REDscript(TM) M-MLV Reverse Transcriptase (H-)

REDscript(TM) M-MLV Reverse Transcriptase (H-)

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REDScript(TM) M-MLV Reverse Transcriptase, encoded by Moloney Murine Leukemia Virus (MMLV RT) is an RNA-dependent DNA polymerase that synthesizes the complementary DNA (cDNA) first strand from a single-stranded RNA template to which a primer has been hybridized. M-MLV RT will also extend primers hybridized to single-stranded DNA. Second strand cDNA synthesis can be achieved from some mRNA templates without an additional DNA polymerase. The difference between this to the general M-MLV RT is that the capacity to endure the heat is enhanced. It can remain the 100% activity at 50℃, it can also keep more than 75% activity at even at 55℃.

Thermal Stable M-MLV Reverse RH-Thermal Stable M-MLV

Upper panel: Template: Mouse total RNA, Reverse transcription at 50℃

Lane 1-2: REDscript(TM) M-MLV 200U

Lane 3-4 :invitrogen SuperScript® III

Lane 5-6:Takara m-mlv

Lane M: Marker

Lower panel: Template: Mouse total RNA, Reverse transcription at 55℃

Lane 1-2:Takara m-mlv

Lane 3-4: REDscript(TM) M-MLV

Lane 5-6:200U invitrogen SuperScript® III

Lane M: Marker

Source: Recombination of E.coli containing Moloney murine leukemia virus reverse transcriptase gene from clone of Moloney murine.
Concentration: 200U/μl
Component: M-MLV (200U/μl)
5×Buffer (with DTT)
Package: Bulk
Features: Weak RNaseH activity High cDNA yield
Storeage: -20℃

Unit Definition: One unit of REDscript(TM) M-MLV RT catalyzes
the incorporation of 1 nmol of dTTP into acidinsoluble material in 10 minutes at 37℃ using oligo(dT)12-18-primed poly(A)n as a template.

Applications: The first-strand cDNA synthesis; RT-PCR.

Storage Buffer: 20 mM Tris-HCl (pH7.5), 200 mM NaCl,0.25 mM EDTA, 0.01% NP-40(v/v),2.5 mM DTT,50% glycerol (v/v).
5X Reaction Buffer:
[5×RT Buffer] 250mM Tris-HCl (pH 8.3), 15mM MgCl2,375 mM KCl,50mM DTT.
Recommended Reaction Conditions:
The first-strand cDNA synthesis
1) Add the following reagents to a RNase free PCR tube at room temperature add the MMLV RT last.
Oligo dT12-18 (1μg/μl) or random primer (50-250ng) 1μl
Total RNA (10ng-5μg) or mRNA(1-500ng) xμl
dNTP (10mM each) 1μl
DEPC ddH2O (14-x)μl
2) Gently mix and incubate 10 Min at 70℃ then chill on ice for 2-10min.
3) Centrifuge for a few seconds then Put the tube into ice and add the next composition :
5×RT Buffer 4μl
RNasin (40U/μl) 1μl
5) Gently mix and incubate at 50℃ for 2 Min(Oligo dT12-18 or sequence especially primer)
or at 25℃ for 10 min for the random primer.
6) Centrifuge for a few seconds. Add 1μl REDscript(TM) M-MLV RT(200U/μl)Incubate at 50℃ for 50min.
7) Inactivate at 70℃ for 10min then get the cDNA.

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