Whether you're new to real-time PCR, also called quantitative PCR (qPCR), or want to learn about new applications for qPCR, we have the learning material, including videos and webinars, to help you understand the technology and get started quickly.
Quantitative PCR defined
Real-time quantitative PCR, also known as qPCR, combines PCR amplification and detection into a single step. This eliminates the need to detect products using gel electrophoresis, but more importantly, enables the method to be truly quantitative. With qPCR, fluorescent dyes are used to label PCR products during thermal cycling. Real-time PCR instruments measure the accumulation of fluorescent signal during the exponential phase of the reaction for fast, precise quantification of PCR products and objective data analysis.
Labeling, Detecting, and Quantifying PCR products
qPCR reaction products are fluorescently labeled using one of two main strategies:
- TaqMan® fluorogenic probes—target-specific oligonucleotides that produce a fluorescent signal only when the target DNA is amplified during qPCR
- SYBR® Green I dye—binds to double-stranded DNA and emits fluorescence only when bound
Reactions are run in real-time PCR instruments with thermal cycling and fluorescence detection capabilities. By using highly efficient PCR primers and optimal conditions for amplification, every target molecule is copied once at each cycle and data are captured throughout the thermal cycling. Since qPCR reactions are set up with a large molar excess of PCR primers and thermostable DNA polymerase, in the early rounds of thermal cycling, target-template is the limiting factor for the reaction thus, fluorescent signal is directly proportional to the amount of target in the input sample.
The Applied Biosystems® family of qPCR instrument and software systems, qPCR master mixes, and TaqMan® probe based assays for gene expression and genetic variation make qPCR more accessible than ever. These systems offer cutting-edge software designed to support your application, and advanced fluorescence detection to accommodate the real-time chemistry of your choice.
- Essentials of Real-Time PCR
- qPCR vs. Digital PCR vs. Traditional PCR
- Absolute vs. Relative Quantification for qPCR
- TaqMan® Assays vs. SYBR® Green Dye for qPCR
- Real-time PCR: Understanding Ct Application Note
- Real-time PCR Troubleshooting Tool
- Amplification Efficiency of TaqMan® Gene Expression Assays
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