Rnase T1 digests

Matt Thomas Thomas at molbio.uoregon.edu
Wed Apr 23 05:15:07 EST 1997

I need to cut an in vitro prepared transcript with RNase T1.  The goal
with this was to cut to completion and anaylize the products on a high
percentage gel.  The problem I'm having is getting the dang enzyme to
cut completely and doing it in a buffer that doesn't make the high
percentage gels all ugly. The crazy idea is too look for G's which
aren't supposed to be there.  But I doubt to many of you really want to
hear the sob story of why I'm doing this.  

My first attempts at this, I used a buffer recomened by people in my lab
who use T1 for protection assays.  This was less the wonderfull.  Does
anyone out there have a protocol that has the activity of T1 optimized
for complete digestions?  Also, I'm useing BRL Rnase T1, is this the
best one out there or can someone recomend a differenct supplyer.  It
seems to me that this enzyme is fairly dirty (at least my + enzyme
pellets are really ugly compared to -enzyme pellets) and that may be
giving me trouble on my gels also.  

thanks for any advise or help.

Matthew Thomas,
Institute of Molecular Biology, University of Oregon
Thomas at molbio.uoregon.edu

Science is a cross dressing dominatrix.
--a fellow graduate student
  Version: 3.1
GS $ d-(--) S: a- C+>++ U@ P? L E? W+ N+ o? k? w--- O M++ !V(--) PS+ PE
PGP? t(++)@ 5+++ X+(++) R tv+() b+++ DI++ D+ G e++$>++++ h- r++ Y+

More information about the Methods mailing list