RT-qPCR

One Step RT-qPCR Kit

● Product Name: Magigen One Step RT-qPCR II MIX Kit 100T   

● Catalog#: A014S

● Size: 100T

● Taq Polymerase Price: 115USD

● Contents

Composition                           Vol

2× Reaction Buffer 3^a            1.25 ml   

RT-qPCR Enzyme Mix II^b        100 µl   

a. Contains dNTPs、Mg²⁺ etc.；

b. Contains MMLV Reverse Transciptase、RNase Inhibitor 以及 Hotstart Taq DNA Polymerase.

● RT qPCR Kit Application

Magigen one step RT qPCR Kit is used for probe-based qPCR with RNA as a template. Recommended for detecting nucleic acids extracted from samples as detection template.

Magigen RT qPCR premix kit makes it possible to complete the detection and gene expression workflow of COVID-19 and other pathogens in one step.

● Advantages and Features

Magigen one step RT-aPCR kit integrates the Real Time RT-PCR reverse transcriptase (MMLV Reverse Transcriptase) and antibody mediated hot start enzyme (Hotstart Taq DNAPolymerase), combined with an optimized buffer system, using gene specific primers (GSP) to complete reverse transcription and qPCR reactions in a single tube without the need for additional tube opening/pipetting operations. qPCR operation is simple, and reduces contamination risk.

● Storage Conditions and Shelf Life

Sealed and stored at -20 ℃,   12 months.

● Applicable instruments

AB series, ROCHE series, SLAN series, MA6000   and other fluorescence quantitative PCR instruments.

● RT qPCR testing steps

Taking SLAN-96P as an example

1. Prepare reaction mixture in RNase free centrifuge tube

Composition                           Vol (μL)

2× Reaction Buffer 3                12.5

RT-qPCR Enzyme Mix II            1

Foward Primer(10μM)               0.5

Reverse Primer(10μM)              0.5

Probe(10μM)                             0.25

RNA template                             2

RNase-free Water                     Up to 25

a. The amount of primers, probes, and templates used in the reaction system can be adjusted independently. RNase free water can be added if necessary;

b. The dosage of the reaction system can be adjusted according to the following principles:

    1) A primer concentration of   0.2 μM in the reaction system could achieve good amplification, but when the amplification performance is not good, you can          adjust the primer concentration range from 0.1 to 1.0 μ M ;

   2) The probe concentration can be adjusted between 50-250nM.

2. Perform one step RT qPCR reaction according to the following conditions:

Cycle step   Temperature   Time       Number of cycles

1                     50℃          15min      1

2                     95℃          2.5min     1

3                     95℃          15s           45

4                     60℃          30s

a. The extension time is adjusted according to the shortest time required for data collection by the Real Time PCR instrument.

● Cautions

1. This RT-qPCR Kit is for scientific research use only. Please read this manual carefully before use.

2. Wait for 2 × Reaction Buffer 3 to thaw and mix thoroughly and centrifuge briefly Before use; RT qPCR Enzyme Mix II can be briefly mixed and centrifuged before use, and accurately aspirated;

3. Please strictly follow the management standards of the gene amplification laboratory for experimental operations. The preparation of the reaction solution should use RNase free gun heads, reaction tubes, etc., and be careful to prevent contamination of the amplification products.

4. After the experiment, the waste generated during the testing process should be disposed of in accordance with relevant regulations.

➢ Why is nuclease free water used?

The presence of nucleases such as DNase and RNase in water can degrade precious molecular samples and even ruin experiments. To prevent DNA and RNA sample loss, it is essential that highly pure, nuclease-free water be used in applications such as PCR, cDNA synthesis, nucleic acid purification, sequencing, and cloning.

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