HomeQuantitative PCR (qPCR)Dual-Labeled DNA Probes: Fluorescent Probes

Dual-Labeled DNA Probes: Fluorescent Probes

What is real-time quantitative PCR?

Real-time quantitative PCR is an innovative and robust technique for quantitative analysis of gene expression, mutation detection, and SNP genotyping. Real-time quantitative PCR was developed to overcome the basic weakness of classical PCR technology: whereas PCR can indicate the presence of a particular nucleic acid in a sample after a completed PCR amplification, it cannot directly quantify the amount of amplicon.

Real-time quantitative PCR systems quantify the amount of amplified product through the detection and quantitation of a fluorescent dye. This dye, upon excitation, emits a light or signal that increases in intensity in direct proportion to the amount of amplified PCR product and, thereby, quantifies the amount of target DNA. The assays are directly performed in a single, closed tube without any purification or separation step, avoiding risks of contamination and the need for post-PCR handling.

Real-time quantitative PCR systems also enable mutation detection via melting-curve analysis, as each double-stranded DNA product has its own specific melting temperature (Tm). Melting-curve data enables the researcher to differentiate between specific PCR products and non-specific PCR products, such as primer-dimers.

Why use sequence-specific probes?

Detection by real-time quantitative PCR can be sequence-specific or non sequence specific, and the method chosen will effect the performance of your real-time quantitative PCR results. For instance, SYBR® Green 1, a non-sequence-specific reagent, can bind to any double-stranded DNA including primer-dimers and other non specific reaction products, and so may result in an over-estimation of the amount of target DNA. However, sequence-specific probes bind only to the target template and therefore achieve accurate and reliable quantification of the amount of target DNA.

Design Service for Fluorescent Probes

Save time by letting our Genosys design your fluorescent probes and associated primers for your real-time quantitative PCR applications or mutation detection needs.

Using our in-house expertise and powerful design tools, we will provide you with the optimal probe design. Should you work with SYBR® Green I and need primer sequences, we recommend that we also design the appropriate fluorescent probe at the same time.

Our team of design specialists is always available to answer any questions you may have about our design service. Please contact us at

Note: This design service applies only to probes and primers for real-time quantitative PCR applications and mutation detection needs. If you have already designed your own primer and/or probe sequence, we can also test your design.