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ZipScript™ One-Step RT-qPCR Kit

Product Description

The ZipScript One-Step RT-qPCR Kit is a highly sensitive and reproducible RT-qPCR solution optimized for real-time PCR. The 25X enzyme mix is accompanied by a 2X reaction buffer. If a passive reference dye, such as ROX, is required for your assay, it can be purchased separately (P/N: Y9390).

Quality Control Analysis

The functionality of the ZipScript is evaluated by amplification of two mRNA transcripts in a one-step quantitative RT-qPCR assay. The amplification threshold (Ct) of the test lot is compared to a reference lot.

Kit Contents:
25X ZipScript Enzyme Mix 2X ZipScript Reaction Buffer I

Notes:
Enzyme components were tested prior to formulation of the master mix and found free of contaminating endonucleases and exonucleases. Enzyme purity was >99% as determined by SDS-PAGE and negligible E.coli genomic DNA contamination was confirmed by qPCR. Specific activity was verified for each enzyme pre-formulation.

Protocol

ZipScript One-Step RT-qPCR Reaction Setup:

  1. All components and reaction set up should be kept on ice.
  2. Thaw the 2X ZipScript Reaction Buffer I completely and vortex for 3-5 seconds to mix thoroughly. Quick spin to collect contents if necessary.
  3. Prepare primer/probe mix. A final concentration of 0.4-0.9 µM for each primer and 0.1-0.5 µM for probe is recommended. However, the optimal concentration for primers/probe needs to be empirically determined for each assay.
  4. Determine the number of reactions to prepare, including No Template Controls (NTCs). Add 10% extra volume to compensate for the pipetting loss.
  5. Follow the table below to set up the reaction mix. It is recommended to make a master mix to minimize variations and potential errors.
    Components Volume/Rxn Final Conc
    Nuclease-Free Water To a final 20 µL reaction volume
    2X ZipScript Reaction Buffer I 10 µL 11X
    50X ROX (optional) 0.4 µL 1X
    Primer/Probe Mix X µL Variable
    25X ZipScript Enzyme Mix 0.8 µL 0.8 µL
    RNA Template Variable
  6. Seal PCR plate and spin briefly to bring down reagents.

Thermal Cycling Conditions:

Program the cycling conditions based on the recommendations below.

Standard Cycling Program*

Steps Temperature Time Cycles
Reverse Transcription 50°C 15 min 1
Taq Activation/InitialDenaturation 95°C 2 min 1
Denaturation 95°C 15 min 40
Annealing/Extension* 60°C 30-60 sec

* Cycling parameters can be modified (especially the annealing/extension condition) to fit specific primer/probe selection.

Product Information

ZipScript™ One-Step RT-qPCR Kit
Part NumberP7640L
Price$1,338
ConcentrationN/A
Unit Size1000 Reactions
SDS Available on request

Product Specification*

Storage Temperature-25°C to -15°C
Assay RT-qPCR
SpecificationAmplification of Test Lot within 1Ct of Reference Lot for two different mRNA targets

* For a detailed summary of assay conditions and data, refer to the Quality Controls Analysis section.

Additional Downloads

Limitations of Use
This product was developed, manufactured, and sold for in vitro use only. The product is not suitable for administration to humans or animals. SDS sheets relevant to this product are available upon request.

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