AMV Reverse Transcriptase 反转录酶 Promega M5101 300u
Description （反转录酶英文描述） Avian Myeloblastosis Virus Reverse Transcriptase （AMV RT） catalyzes the
polymerization of DNA using template DNA, RNA or RNA:DNA hybrids （1）. It requires a primer （DNA primers are more efficient than RNA primers） as well as Mg2+ or Mn2+. The enzyme possesses an intrinsic RNase H activity. Both nonionic detergents and sulfhydryl compounds stabilize the enzyme activity in vitro.
Features
Available at High Concentration:Cat.# M9004 contains 600 units of AMV RT at 20–25u/μl.
Provided with 5X Reaction Buffer:250mM Tris-HCl （pH 8.3 at25°C）, 250mM KCl, 50mM MgCl2,
2.5mMspermidine, 50mM DTT.
Temperature Stability:AMV RT is the preferred reverse transcriptase for templates with high secondary structure due to its stability at higher reaction temperatures （37–58°C）
Applications First- and second-strand synthesis of cDNA.
Primer extensions and RNA sequencing （2）.
RT-PCR. Up to 10μl of an RT reaction containing AMV RT and the supplied AMV
RT Reaction Buffer can be added to a 50μl PCR amplification reaction that uses TaqDNA polymerase. If GoTaq® DNA Polymerase or PCR Master Mix are used, up to 25μl of an RT reaction can be added per 50μl PCR.
Protocol
Storage Conditions Store at –20°C.
Storage Buffer 200mM potassium phosphate （pH 7.2 at 4°C）, 0.2% Triton® X-100, 2mM DTT and
50% gycerol.
Unit Definition One unit is defined as the amount of enzyme required to catalyze the transfer of
1nmol of dTTP into acid-insoluble form in 10 minutes at 37°C. The reaction conditions are: 50mM Tris-HCl （pH 8.3 at 25°C）, 40mM KCl, 8.75mM MgCl2, 10mM DTT, 0.1mg/ml acetylated BSA, 1mM radiolabeled dTTP and 0.25mM poly（A）:oligo（dT）.
Quality Control Tests Activity, SDS-PAGE/purity, DNase, RNase, endonuclease, RT-PCR,
first-strand cDNA synthesis.
Source Purified Avian Myeloblastosis Virus particles.
References
1、Kacian, D.L. （1977） Meth. Virol. 6, 143.
2、Mierendorf, R.C. and Pfeffer, D. （1987） Meth. Enzymol. 152, 563–6. |
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