Description

ReverTra Ace® qPCR RT Kit is an efficient and convenient kit to synthesize high quality cDNAs for real-time PCR. This kit contains the highly efficient reverse transcriptase "ReverTra Ace®" and a RT buffer optimized for the highly efficient synthesis of short-chain cDNAs suitable for real-time PCR. The protocol is simple, and the reaction can be completed in 15 min.
ReverTra Ace® is a mutant-type M-MLV reverse transcriptase shows excellent efficiency.
ReverTra Ace® qPCR RT Master Mix (Code No. FSQ-201) is a premix version of ReverTra Ace® qPCR RT Kit. The master mix reagent (5x) contains the highly efficient reverse transcriptase "ReverTra Ace®" , primers and buffer optimized for highly efficient synthesis of short-chain cDNAs suitable for real-time PCR.

Features

  • The optimized RT buffer and Primer Mix enable the highly efficient reverse transcription.
  • The reaction can be completed in 15 min. The protocol does not contain an additional RNase H treatment step to remove residual RNA after reverse transcription (Patent Pending).
  • Suitable for the detection of low-expressing mRNAs. Since the RT buffer is optimized for real-time PCR, the addition of 20% (v/v) of the synthesized cDNA solution to the PCR solution does not inhibit the PCR reaction.

Applications

Reverse transcriptation for real-time PCR

Storage condition

Store at -20ºC

Components

ReverTra Ace® qPCR RT Kit

5 x RT Buffer 400 µl
Enzyme Mix 100 µl
Primer Mix 100 µl
Nuclease-free water 1000 µl × 2

ReverTra Ace® qPCR RT Master Mix

5 x RT Master Mix 400 µl
5 x RT Master Mix no RT-Control 40 µl
Nuclease-free water 1000 µl × 2

Application data

Example 1.Comparison of cDNA yields using various reverse transcription kits

cDNA was synthesized from human cell line (HeLa cell) total RNA with the ReverTra Ace® qPCR RT kit. Subsequently, quantification of β-actin and polymerase ε mRNA were performed by real-time PCR with 100 ng cDNA, in conjunction with the SYBR® Green Realtime PCR Master Mix [Code No. QPK-201].
As shown in Fig. 1, the relative cDNA yields from the ReverTra Ace® qPCR RT kit were greater than from the other commercial kits.

Fig. 1 Comparison of relative cDNA concentrations

The reaction temperature conditions of the reverse transcription are indicated under the figure.

Example 2.Detection of a rare expressing gene

Several 1-µl cDNA aliquots were prepared from 100 ng human cell line total RNA in a volume of 20 µl using various reverse transcription kits. Subsequently, a low expressing gene (TNF-α) was detected with real-time PCR kit (SYBR® Green Realtime PCR Master Mix [Code No. QPK-201]. The Ct values and success rates of the ReverTra Ace qPCR RT kit were greater than those from the other commercial kits (Fig. 2).

Fig. 2 Comparison of Ct values and success rates when detecting rare expressing genes