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CAPITAL™ qPCR Probe Mix, 4×

Features

  • Best in-class performance for both single and multiplex detection
  • Convenient master mix with high specificity in pathogen detection
  • Highly sensitive for low-abundance DNA targets

Applications

  • Standard and fast cycling qPCR with rapid extension rate for early Ct values
  • For use on a wide range of probe technologies including Taqman®, Molecular Beacons® and Scorpion® probes

Purchase CAPITAL™ qPCR Probe Mix, 4×

Availability: In stock Bulk requests please inquire at oem@biotechrabbit.com

Catalog # Size Package Content Price* Qty
BR0501401 200 rxn of 20 µl 1 ml CAPITAL qPCR Probe Mix
€84.00
BR0501402 1000 rxn of 20 µl 5 × 1 ml CAPITAL qPCR Probe Mix
€270.00
BR0501403 4000 rxn of 20 µl 4 kits BR0501402 CAPITAL qPCR Probe Mix (1000 rxn each)
€972.00
BR0501501 200 rxn of 20 µl 1 ml CAPITAL qPCR Probe Mix LRox
€84.00
BR0501502 1000 rxn of 20 µl 5 × 1 ml CAPITAL qPCR Probe Mix LRox
€270.00
BR0501503 4000 rxn of 20 µl 4 kits BR0501502 CAPITAL qPCR Probe Mix LRox (1000 rxn each)
€972.00
BR0501601 200 rxn of 20 µl 1 ml CAPITAL qPCR Probe Mix HRox
€84.00
BR0501602 1000 rxn of 20 µl 5 × 1 ml CAPITAL qPCR Probe Mix HRox
€270.00
BR0501603 4000 rxn of 20 µl 4 kits BR0501602 CAPITAL qPCR Probe Mix HRox (1000 rxn each)
€972.00

*Does not include applicable tax, shipping & handling, or other charges.

OR

biotechrabbit™ CAPITAL qPCR Probe Mix for quantifying genomic, cDNA and viral sequences provides outstanding performance in single and multiplex qPCR. The high sensitivity provided by the mix is ideal for detection of low-abundance DNA targets in various applications, such as pathogen detection.

CAPITAL qPCR Probe Mix uses proprietary combination of enzyme and buffer chemistry for efficient extension and early Ct in single and multiplex qPCR. To enable the use of the kit on qPCR platforms with different reference dye concentration requirements, three kit formats are available: a one-step kit containing no ROX, as well as LRox and HRox versions containing ROX in the corresponding concentrations.

 

Info: Recommended annealing temperature is 2°C above primer Tm (use gradient PCR to optimize the annealing temperature).

 

Component

Composition

CAPITAL qPCR Probe Master Mix

Optimized 4× QPCR Probe Master Mix

LRox / HRox mixes

Rox incorporated in the mix in low / high concentration

 

STORAGE

−20°C (until expiry date – see product label).
Avoid multiple freeze thaw cycles by preparing aliquots.

 

Quality Control
Functional assay

Mix tested functionally in qPCR.

ROX reference dye
  • See PCR cycler instruction for recommended concentration of ROX passive reference dye.
Notes
  • For efficient amplification under fast cycling conditions use amplicon lengths between 80 bp and 200 bp.
  • The shorter the amplicon length the faster the reaction can be cycled.
  • Amplicon lengths should not exceed 400 bp.
  • Primers should have a predicted melting temperature of around 60°C, using default Primer 3 settings (http://frodo.wi.mit.edu/primer3/).
  • For TaqMan® probes choose probe close to 5’ primer, avoid terminal guanosine residues.
Prevention of PCR contamination

When assembling the amplification reactions, care should be taken to eliminate the possibility of contamination with undesired DNA.

  • Use separate clean areas for preparation of samples and reaction mixtures and for cycling.
  • Wear fresh gloves. Use sterile tubes and pipette tips with aerosol filters for PCR setup.
  • Use only water and reagents that are free of DNA and nucleases.
  • With every PCR setup, perform a contamination control reaction that does not include template DNA.
Basic Protocol
  • Keep the master mix protected from light until you use it.
  • Aliquot the master mix to minimize freeze-thaw cycles and light exposure.
  • Thaw on ice and mix very well all reagents. Assemble and keep all reactions on ice.
  • Use only high quality optically clear reaction plates and seals designed for fluorescence applications.
  • Do not use corner wells or use a more robust seal.
  • Reserve plate positions for positive (control DNA) and negative (water or buffer) controls.
  • First pipette the primer mixture, then add the template and last the Master Mix.
  • Before preparing mixes, calculate the volume needed according to the reaction number plus one extra.
  • To have a better correlation, run the reactions in triplets.

Component

Volume

Final concentration

Primer Mix (Reverse and Forward)

Variable

100–400 nM

Too high primer concentrations result in unspecific amplification and should be avoided.

Specific Probe

Variable

200 nM

Template DNA

Variable

10 pg – 100 ng

Use diluted or undiluted cDNA from less than 1 µg RNA

CAPITAL qPCR Probe Mix, 4×

5 µl

Nuclease free water

Variable

 

Total volume

20 µl

 

  • Gently mix the reactions without creating bubbles (do not vortex). Bubbles will interfere with fluorescence detection.
  • Place the reaction into the PCR cycler.
Cycling Program

Step

Temperature

Time

Cycles

Initial activation

95°C

2-3 min

1

Denaturation

95°C

10–15 s

40–45

Annealing/Extension*

(60-68°C)

30 s


*Recommended annealing/extension temperature is primer Tm +2°C. Use gradient PCR to optimize the annealing temperature. Do not use temperatures below 60°C. Do not exceed 30 seconds.
For melt analysis refer to instrument instructions. 
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