Product info

scorpions primers conceptual illustration

Scorpions™ primers combine primer and probe into a single bifunctional molecule. They contain a primer sequence at the 3′ end and a hairpin loop structure at the 5′ end. Like molecular beacons, the hairpin brings the reporter and quencher into proximity and the loop contains a sequence complementary to the target.

These primers utilise a unimolecular mechanism that acts faster in solution for instantaneous fluorescence in real-time PCR. Similar to molecular beacons, Scorpions primers do not require enzymatic cleavage of the probe during PCR cycling. These qualities make Scorpions primers valuable tools for rapid, real-time PCR, endpoint PCR, SNP detection and gene quantification.

 

Scorpions primer structure

These molecules incorporate two distinct structures: a target-specific probe and a target-specific PCR primer.

Probe sequence:

  • 5′ fluorophore: The reporter dye emits fluorescence when the probe is linearised and hybridised to the target, separating the dye and quencher.
  • 3′ quencher: When the probe is in its hairpin structure, the proximity of the quencher to the reporter prevents fluorescence emission.
  • Stem: The stem is a double-stranded region formed by binding the complementary sequences (5-7 nt) at both ends of the probe.
  • Loop: The loop is a 18-30 nt sequence that is complementary to the target sequence.

Primer sequence:

  • Primer region: A target-specific PCR primer is covalently linked to the 3' end of the probe sequence through a blocking moiety, often a hexaethylene glycol (HEG) modification. This moiety prevents polymerase extension into the probing element, which could generate false positives.

 

Scorpions primer mechanism

During the first PCR cycle, the primer region hybridises to the complementary DNA template sequence and extends to form a primer extension product. The primer extension product contains the desired probe target sequence. At this stage, the probe region remains in a closed hairpin conformation. The presence of a PCR blocker in the linker sequence prevents the polymerase-mediated duplication of the probe region.

During the second PCR cycle, the target-binding region (loop region) of the probe sequence hybridises to the complementary sequence within the primer extension product. Hybridization forces the hairpin open, separating the fluorophore and quencher, and releasing fluorescence.

Key benefits:

  • Enhanced specificity: The stem-loop conformation confers high specificity, making Scorpions primers ideal for SNP/mismatch discrimination.
  • High signal-to-noise: Scorpions primers offer superior fluorophore quenching due to the proximity and the direct energy transfer between reporter and quencher while in the closed hairpin conformation.
  • Melt curve analysis suitability: Because Scorpions primers generate fluorescence under non-hydrolytic conditions by target hybridisation, post-PCR melt curve analysis can be performed.
  • Rapid hybridisation: The proximity of the probe region and the target sequence kinetically favours formation of the probe-template hybrid over template duplex re-annealing. The unimolecular event enables rapid signal generation during hybridization.
 

Select the best dye for your application

We offer Scorpions primers labelled with a wide selection of dyes, enabling you to select options based on instrumentation and assay design. Quencher options for this probe type include Black Hole Quenchers, BHQ-1 and BHQ-2.

5' fluorescent dye

Abs (nm)

Em (nm)

Internal quencher

PCR blocker

  FAM 495 520 BHQ-1 HEG
  CAL Fluor Gold 540 522 544 BHQ-1 HEG
  CAL Fluor Orange 560 538 559 BHQ-1 HEG
  Quasar 570 548 566 BHQ-2 HEG
  TAMRA 557 583 BHQ-2 HEG
  CAL Fluor Red 590 569 591 BHQ-2 HEG
  CAL Fluor Red 610 590 610 BHQ-2 HEG
  CAL Fluor Red 635 618 637 BHQ-2 HEG
  Quasar 670 647 670 BHQ-2 HEG
  Quasar 705 690 705 BHQ-2 HEG

Use the spectral overlay tool to view compatible dyes for your instrument. Learn more

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  • Oligos for North America, South America, and APAC will be supplied out of our Petaluma, US manufacturing facility while oligos for EMEA will be supplied out of our Lystrup, Denmark manufacturing facility unless you have an agreement in place.
  • Our advertised turnaround time applies to orders received before 12PM for the local manufacturing site. Turnaround time is defined as the time of checkout until shipment.

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Technical Specs

Scorpions primers specifications
Dyes FAM, CAL Fluor gold 540, CAL Fluor Orange 560, Quasar 570, TAMRA, CAL Fluor Red 590, CAL Fluor Red 610, CAL Fluor Red 635, Quasar 670, Quasar 705
Quenchers BHQ-1, BHQ-2
PCR blocker HEG
Yield Dye dependent (see product listing)
Purification DUAL HPLC
Delivery format Dry or in solution (water, TrisHCl, T10E0.1, T10E1)
Quality control MS and UHPLC
Quality standard ISO 9001 or ISO 13485
Shelf life 12 months from date of manufacture

 

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