Guidelines and Considerations
Real-Time PCR Assays Real-time Polymerase Chain Reaction (PCR) is the ability to monitor the progress of the PCR as it occurs (i.e., in real time). Data is therefore collected throughout the PCR process, rather than at the end of the PCR. This completely revolutionizes the way one approaches PCR-based quantitation of DNA and RNA. In real-time PCR, reactions are characterized by the point in time during cycling when amplification of a target is first detected rather than the amount of target accumulated after a fixed number of cycles. The higher the starting copy number of the nucleic acid target, the sooner a significant increase in fluorescence is observed. In contrast, an endpoint assay (also called a “plate read assay”) measures the amount of accumulated PCR product at the end of the PCR cycle.
Dive into the basics of qPCR and find information about Sequence Detection Chemistries and Quantitation Assays, download the guide.
Absolute Standard Curve Method versus Relative Standard Curve Method
The standard curve method in real-time quantitative PCR can be used to determine either the absolute quantity (AQ) or the relative quantity (RQ) of target sequence.
This quick reference highlights the differences between the methods. A schematic diagram is provided to illustrate how relative target quantity is determined by the software in an easy-to-understand manner.
Real-time PCR can be challenging to learn, but it doesn’t have to be.
Become a qPCR expert – watch the Taq Talk video series at YouTube.
Here are some of our favorites:
The purpose of ROX reference dye in real-time PCR
How real-time PCR assay efficiency can affect your data
Finding the Right TaqMan Gene Expression Assay
