included with this system, allowing users to add reference dye to amplification reactions if desired. The GoTaq® Probe qPCR Master Mix is designed to provide resistance to a wide range of PCR inhibitors. This formulation uses antibody-mediated hot-start chemistry, allowing reaction setup to be performed at room temperature. The master mix also employs rapid hot-start activation and processive enzymes, making it compatible with both standard and fast instrument cycling programs. Features - Benefits • Superior Performance: Sensitive detection on any real-time instrument • Enhanced Stability: Exceptional room-temperature setup makes it suitable for automation and high-throughput detection • Versatility: Compatible with both fast and standard cycling methods • Robustness: High-efficiency, full-length cDNA synthesis in the presence of inhibitors • Confidence: The Promega PCR Performance Guarantee Applications • Sensitive sequence detection and quantification • Gene expression analysis • Detection of sequence variants • Fluorescent detection
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GoTaq® Long PCR Master Mix
Promega Corp.
GoTaq® Long PCR Master Mix contains the high-performance GoTaq® Hot Start Polyemrase in a specially formulated mixture with a proprietary thermostable proofreading polyemrase. This optimized enzyme mixture allows efficient amplification of up to 40kb from lambda DNA or 30kb from human genomic DNA. The presence of a proofreading enzyme to repair DNA mismatches and a highly processive polyemrase allows the polyemrase to contninue to elongate the DNA much farther, resulting in longer DNA amplification. The optimized formulation of the GoTaq(R) Long PCR Master MIx components enables simple reaction setup and provides consistently efficient, accurate and robust amplification of long DNA amplicons. GoTaq® Long PCR Master Mix Features: Easy Setup - Handle reactions at room temperature. Enhanced Yields - Optimized components maximize results. Improved Processivity - Enzyme blend provides robust amplification. Assured Confidence - Includes control primer pair and gDNA template. Improved Sensitivity - Amplify as little as 2ng with high specificity.
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GoTaq Real-Time PCR Systems
Promega Corp.
The GoTaq®Real-Time PCR Systems are ready-to-use 2X master mixes optimized for quantitative PCR in dye (using BRYT(R) Green Dye) or hydrolysis probe detection format. The 1-Step and 2-Step RT-qPCR Master Mixes contain GoScript™ Reverse Transcriptase for efficient first-strand cDNA synthesis. GoTaq Real-Time PCR Systems Features & Benefits: Higher fluorescence, often resulting in earlier Cqs Direct substitute for SYBR® Green I products Enhanced stability for automated setup Choose from qPCR, 1-Step or 2-Step RT-qPCR options Compatible with both fast and standard cycling methods Sensitive quantitation on most real-time instruments
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PCR and Thermal CyclingGene Expression and Molecular CloningDNA / RNA QuantificationFluorescenceqPCRGene ExpressionRNATranscriptomicsRT qPCRReal-Time PCR

GoTaq® Probe 1-Step RT-qPCR System

Promega Corp.

The GoTaq® Probe 1-Step RT-qPCR System is optimized for quantitative PCR assays in the hydrolysis probe detection format. The system enables detection and relative quantification of RNA expression levels using a one-step RT-qPCR method, combining GoScript™ Reverse Transcriptase and GoTaq® Probe qPCR Master Mix in single-step real-time amplification reactions. The GoScript™ RT Mix for 1-Step RT-qPCR (50X) combines optimized amounts of GoScript™ Reverse Transcriptase, RNasin® Plus RNase Inhibitor and additives to enhance single-step reactions. The GoTaq® Probe qPCR Master Mix is provided as a ready-to-use, stabilized 2X formulation that includes all components for qPCR (except template, primers and probe). This master mix does not contain a reference dye; however, a separate tube of carboxy-X-rhodamine (CXR) reference dye is included with this system, allowing users to add reference dye to amplification reactions if desired. The GoTaq® Probe qPCR Master Mix is designed to provide resistance to a wide range of PCR inhibitors. This formulation uses antibody-mediated hot-start chemistry, allowing reaction setup to be performed at room temperature. The master mix also employs rapid hot-start activation and processive enzymes, making it compatible with both standard and fast instrument cycling programs. Features - Benefits • Superior Performance: Sensitive detection on any real-time instrument • Enhanced Stability: Exceptional room-temperature setup makes it suitable for automation and high-throughput detection • Versatility: Compatible with both fast and standard cycling methods • Robustness: High-efficiency, full-length cDNA synthesis in the presence of inhibitors • Confidence: The Promega PCR Performance Guarantee Applications • Sensitive sequence detection and quantification • Gene expression analysis • Detection of sequence variants • Fluorescent detection

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RNA Quantification Using the GoTaq® RT-qPCR Systems

Related products

GoTaq® 2-Step RT-qPCR System

GoTaq® Probe 1-Step RT-qPCR System

GoTaq® Long PCR Master Mix

GoTaq Real-Time PCR Systems

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GoTaq® 2-Step RT-qPCR System

GoTaq® Probe 1-Step RT-qPCR System

GoTaq® Long PCR Master Mix

GoTaq Real-Time PCR Systems