AMV Reverse Transcriptase   反轉(zhuǎn)錄酶 Promega M5101  300u

Description （反轉(zhuǎn)錄酶英文描述）  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 Taq DNA 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.