Home  Contacts  Order  Catalog  Support  Search  Alphabetical Index  Numerical Index  Restriction Enzymes  Modifying Enzymes  PCR, qPCR, RT-PCR & dNTPs  Molecular Cloning  Nucleic Acid Purification  Molecular Labeling & Detection  In vitro Transcription  Electrophoresis Products  Nucleotides  Transfection Reagents  Reagents C A T A L O G

RevertAid™ M-MuLV Reverse Transcriptase
Not available in the USA

#EP0441 Supplied with: 5X Reaction Buffer	10,000 u (200 u/µl) 1 ml
#EP0442 Supplied with: 5X Reaction Buffer	5x10,000 u (200 u/µl) 5x1 ml
Related Documents (in pdf, ~40-60 KB): Certificate of Analysis: #EP0441, #EP0442 MSDS (English) MSDS (English-USA) MSDS (German)

 

Features

• Suitable for synthesis of full-length cDNA as long as 13 kb.
• Temperature optimum of activity at 42°C.
• Activity up to 50°C.
• Incorporates modified nucleotides (e.g. Cy3-, Cy5-, rhodamine-, aminoallyl-, fluorescein-labeled nucleotides).

Description
The RevertAid™ M-MuLV Reverse Transcriptase (RT) is a genetically modified 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 ribonuclease H activity specific to RNA in RNA-DNA hybrids (1, 2).

Source
E.coli cells with a cloned fragment of the pol gene encoding Moloney Murine Leukemia Virus reverse transcriptase.

Molecular Weight
76.1 kDa monomer.

Applications

• Generation of first strand cDNA (3) for use in:
  • PCR, see Protocol for First-strand cDNA Synthesis;
  • real-time PCR;
  • second strand cDNA synthesis.
• DNA labeling (3).
• Analysis of RNA by primer extension (3).

Quality Control
The absence of endodeoxyribonucleases, exodeoxyribonucleases, phosphatases and ribonucleases confirmed by appropriate quality tests. Functionally tested in first strand cDNA synthesis.

Concentration
200 u/µl

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), 6 mM MgCl₂, 10 mM DTT, 40 mM KCl, 0.5 mM dTTP, 0.4 MBq/ml [³H]-dTTP, 0.4 mM polyA•oligo(dT)_12-18.

Storage Buffer
The enzyme is supplied in: 50 mM Tris-HCl (pH 8.3), 0.1 M NaCl, 1 mM EDTA, 5 mM DTT, 0.1% (v/v) Triton X-100 and 50% (v/v) glycerol.

5X Reaction Buffer
250 mM Tris-HCl (pH 8.3 at 25°C), 250 mM KCl, 20 mM MgCl₂, 50 mM DTT.

Inhibition and Inactivation

• Inhibitors: metal chelators, inorganic phosphate, pyrophosphate and polyamines (2).
• Inactivated by heating at 70°C for 10 min.

Note
RevertAid™ M-MuLV Reverse Transcriptase has significantly lower RNase H activity than Avian Myeloblastosis Virus (AMV) reverse transcriptase.

Related Products

dNTP Mix, 10 mM each RiboLock™ RNase Inhibitor RevertAid™ First Strand cDNA Synthesis Kit Primers Modified Nucleotides (molecular biology grade): Aminoallyl-dUTP  Biotin-11-dUTP Fluorescein-12-dUTP dm6ATP dm4CTP dm5CTP Taq DNA Polymerase (recombinant) Taq DNA Polymerase (native): without BSA with BSA 2X PCR Master Mix High Fidelity PCR Enzyme Mix Long PCR Enzyme Mix DNase I, RNase-free DNA Polymerase I, E.coli RNase H, E.coli T4 DNA Ligase Pyrophosphatase, Inorganic (from yeast) DNA ladders RiboRuler™ RNA ladders DEPC-treated Water

References

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

2. Gerard, G.F., D’Alessio, J.M., Methods in Molecular Biology, 16, Humana Press, Totowa, N.J., 73-93, 1993.
3. Sambrook, J., Russell, D.W., Molecular Cloning: A Laboratory Manual, the third edition, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, New York, 2001.

Updated kovo 18, 2008 09:56