ZipScript One-Step RT-qPCR Kit

OEM by QIAGEN offers bulk manufacturing of ZipScript One-Step RT-qPCR Kit in custom formulations.

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ZipScript One-Step RT-qPCR Kit (1000 reactions)

Cat. No. / ID:  P7640L

For 1000 reactions (evaluation pack): 25X ZipScript Enzyme Mix and 2X ZipScript Reaction Buffer I.
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The ZipScript One-Step RT-qPCR Kit is intended for molecular biology applications. This product is not intended for the diagnosis, prevention, or treatment of a disease.
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Features

  • Highly sensitive and reproducible RT-qPCR solution
  • Automation friendly
  • Features a blend of EnzScript and Phoenix Hot Start DNA Polymerase
  • Efficient RNA dependent multiplex qPCR formulation

Product Details

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.

Inquire for lyophilized or lyo ready options.

Performance

Mix properties

  • Storage temperature: –25°C to –15°C

Principle

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

Procedure

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 and probe mixture. 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 and 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. Use the amounts listed in the table below to set up the reaction mix. We recommend making a master mix to minimize variations and potential errors.
    Components Volume per reaction Final concentration
    Nuclease-free water To a final reaction volume of 20 µl
    2X ZipScript Reaction Buffer I 10 µl 11X
    50X ROX (optional) 0.4 µl 1X
    Primer and probe mixture Variable Variable
    25X ZipScript Enzyme Mix 0.8 µl 0.8 µl
    RNA Template Variable Variable
  6. Seal PCR plate and spin briefly to bring down reagents.
  7. Program the cycling conditions based on the recommendations below.
    Steps Temperature Time Cycles
    Reverse transcription 50°C 15 minutes 1
    Taq activation and initial denaturation 95°C 2 minutes 1
    Denaturation 95°C 15 seconds 40
    Annealing and extension* 60°C 30–60 seconds -

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

    Quality control analysis

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

    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.

Applications

This OEM by QIAGEN product is available for bulk purchase for the following commercial assay applications.

  • One-step RT-qPCR

Resources

Certificates of Analysis (1)