AMV vs. MMLV reverse transcriptase
Marc Gregory Busch
hexamer primers. The old protocol uses MMLV. However AMV reverse
transcriptase is cheaper. Is there a difference and what's better for
certain conditions. After looking the most I could find is that AMV is
stable at a slightly higher temperature. Thank you an advance for any
assistance you may have in the manner.
Marc
Gregory Busch
theo...@medlib.georgetown.edu
I'm probably showing my age, but when I was an undergrad., people used to get
AMV RT for free.
As far as MMLV vs. AMV, many people (including myself) use a certain MMLV RT
that has been monkeyed around with so that it has no RNAse H activity. AFAIK
(admittedly, not very far) I don't think the equivalent product based on AMV
RT is sold (but please feel free to correct me if I'm wrong). So if the
choice is between the modified murine vs. avian, you might get better cDNA
with the modified murine enzyme. If using wild-type (RNAse H positive), I
don't have a good recommmendation.
Nick Theodorakis
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Neil Saunders
Marc Gregory Busch wrote:
> I am currently trying to made a cDNA library of total RNA using random
> hexamer primers. The old protocol uses MMLV. However AMV reverse
> transcriptase is cheaper. Is there a difference and what's better for
> certain conditions. After looking the most I could find is that AMV is
> stable at a slightly higher temperature. Thank you an advance for any
> assistance you may have in the manner.
> Marc
> Gregory Busch
>
> mgb...@ucdavis.edu
As I remember, AMV-RT is more processive than MMLV-RT, meaning that less units
are required in the reaction; I think typically 1 microlitre of MMLV-RT in a
reaction is about 200U, whereas 1 microlitre of AMV-RT is a few tens of units
(depending on the concentration from the supplier, obviously). AMV-RT is
supposed to be more stable at higher temperatures, meaning that you should be
able to raise the incubation temperature and so lessen the truncation of
cDNA's due to pausing at secondary structure, although MMLV-RT is also usable
to at least 42C. I've used them both, for 5'-RACE, primer extension and
RT-PCR and have found one about as good as the other, to be honest.
Neil Saunders
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