tet-Z: request for any experiences or tricks for using.

David Steffen steffen at mbir.bcm.tmc.edu
Mon Feb 10 11:57:56 EST 1992


  We have attempted to use tet-Z from Amersham as a reverse
transcriptase, and not had much luck.  I am posting to ask if anyone
has used this polymerase as a reverse transcriptase, and if so, how it
compared to MoMLV reverse transcriptase.

  We are amplifying a low abundance RNA with reverse transcriptase
prior to amplifying it by PCR.  We are using a sequence specific
primers for the reverse transcription.  This works well with MoMLV
reverse transcriptase, except that we are concerned that we might not
be getting the whole RNA copied due to RNA secondary structure.  As a
way around this, we attempted to use tet-Z, which works at high
temperature, where secondary structure should be reduced, as an
alternative.  Using tet-Z (under conditions specified by the
manufacturer for use as a reverse transcriptase) in place of MoMLV
reverse transcriptase gives no amplified PCR product.  Out of fairness
to Amersham, we have not extensively tested tet-Z; e.g. on other
templates, nor do w intend to; we cannot spare the time.  Before
giving up on this approach completely, however, I thought I would poll
the net for experiences with tet-Z, good or bad, along with any tips.
(We have already talked to Amersham, who could suggest nothing we
hadn't tried).


David Steffen
Department of Cell Biology, Baylor College of Medicine, Houston TX 77030
Telephone = (713) 798-6655, FAX = (713) 790-0545
Internet = steffen at mbir.bcm.tmc.edu

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