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Real-time PCR (qPCR) Probes

The Real-time PCR (qPCR) or 5'-Nuclease assay is a sensitive assay for online monitoring of amplification kinetics. Real-time PCR Probes consist of an oligonucleotide complementary to the amplified probe. The probe is labeled at the 5'-end with the fluorescence donor (usually Fluorescein, HEX or TET) and a few bases downstream or on the 3'-end with a quencher, usually TAMRA. When no complementary sequence is available, the fluorescence of the donor is quenched. During PCR the labeled oligonucleotide hybridizes to the target sequence and the 5'-dye is removed by 5'3'-exonuclease activity of Taq. The fluorescence of the donor is no longer quenched and can be measured. The fluorescence intensity is proportional to the amount of PCR-product formed during the early exponential phase of PCR (threshold value). 

Real-time PCR principle
Real-time PCR principle


In the probe design it is important that the melting point of the probe is higher than that of the upstream primer. The 3'-end of the probe must be protected against chain elongation during PCR. For blocking the 3'-end phosphate, cordycepin, 2',3'-dideoxynucleosides, inverseT or the quencher dye itself can be used. Labeling the 3'-end with TAMRA or another quencher can be achieved by using a 3'-amino-linker. Labeling internal positions of the sequence is possible by incorporating a modified thymidine residue which contains an amino-link at the 5-position of the base. This attachment does not interfere with hybridization.

We offer

  • great variety of oligonucleotides with different labels and their combinations (see our list of 200 dyes)
  • 5´- , 3´- or internal labels
  • Click chemistry for flexible dual labeling
  • high purity probes and thorough quality control

 

We offer a large selection of dye combinations suitable for Real-time PCR probes. Please make a request if your combination of interest is not listed below! 

Product

Amount

Cat. no.

Price/coupling

Real-time PCR probe

5' FAM 3' TAMRA

5 nmol

5-0901-002

142.00 EUR

25 nmol

5-0901-003

184.00 EUR

100 nmol

5-0901-004

347.00 EUR

Real-time PCR probe

5' HEX 3' TAMRA

5 nmol

5-0901-012

194.00 EUR

25 nmol

5-0901-013

268.00 EUR

100 nmol

5-0901-014

446.00 EUR

Real-time PCR probe

5' TET 3' TAMRA

5 nmol

5-0901-022

194.00 EUR

25 nmol

5-0901-023

268.00 EUR

100 nmol

5-0901-024

373.00 EUR