Description

ReverTra Ace-α-® is an efficient and convenient kit to synthesize high quality cDNA. This kit contains the highly efficient reverse transcriptase "ReverTra Ace®", as well as other components optimized for the synthesis of long cDNAs suitable for RT-PCR. ReverTra Ace® is an M-MLV reverse transcriptase that has been improved by point mutation technology. ReverTra Ace® has two mutations in the polymerase and RNase H domains.

Features

  • Contains all components for reverse transcription.
  • Enables the synthesis of ≥ 14 kb cDNA.

Applications

  • cDNA synthesis
  • RT-PCR

Storage condition

Store at -20ºC

Components

ReverTra Ace® 100 µl
5xRT buffer (contains 25 mM Mg2+) 400 µl
Rnase inhibitor (10 U/µl) 100 µl
dNTPs mixture (10 mM) 200 µl
RNase-free H2O 1200 µl
Oligo (dT)20 (10 pmol/µl) 100 µl
Random primers (25 pmol/µl) 100 µl
Control Primer F (10 pmol/µl) 50 µl
Control Primer R (10 pmol/µl) 50 µl
Positive control RNA (105 copies/µl) 50 µl

Application data

Example 1.Detection of human β-actin mRNA by RT-PCR utilizing ReverTra Ace-α-® and various PCR enzymes

cDNA was synthesized using 1 mg total RNA from the human cell line (HeLa cell) at 42ºC for 20 minutes, followed by inactivation with ReverTra Ace-α-® in a volume of 20 ml at 99ºC for 5 minutes. Subsequently, using the attained cDNA, the target gene (β-actin: 838 bp) was amplified with various PCR enzymes.As shown in Fig. 1, the target genes were successfully amplified using the various PCR enzymes tested.

Fig. 1 Amplification of the 838-bp β-actin gene

M: 100 bp Ladder Marker
1: rTth DNA polymerase
2: rTaq DNA polymerase
3: High efficient Taq DNA polymerase (Company A)
4: High efficient Taq DNA polymerase (Company B)
5: KOD Dash [Code No. LDP-101]

References

  • W.M. Barns, PCR amplification of up to 35-kb DNA with high fidelity and high yield from lambda bacteriophage templates.Proc. Natl. Acad. Sci. USA, 91: 2216-2220 (1994)