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RevertAid Reverse Transcriptase


A genetically modified M-MuLV RT for routine performance in reverse transcription.
  
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Thermo Scientific RevertAid Reverse Transcriptase (RT) is a recombinant M-MuLV RT. It differs from the M-MuLV RT by its structure and catalytic properties. The enzyme possesses an RNA-dependent and DNA-dependent polymerase activity and a RNase H activity specific to RNA in RNA-DNA hybrids which is significantly lower than that of Avian Myeloblastosis Virus (AMV) reverse transcriptase (see References 1, 2).

Highlights

  • Efficient synthesis of full-length first strand cDNA up to 13 kb
  • Optimum activity at 42°C
  • Active up to 50°C
  • Incorporates modified nucleotides (e.g., Cy3-, Cy5-, rhodamine-, aminoallyl-, fluorescein-labeled nucleotides)

Applications

  • First strand cDNA synthesis for RT-PCR and real-time RT-PCR
  • Synthesis of cDNA for cloning and expression
  • Generation of labeled cDNA probes for microarrays
  • DNA labeling (see Reference 3)
  • Analysis of RNA by primer extension (see​ Reference 3)

Includes

  • RevertAid Reverse Transcriptase (200 U/µL)
  • 5X Reaction Buffer

Note

Also available: RevertAid First Strand cDNA Synthesis Kit

  
Storage Condition-20 C
HazardousNo
Molecular Weight76.1 kDa monomer
5x Reaction Buffer250 mM Tris-HCl (pH 8.3 at 25°C), 250 mM KCl, 20 mM MgCl2, 50 mM DTT
Definition of Activity Unit
  • One unit of the enzyme incorporates 1 nmol of dTMP into a polynucleotide fraction (adsorbed on DE-81) in 10 min at 37°C.
  • Enzyme activity is assayed in the following mixture: 50 mM Tris-HCl (pH 8.3), 4 mM MgCl2, 10 mM DTT, 50 mM KCl, 0.5 mM dTTP, 0.4 MBq/mL [3H]-dTTP, 0.4 mM polyA·oligo(dT)12-18.
InactivationInactivated by heating at 70°C for 10 min.
InhibitionMetal chelators, inorganic phosphate, pyrophosphate and polyamines (see Reference 2)
Quality Control
  • The absence of endo-, exodeoxyribonucleases, phosphatases, and ribonucleases confirmed by appropriate quality tests.
  • Functionally tested in first strand cDNA synthesis and RT-PCR.
RevertAid Reverse Transcriptase(200 U/µL) is supplied in 50 mM Tris-HCl (pH 7.5), 0.1 M NaCl, 1 mM EDTA, 5 mM DTT, 0.1% (v/v) Triton X-100, and 50% (v/v) glycerol
SourceE.coli cells with a cloned fragment of the pol gene encoding Moloney Murine Leukemia Virus reverse transcriptase.

References

  1. I. M. Verma, Reverse transcriptase, The Enzymes (P.D. Boyer, ed, Academic Press Inc.,vol. 14, 1981) pp. 87-103.

  2. G. F. Gerard, J. M. D’Alessio, Methods in Molecular Biology (Humana Press, N.J. Totowa vol. 16, 1993) pp. 73-93.

  3. J. Sambrook, D. W. Russell, Molecular Cloning: A Laboratory Manual, the third edition (Cold Spring Harbor Laboratory Press, Cold Spring Harbor, New York, 2001).