One Step Fast RT-qPCR Probe Premix-UNG

Cat No: HCR5143A

Package: 100RXN/1000RXN/10000RXN

One Step Fast RT-qPCR Probe Kit U+ is designed for one-step real-time RT-PCR using probe detection, all RT-PCR steps can be performed in a single tube.

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Product Description

Product detail

Cat No: HCR5143A

One Step RT-qPCR Probe Kit (FOR FAST) is a probe-based RT-qPCR fast detection kit suitable for single-plex or multiplex quantitative PCR using RNA as a template (such as RNA virus). This product uses a new generation of antibody-modified Taq DNA Polymerase and one-step dedicated Reverse Transcriptase, with an optimized buffer for rapid amplification, which has faster amplification speed, higher amplification efficiency and specificity. It supports balanced amplification in both single-plex and multiplex of low and high concentration samples in a short time.

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Components

1. 5×RT-qPCR buffer (U+)

2. Enzyme mix (U+)

Notes:

a. 5×RT-qPCR buffer (U+)includes dNTP and Mg²⁺;

b. Enzyme mix (U+) includes reverse transcriptase, Hot Start Taq DNA polymerase, RNase inhibitor and UDG;

c. Use RNase-Free tips, EP tubes, etc.

Before use, thoroughly mix the 5×RT-qPCR buffer (U+). If there is any precipitation after thawing, wait for the buffer to return to room temperature, mix and dissolve, and then use them normally.

Storage Conditions

The product is shipped with dry ice and can be stored at -25~-15℃ for 1 year.

Instructions

1. Reaction System

Components	Volume (20 μL reaction)
2 ×RT-qPCR buffer	4μL
Enzyme mix (U+)	0.8μL
Primer Forward	0.1~ 1.0μM
Primer Reverse	0.1~ 1.0μM
TaqMan Probe	0.05~0.25μM
Template	X μL
RNase-Free Water	up to 25μL

Notes: Reaction volume is 10-50μL.

2. Cycling Protocol (Standard)

Cycle step	Temp.	Time	Cycles
Reverse Transcription	55 ℃	10 min	1
Initial Denaturation	95 ℃	30 sec	1
Denaturation	95 ℃	10 sec	45
Annealing/Extension	60 ℃	30 sec	45

Cycling Protocol (Fast) Cycle step	Temp.	Time	Cycles
Reverse Transcription	55 ℃	5 min	1
Initial Denaturation	95 ℃	5 s	1
Denaturation	95 ℃	3 s	43
Annealing/Extension	60 ℃	10 s	43

Notes:

a. Reverse transcription temperature is between 50℃ to 60℃, increasing the temperature helps to amplify complex structures and high CG content templates;

b. The optimal annealing temperature needs to be adjusted based on the Tm value of the primer, and select the shortest time for fluorescence signal collection based on the Real Time PCR instrument.

Notes

Please wear the necessary PPE, such lab coat and gloves, to ensure your health and safety!