Dual-labeled Probes Services

With the development of nucleic acid quantitative technology in the past decades, competitive reverse transcription PCR (RT-PCR) has become the most advanced method. This technology can achieve the quantification of nucleic acids with higher specificity and accuracy as well as lower template copy numbers, compared with earlier developed technologies such as the Southern and Northern blot analysis and RNase-protection assays. Among all kinds of fluorescence-based assay systems, dual-labeled fluorescent oligonucleotide probes can provide the highest template specificity. A dual-labeled probe is normally 20 - 30 bases in length modified with a fluorophore at 5'-end and a quencher at 3'-end.

Working mechanism

As shown in Figure 1, there are three states of dual-labeled probes, including (A) unhybridized state where the fluorescence cannot be emitted as the fluorophore is in a close distance with quencher, (B) denatured state where the probe is melted or denatured by heating and behaves in a random coil conformation, and (C) hybridized state where the probe and target sequences are complimentarily bound. During elongation, as DNA polymerase incorporates nucleotides in a 5' to 3' direction from the primer, it will encounter the 5'-end of the probe, followed by cutting the nucleotide with fluorophore; consequently, the fluorophore and quencher are separated permanently and fluorescence is released.

Dual-labeled Probes ServicesFigure 1. Different states of dual-labeled probes.

Our services

There are many excellent properties of dual-labeled probes. Besides, dual-labeled probes can also be applied for SNP detection, quantitative and qualitative analysis, real-time and endpoint PCR analyses and allelic discrimination, etc. Our company can design and produce kinds of highly efficient dual-labeled probes.

  • Related fluorophores include but are not limited to FAM, HEX, JOE, ROX, TET, TAMRA, CIV, CAL Fluor Red 610, Quasar 670/570/705 and CAL Fluor Gold 540/590, etc.
  • Different quenchers are available based on different requirements, such as TAMRA, DABCYL, DDQ, etc.
  • The best length of this kind of probe is 20-30 bases.
  • All probes will be monitored by QC with MALDI-TOF MS and analytical HPLC

References:

  1. Johansson M K, Fidder H, Dick D, Cook R M. Intramolecular dimers: a new strategy to fluorescence quenching in dual-labeled oligonucleotide probes. American Chemical Society, 2002, 124(24): 6950-6956.
  2. Yeung A T, Holloway B P, Adams P S, Shipley G L. Evaluation of dual-labeled fluorescent DNA probe purity versus performance in real-time PCR. BioTechniques, 2004, 36(2): 266-275.
  3. Navarro E, Serrano-Heras G, Castaño M J, Solera J. Real-time PCR detection chemistry. Clinica Chimica Acta, 2015, 439: 231-250.
* It should be noted that our service is only used for research, not for clinical use.

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